Sample records for mec gene complex
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Katayama, Yuki; Ito, Teruyo; Hiramatsu, Keiichi
2001-01-01
We report on the structural diversity of mecA gene complexes carried by 38 methicillin-resistant Staphylococcus aureus and 91 methicillin-resistant coagulase-negative Staphylococcus strains of seven different species with a special reference to its correlation with phenotypic expression of methicillin resistance. The most prevalent and widely disseminated mec complex had the structure mecI-mecR1-mecA-IS431R (or IS431mec), designated the class A mecA gene complex. In contrast, in S. haemolyticus, mecA was bracketed by two copies of IS431, forming the structure IS431L-mecA-IS431R. Of the 38 S. haemolyticus strains, 5 had low-level methicillin resistance (MIC, 1 to 4 mg/liter) and characteristic heterogeneous methicillin resistance as judged by population analysis. In these five strains, IS431L was located to the left of an intact mecI gene, forming the structure IS431L-class A mecA-gene complex. In other S. haemolyticus strains, IS431L was associated with the deletion of mecI and mecR1, forming the structure IS431L-ΔmecR1-mecA-IS431mec, designated the class C mecA gene complex. Mutants with the class C mecA gene complex were obtained in vitro by selecting strain SH621, containing the IS431L-class A mecA gene complex with low concentrations of methicillin (1 and 3 mg/liter). The mutants had intermediate level of methicillin resistance (MIC, 16 to 64 mg/liter). The mecA gene transcription was shown to be derepressed in a representative mutant strain, SH621-37. Our study indicated that the mecI-encoded repressor function is responsible for the low-level methicillin resistance of some S. haemolyticus clinical strains and that the IS431-mediated mecI gene deletion causes the expression of methicillin resistance through the derepression of mecA gene transcription. PMID:11408208
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Szczuka, Ewa; Krajewska, Magdalena; Lijewska, Dagmara; Bosacka, Karolina; Kaznowski, Adam
2016-11-01
Staphylococcus haemolyticus is the second, most frequently isolated coagulase-negative staphyloccus (CoNS) from patients with hospital-acquired infections, and it is usually resistant to methicillin and other semisynthetic penicillins. The purpose of this study was to characterize staphylococcal cassette chromosome mec (SCCmec) elements and assess the in-vitro activity of antibiotics against 60 S. haemolyticus strains recovered from hospitalized patients. All these strains expressed methicillin resistance and carried a mecA gene. Moreover, all strains possessed a multiresistant phenotype, i.e., exhibited resistance to more than three classes of antibiotics. Eleven strains (18 %) harbored the SCCmec type V, containing ccrC and mec complex C. Three isolates harboring the ccrC gene did not contain a known mec complex. One strain positive for mec complex C was not typeable for ccr. This suggests that ccrC and mec complex C may exist autonomously. Only four strains carried mec complex B, whereas none of the S. haemolyticus harboured mec complex A. A new combination, which is mec complex B-ccrAB ship , was found in S. haemolitycus. The ccrAB ship was also identified in two strains of S. haemolitycus in which the mec gene complex was not identified. The results of the present study indicate that in S. haemolyticus the mec gene complex and the ccr genes are highly divergent. However, ccr sequence analysis does not allow the identification of a new allotype, based on a cut-off value of 85 % identity. The ccr genes in the S. haemolitycus strain showed ≥96 % sequence identity to the ccrAB2 genes.
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Ito, Teruyo; Ma, Xiao Xue; Takeuchi, Fumihiko; Okuma, Keiko; Yuzawa, Harumi; Hiramatsu, Keiichi
2004-01-01
Staphylococcal cassette chromosome mec (SCCmec) is a mobile genetic element composed of the mec gene complex, which encodes methicillin resistance, and the ccr gene complex, which encodes the recombinases responsible for its mobility. The mec gene complex has been classified into four classes, and the ccr gene complex has been classified into three allotypes. Different combinations of mec gene complex classes and ccr gene complex types have so far defined four types of SCCmec elements. Now we introduce the fifth allotype of SCCmec, which was found on the chromosome of a community-acquired methicillin-resistant Staphylococcus aureus strain (strain WIS [WBG8318]) isolated in Australia. The element shared the same chromosomal integration site with the four extant types of SCCmec and the characteristic nucleotide sequences at the chromosome-SCCmec junction regions. The novel SCCmec carried mecA bracketed by IS431 (IS431-mecA-ΔmecR1-IS431), which is designated the class C2 mec gene complex; and instead of ccrA and ccrB genes, it carried a single copy of a gene homologue that encoded cassette chromosome recombinase. Since the open reading frame (ORF) was found to encode an enzyme which catalyzes the precise excision as well as site- and orientation-specific integration of the element, we designated the ORF cassette chromosome recombinase C (ccrC), and we designated the element type V SCCmec. Type V SCCmec is a small SCCmec element (28 kb) and does not carry any antibiotic resistance genes besides mecA. Unlike the extant SCCmec types, it carries a set of foreign genes encoding a restriction-modification system that might play a role in the stabilization of the element on the chromosome. PMID:15215121
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Urushibara, Noriko; Paul, Shyamal Kumar; Hossain, Mohammad Akram; Kawaguchiya, Mitsuyo; Kobayashi, Nobumichi
2011-06-01
Methicillin resistance in staphylococci is conferred by the acquisition in its chromosome of the mecA gene, which is located on a mobile genetic element called staphylococcal cassette chromosome mec (SCCmec). Genetic type of SCCmec is defined by combination of mec gene complex class and cassette chromosome recombinase gene (ccr) allotype. In this study, we analyzed genetic diversity of the SCCmec in 11 Staphylococcus haemolyticus strains and a Staphylococcus sciuri strain, which were recently isolated from clinical specimens in Bangladesh. Among these strains, only two S. haemolyticus strains were proved to have the known types of SCCmec, that is, SCCmec V (class C2 mec-ccrC) and VII (class C1 mec-ccrC). Five S. haemolyticus strains were assigned two unique mec-ccr gene complexes combination; that is, class C1 mec-ccrA4B4 (four isolates) and class A mec-ccrC (one isolate). In the remaining four S. haemolyticus strains with class C1 mec, no known ccr allotypes could be detected. A single S. sciuri strain with class A mec complex carried a ccrA gene belonging to a novel allotype designated ccrA7, together with ccrB3. The ccrA7 gene in the S. sciuri strain showed 61.7%-82.7% sequence identity to the ccrA gene sequences published so far, and 75.3% identity to ccrA3, which is a component of the type 3 ccr complex (ccrA3-ccrB3) in methicillin-resistant Staphylococcus aureus. The results of the present study indicated that mec gene complex and ccr genes in coagulase-negative staphylococci are highly divergent, and distinct from those of common methicillin-resistant S. aureus. Identification of the novel ccrA7 allotype combined with ccrB3 suggested an occurrence of recombination between different ccr complexes in nature.
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Gómez-Sanz, Elena; Schwendener, Sybille; Thomann, Andreas; Gobeli Brawand, Stefanie
2015-01-01
A methicillin-resistant mecB-positive Macrococcus caseolyticus (strain KM45013) was isolated from the nares of a dog with rhinitis. It contained a novel 39-kb transposon-defective complete mecB-carrying staphylococcal cassette chromosome mec element (SCCmecKM45013). SCCmecKM45013 contained 49 coding sequences (CDSs), was integrated at the 3′ end of the chromosomal orfX gene, and was delimited at both ends by imperfect direct repeats functioning as integration site sequences (ISSs). SCCmecKM45013 presented two discontinuous regions of homology (SCCmec coverage of 35%) to the chromosomal and transposon Tn6045-associated SCCmec-like element of M. caseolyticus JCSC7096: (i) the mec gene complex (98.8% identity) and (ii) the ccr-carrying segment (91.8% identity). The mec gene complex, located at the right junction of the cassette, also carried the β-lactamase gene blaZm (mecRm-mecIm-mecB-blaZm). SCCmecKM45013 contained two cassette chromosome recombinase genes, ccrAm2 and ccrBm2, which shared 94.3% and 96.6% DNA identity with those of the SCCmec-like element of JCSC7096 but shared less than 52% DNA identity with the staphylococcal ccrAB and ccrC genes. Three distinct extrachromosomal circularized elements (the entire SCCmecKM45013, ΨSCCmecKM45013 lacking the ccr genes, and SCCKM45013 lacking mecB) flanked by one ISS copy, as well as the chromosomal regions remaining after excision, were detected. An unconventional circularized structure carrying the mecB gene complex was associated with two extensive direct repeat regions, which enclosed two open reading frames (ORFs) (ORF46 and ORF51) flanking the chromosomal mecB-carrying gene complex. This study revealed M. caseolyticus as a potential disease-associated bacterium in dogs and also unveiled an SCCmec element carrying mecB not associated with Tn6045 in the genus Macrococcus. PMID:25987634
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Safo, Martin K., E-mail: msafo@vcu.edu; Ko, Tzu-Ping; Musayev, Faik N.
The up-and-down binding of dimeric MecI to mecA dyad DNA may account for the cooperative effect of the repressor. The dimeric repressor MecI regulates the mecA gene that encodes the penicillin-binding protein PBP-2a in methicillin-resistant Staphylococcus aureus (MRSA). MecI is similar to BlaI, the repressor for the blaZ gene of β-lactamase. MecI and BlaI can bind to both operator DNA sequences. The crystal structure of MecI in complex with the 32 base-pair cognate DNA of mec was determined to 3.8 Å resolution. MecI is a homodimer and each monomer consists of a compact N-terminal winged-helix domain, which binds to DNA,more » and a loosely packed C-terminal helical domain, which intertwines with its counter-monomer. The crystal contains horizontal layers of virtual DNA double helices extending in three directions, which are separated by perpendicular DNA segments. Each DNA segment is bound to two MecI dimers. Similar to the BlaI–mec complex, but unlike the MecI–bla complex, the MecI repressors bind to both sides of the mec DNA dyad that contains four conserved sequences of TACA/TGTA. The results confirm the up-and-down binding to the mec operator, which may account for cooperative effect of the repressor.« less
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Safo,M.; Ko, T.; Musayev, F.
The dimeric repressor MecI regulates the mecA gene that encodes the penicillin-binding protein PBP-2a in methicillin-resistant Staphylococcus aureus (MRSA). MecI is similar to BlaI, the repressor for the blaZ gene of {beta}-lactamase. MecI and BlaI can bind to both operator DNA sequences. The crystal structure of MecI in complex with the 32 base-pair cognate DNA of mec was determined to 3.8 Angstroms resolution. MecI is a homodimer and each monomer consists of a compact N-terminal winged-helix domain, which binds to DNA, and a loosely packed C-terminal helical domain, which intertwines with its counter-monomer. The crystal contains horizontal layers of virtualmore » DNA double helices extending in three directions, which are separated by perpendicular DNA segments. Each DNA segment is bound to two MecI dimers. Similar to the BlaI-mec complex, but unlike the MecI-bla complex, the MecI repressors bind to both sides of the mec DNA dyad that contains four conserved sequences of TACA/TGTA. The results confirm the up-and-down binding to the mec operator, which may account for cooperative effect of the repressor.« less
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A Staphylococcus xylosus Isolate with a New mecC Allotype
Harrison, Ewan M.; Paterson, Gavin K.; Holden, Matthew T. G.; Morgan, Fiona J. E.; Larsen, Anders Rhod; Petersen, Andreas; Leroy, Sabine; De Vliegher, Sarne; Perreten, Vincent; Fox, Lawrence K.; Lam, Theo J. G. M.; Sampimon, Otlis C.; Zadoks, Ruth N.; Peacock, Sharon J.; Parkhill, Julian
2013-01-01
Recently, a novel variant of mecA known as mecC (mecALGA251) was identified in Staphylococcus aureus isolates from both humans and animals. In this study, we identified a Staphylococcus xylosus isolate that harbors a new allotype of the mecC gene, mecC1. Whole-genome sequencing revealed that mecC1 forms part of a class E mec complex (mecI-mecR1-mecC1-blaZ) located at the orfX locus as part of a likely staphylococcal cassette chromosome mec element (SCCmec) remnant, which also contains a number of other genes present on the type XI SCCmec. PMID:23274660
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2013-01-01
Background Methicillin resistance determinant mecA is generally transferred by SCCmec elements. However, the mecA gene might not be carried by a SCCmec in a Staphylococcus haemolyticus clinical isolate, WCH1, as no cassette chromosome recombinase genes were detected. Therefore, the genetic context of mecA in WCH1 was investigated. Results A 40-kb region containing mecA was obtained from WCH1, bounded by orfX at one end and several orfs of S. haemolyticus core chromosome at the other. This 40-kb region was very complex in structure with multiple genetic components that appeared to have different origins. For instance, the 3.7-kb structure adjacent to orfX was almost identical to that on the chromosome of Staphylococcus epidermidis RP62a but was absent from S. haemolyticus JCSC1435. Terminal inverted repeats of SCC were found but no ccr genes could be detected. mecA was bracketed by two copies of IS431, which was flanked by 8-bp direct target repeat sequence (DR). Conclusions The presence of 8-bp DR suggests that the two copies of IS431 might have formed a composite transposon for mobilizing mecA. This finding is of significance as multiple copies of IS431 are commonly present in the contexts of mecA, which might have the potential to form various composite transposons that could mediate the mobilization of mecA. This study also provides an explanation for the absence of ccr in some staphylococci isolates carrying mecA. PMID:23521926
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Zong, Zhiyong
2013-03-22
Methicillin resistance determinant mecA is generally transferred by SCCmec elements. However, the mecA gene might not be carried by a SCCmec in a Staphylococcus haemolyticus clinical isolate, WCH1, as no cassette chromosome recombinase genes were detected. Therefore, the genetic context of mecA in WCH1 was investigated. A 40-kb region containing mecA was obtained from WCH1, bounded by orfX at one end and several orfs of S. haemolyticus core chromosome at the other. This 40-kb region was very complex in structure with multiple genetic components that appeared to have different origins. For instance, the 3.7-kb structure adjacent to orfX was almost identical to that on the chromosome of Staphylococcus epidermidis RP62a but was absent from S. haemolyticus JCSC1435. Terminal inverted repeats of SCC were found but no ccr genes could be detected. mecA was bracketed by two copies of IS431, which was flanked by 8-bp direct target repeat sequence (DR). The presence of 8-bp DR suggests that the two copies of IS431 might have formed a composite transposon for mobilizing mecA. This finding is of significance as multiple copies of IS431 are commonly present in the contexts of mecA, which might have the potential to form various composite transposons that could mediate the mobilization of mecA. This study also provides an explanation for the absence of ccr in some staphylococci isolates carrying mecA.
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Chatterjee, Som S; Chen, Liang; Joo, Hwang-Soo; Cheung, Gordon Y C; Kreiswirth, Barry N; Otto, Michael
2011-01-01
The phenol-soluble modulin PSM-mec is the only known staphylococcal toxin that is encoded on a mobile antibiotic resistance determinant, namely the staphylococcal cassette chromosome (SCC) element mec encoding resistance to methicillin. Here we show that the psm-mec gene is found frequently among methicillin-resistant Staphylococcus aureus (MRSA) strains of SCCmec types II, III, and VIII, and is a conserved part of the class A mec gene complex. Controlled expression of AgrA versus RNAIII in agr mutants of all 3 psm-mec-positive SCCmec types demonstrated that expression of psm-mec, which is highly variable, is controlled by AgrA in an RNAIII-independent manner. Furthermore, psm-mec isogenic deletion mutants showed only minor changes in PSMα peptide production and unchanged (or, as previously described, diminished) virulence compared to the corresponding wild-type strains in a mouse model of skin infection. This indicates that the recently reported regulatory impact of the psm-mec locus on MRSA virulence, which is opposite to that of the PSM-mec peptide and likely mediated by a regulatory RNA, is minor when analyzed in the original strain background. Our study gives new insight in the distribution, regulation, and role in virulence of the PSM-mec peptide and the psm-mec gene locus.
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A novel hybrid SCCmec-mecC region in Staphylococcus sciuri
Harrison, Ewan M.; Paterson, Gavin K.; Holden, Matthew T. G.; Ba, Xiaoliang; Rolo, Joana; Morgan, Fiona J. E.; Pichon, Bruno; Kearns, Angela; Zadoks, Ruth N.; Peacock, Sharon J.; Parkhill, Julian; Holmes, Mark A.
2014-01-01
Objectives Methicillin resistance in Staphylococcus spp. results from the expression of an alternative penicillin-binding protein 2a (encoded by mecA) with a low affinity for β-lactam antibiotics. Recently, a novel variant of mecA known as mecC (formerly mecALGA251) was identified in Staphylococcus aureus isolates from both humans and animals. In this study, we identified two Staphylococcus sciuri subsp. carnaticus isolates from bovine infections that harbour three different mecA homologues: mecA, mecA1 and mecC. Methods We subjected the two isolates to whole-genome sequencing to further understand the genetic context of the mec-containing region. We also used PCR and RT–PCR to investigate the excision and expression of the SCCmec element and mec genes, respectively. Results Whole-genome sequencing revealed a novel hybrid SCCmec region at the orfX locus consisting of a class E mec complex (mecI-mecR1-mecC1-blaZ) located immediately downstream of a staphylococcal cassette chromosome mec (SCCmec) type VII element. A second SCCmec attL site (attL2), which was imperfect, was present downstream of the mecC region. PCR analysis of stationary-phase cultures showed that both the SCCmec type VII element and a hybrid SCCmec-mecC element were capable of excision from the genome and forming a circular intermediate. Transcriptional analysis showed that mecC and mecA, but not mecA1, were both expressed in liquid culture supplemented with oxacillin. Conclusions Overall, this study further highlights that a range of staphylococcal species harbour the mecC gene and furthers the view that coagulase-negative staphylococci associated with animals may act as reservoirs of antibiotic resistance genes for more pathogenic staphylococcal species. PMID:24302651
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Monecke, Stefan; Gavier-Widen, Dolores; Mattsson, Roland; Rangstrup-Christensen, Lena; Lazaris, Alexandros; Coleman, David C.; Shore, Anna C.; Ehricht, Ralf
2013-01-01
Recently, a novel mec gene conferring beta-lactam resistance in Staphylococcus aureus has been discovered. This gene, mecC, is situated on a SCCmec XI element that has to date been identified in clonal complexes 49, 130, 425, 599 and 1943. Some of the currently known isolates have been identified from animals. This, and observations of mecA alleles that do not confer beta-lactam resistance, indicate that mec genes might have a reservoir in Staphylococcus species from animals. Thus it is important also to screen wildlife isolates for mec genes. Here, we describe mecC-positive Staphylococcus aureus (ST130-MRSA-XI) and the lesions related to the infection in two diseased free-ranging European hedgehogs (Erinaceus europaeus). One was found dead in 2003 in central Sweden, and suffered from S. aureus septicaemia. The other one, found on the island of Gotland in the Baltic Sea in 2011, showed a severe dermatitis and was euthanised. ST130-MRSA-XI isolates were isolated from lesions from both hedgehogs and were essentially identical to previously described isolates from humans. Both isolates carried the complete SCCmec XI element. They lacked the lukF-PV/lukS-PV and lukM/lukF-P83 genes, but harboured a gene for an exfoliative toxin homologue previously described from Staphylococcus hyicus, Staphylococcus pseudintermedius and other S. aureus of the CC130 lineage. To the best of our knowledge, these are the first reported cases of CC130-MRSA-XI in hedgehogs. Given that one of the samples was taken as early as 2003, this was the earliest detection of this strain and of mecC in Sweden. This and several other recent observations suggest that CC130 might be a zoonotic lineage of S. aureus and that SCCmec XI/mecC may have originated from animal pathogens. PMID:23776626
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Monecke, Stefan; Gavier-Widen, Dolores; Mattsson, Roland; Rangstrup-Christensen, Lena; Lazaris, Alexandros; Coleman, David C; Shore, Anna C; Ehricht, Ralf
2013-01-01
Recently, a novel mec gene conferring beta-lactam resistance in Staphylococcus aureus has been discovered. This gene, mecC, is situated on a SCCmec XI element that has to date been identified in clonal complexes 49, 130, 425, 599 and 1943. Some of the currently known isolates have been identified from animals. This, and observations of mecA alleles that do not confer beta-lactam resistance, indicate that mec genes might have a reservoir in Staphylococcus species from animals. Thus it is important also to screen wildlife isolates for mec genes. Here, we describe mecC-positive Staphylococcus aureus (ST130-MRSA-XI) and the lesions related to the infection in two diseased free-ranging European hedgehogs (Erinaceus europaeus). One was found dead in 2003 in central Sweden, and suffered from S. aureus septicaemia. The other one, found on the island of Gotland in the Baltic Sea in 2011, showed a severe dermatitis and was euthanised. ST130-MRSA-XI isolates were isolated from lesions from both hedgehogs and were essentially identical to previously described isolates from humans. Both isolates carried the complete SCCmec XI element. They lacked the lukF-PV/lukS-PV and lukM/lukF-P83 genes, but harboured a gene for an exfoliative toxin homologue previously described from Staphylococcus hyicus, Staphylococcus pseudintermedius and other S. aureus of the CC130 lineage. To the best of our knowledge, these are the first reported cases of CC130-MRSA-XI in hedgehogs. Given that one of the samples was taken as early as 2003, this was the earliest detection of this strain and of mecC in Sweden. This and several other recent observations suggest that CC130 might be a zoonotic lineage of S. aureus and that SCCmec XI/mecC may have originated from animal pathogens.
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Basrai, M A; Velculescu, V E; Kinzler, K W; Hieter, P
1999-10-01
Analysis of global gene expression in Saccharomyces cerevisiae by the serial analysis of gene expression technique has permitted the identification of at least 302 previously unidentified transcripts from nonannotated open reading frames (NORFs). Transcription of one of these, NORF5/HUG1 (hydroxyurea and UV and gamma radiation induced), is induced by DNA damage, and this induction requires MEC1, a homolog of the ataxia telangiectasia mutated (ATM) gene. DNA damage-specific induction of HUG1, which is independent of the cell cycle stage, is due to the alleviation of repression by the Crt1p-Ssn6p-Tup1p complex. Overexpression of HUG1 is lethal in combination with a mec1 mutation in the presence of DNA damage or replication arrest, whereas a deletion of HUG1 rescues the lethality due to a mec1 null allele. HUG1 is the first example of a NORF with important biological functional properties and defines a novel component of the MEC1 checkpoint pathway.
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Bortolami, Alessio; Verin, Ranieri; Chantrey, Julian; Corrò, Michela; Ashpole, Ian; Lopez, Javier; Timofte, Dorina
2017-10-01
Little is known about the characteristics and diseases associated with methicillin-resistant Staphylococcus aureus (MRSA) in nondomestic animals. Four presumptive MRSA isolates, obtained from clinical (n = 3) and surveillance specimens (n = 1) from dwarf (Helogale parvula) and yellow mongooses (Cynictis penicillata) from a United Kingdom zoo, were analyzed by PCR for detection of mecA and mecC-mediated methicillin resistance, and virulence genes. Isolates were genotyped by multilocus sequence typing (MLST) and staphylococcal cassette chromosome mec (SCCmec) and spa sequence typing. Three isolates, obtained from the dwarf mongooses, carried mecA, tetK, and fexA resistance and virulence genes (icaA, icaD, and sec) and were typed to SCCmec IVa, spa type t899, and clonal complex (CC) 398. The fourth MRSA isolate, obtained from the femoral bone marrow of a yellow mongoose showing postmortem findings consistent with septicemia, carried mecC and was oxacillin/cefoxitin susceptible, when tested at 37°C but showed a characteristic MRSA susceptibility profile at 25°C ± 2°C. Furthermore, this isolate exhibited a different genetic background (SCCmecXI/t843/CC130) and had biofilm-associated genes (bap, icaA, and icaD) and tetK tetracycline resistance genes. This work describes the first isolation of livestock-associated MRSA CC398 from two zoo mongoose species where it was associated with both clinical disease and colonization, and the first isolation of mecC MRSA from a zoo species in the United Kingdom. Both reports highlight the potential for zoo species to act as reservoirs for these zoonotic agents.
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Hosseinkhani, Farideh; Tammes Buirs, Matthias; Jabalameli, Fereshteh; Emaneini, Mohammad; van Leeuwen, Willem B
2018-06-06
Staphylococcus haemolyticus has emerged as a highly antimicrobial-resistant healthcare-associated pathogen, in particular for patients admitted to neonatal intensive care. The objective of this study was to study the nature of SCCmec types among MDR-SH strains isolated from paediatric patients. S. haemolyticus strains (n=60) were isolated from paediatric patients. Antibiotic resistance patterns were established using the disk agar diffusion and micro-broth dilution methods. SCCmec typing was performed using whole-genome sequencing (WGS) and an additional PCR analysis. All S. haemolyticus isolates demonstrated multidrug resistance. Using WGS, various novel mec types and combinations of SCCmec types were found, including a new composite island [SCCmec type V (Vd)+SCC cad/ars/cop] comprising 30 % of the strains. SCCmec type V was identified in 23 % of the isolates. A combination of the mecA gene enclosed by two copies of IS431 and absence of the mecRI and ccr genes was identified in 11 strains. In total, mecA regulatory genes were absent in all SH isolates used in this study. A high diversity of SCCmec elements with the prevalence of a new composite island was determined among MRSH strains. The structure of the composite island represented by MDR-SH strains in this study, in combination with the presence of a restriction-modification system type III, is described for the first time in this study. The presence of an 8 bp direct repeat (DR) and the sequences flanking the DR may support the integration of the mecA gene complex as a composite transposon (IS431-mecA-IS431) independently from recombinase genes.
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Chanchaithong, Pattrarat; Prapasarakul, Nuvee; Schwendener, Sybille
2015-01-01
A novel staphylococcal cassette chromosome mec (SCCmec) composite island (SCCmecAI16-SCCczrAI16-CI) was identified in Staphylococcus pseudintermedius. Four integration site sequences for SCC subdivided the 60,734-bp island into 41,232-bp SCCmecAI16, 19,400-bp SCCczrAI16, and 102-bp SCC-likeAI16 elements. SCCmecAI16 represents a new combination of ccrA1B3 genes with a class A mec complex. SCCczrAI16 contains ccrA1B6 and genes related to restriction modification and heavy metal resistance. SCCmecAI16-SCCczrAI16-CI was found in methicillin-resistant S. pseudintermedius sequence type 112 (ST112) and ST111 isolated from dogs and veterinarians in Thailand. PMID:26643350
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Giacinti, G; Carfora, V; Caprioli, A; Sagrafoli, D; Marri, N; Giangolini, G; Amoruso, R; Iurescia, M; Stravino, F; Dottarelli, S; Feltrin, F; Franco, A; Amatiste, S; Battisti, A
2017-10-01
Between January and May 2012, a total of 286 bulk tank milk samples from dairy sheep farms located in central Italy were tested for the presence of Staphylococcus aureus. One hundred fifty-three samples were positive for S. aureus (53.5%), with an average count of 2.53 log cfu/mL. A total of 679 S. aureus colonies were screened for methicillin resistance by the cefoxitin disk diffusion test, and 104 selected cefoxitin-susceptible isolates were also tested for their susceptibility to other antimicrobials representative of the most relevant classes active against Staphylococcus spp. by using the Kirby-Bauer disk diffusion method. Two methicillin-resistant Staphylococcus aureus (MRSA) isolates, carrying respectively the mecA and the mecC genes, were detected in 2 samples from 2 different farms (prevalence 0.7%). The mecA-positive MRSA isolate was blaZ positive, belonged to spa type t127, sequence type (ST)1, clonal complex (CC)1, carried a staphylococcal cassette chromosome mec (SCCmec) type IVa, and was phenotypically resistant to all the β-lactams tested and to erythromycin, streptomycin, kanamycin, and tetracycline. The mecC-positive MRSA isolate was negative for the chromosomally or plasmid-associated blaZ gene but positive for the blaZ allotype associated with SCCmec XI (blaZ-SCCmecXI), belonged to spa type 843, ST(CC)130, carried a SCCmec type XI, and was resistant only to β-lactams. Both MRSA were negative for the presence of specific immune-evasion and virulence genes such as those coding for the Panton-Valentine leucocidin, the toxic shock syndrome toxin 1, and the immune evasion cluster genes. Regarding the presence of the major S. aureus enterotoxin genes, the mecC-positive MRSA tested negative, whereas the ST (CC)1 mecA-positive MRSA harbored the seh gene. Among the 104 methicillin-susceptible S. aureus isolates examined for antimicrobial susceptibility, 63 (60.58%) were susceptible to all the antimicrobials tested, and 41 (39.42%) were resistant to at least 1 antimicrobial. In particular, 23 isolates (22.12%) were resistant to tetracycline, 16 (15.38%) to sulfonomides, 14 (13.46%) to trimethoprim and sulfamethoxazole, and 9 (8.65%) to ampicillin, whereas only 1 isolate was resistant to both fluoroquinolones and aminoglycosides. The high prevalence of S. aureus found in bulk tank milk samples and the isolation of MRSA, although at a low prevalence, underlines the importance of adopting control measures against S. aureus in dairy sheep farms to minimize the risks for animal and public health. Moreover, this study represents the first report of mecC-positive MRSA isolation in Italy and would confirm that, among livestock animals, sheep might act as a mecC-MRSA reservoir. Although this lineage seems to be rare in dairy sheep (0.35% of farms tested), because mecC-positive MRSA are difficult to detect by diagnostic routine methods employed for mecA-positive livestock-associated MRSA, diagnostic laboratories should be aware of the importance of searching for the mecC gene in all the mecA-negative S. aureus isolates displaying resistance to oxacillin, cefoxitin, or both. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Ainsworth, William B.; Hughes, Bridget Todd; Au, Wei Chun
2013-10-04
Highlights: •Hug1p overexpression sensitizes wild-type cells to DNA damage and hydroxyurea (HU). •Expression of Hug1p in response to HU treatment is delayed relative to Rnr3p. •MEC1 pathway genes are required for cytoplasmic localization of Hug1p. •Hug1p subcellular compartmentalization to the cytoplasm coincides with Rnr2p–Rnr4p. -- Abstract: The evolutionarily conserved MEC1 checkpoint pathway mediates cell cycle arrest and induction of genes including the RNR (Ribonucleotide reductase) genes and HUG1 (Hydroxyurea, ultraviolet, and gamma radiation) in response to DNA damage and replication arrest. Rnr complex activity is in part controlled by cytoplasmic localization of the Rnr2p–Rnr4p subunits and inactivation of negative regulatorsmore » Sml1p and Dif1p upon DNA damage and hydroxyurea (HU) treatment. We previously showed that a deletion of HUG1 rescues lethality of mec1Δ and suppresses dun1Δ strains. In this study, multiple approaches demonstrate the regulatory response of Hug1p to DNA damage and HU treatment and support its role as a negative effector of the MEC1 pathway. Consistent with our hypothesis, wild-type cells are sensitive to DNA damage and HU when HUG1 is overexpressed. A Hug1 polyclonal antiserum reveals that HUG1 encodes a protein in budding yeast and its MEC1-dependent expression is delayed compared to the rapid induction of Rnr3p in response to HU treatment. Cell biology and subcellular fractionation experiments show localization of Hug1p-GFP to the cytoplasm upon HU treatment. The cytoplasmic localization of Hug1p-GFP is dependent on MEC1 pathway genes and coincides with the cytoplasmic localization of Rnr2p–Rnr4p. Taken together, the genetic interactions, gene expression, and localization studies support a novel role for Hug1p as a negative regulator of the MEC1 checkpoint response through its compartmentalization with Rnr2p–Rnr4p.« less
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Liu, Qian; Ren, Zhiyong Jason; Huang, Cong; Liu, Bingfeng; Ren, Nanqi; Xing, Defeng
2016-01-01
Biohythane is a new and high-value transportation fuel present as a mixture of biomethane and biohydrogen. It has been produced from different organic matters using anaerobic digestion. Bioenergy can be recovered from waste activated sludge through methane production during anaerobic digestion, but energy yield is often insufficient to sludge disposal. Microbial electrolysis cell (MEC) is also a promising approach for bioenergy recovery and waste sludge disposal as higher energy efficiency and biogas production. The systematic understanding of microbial interactions and biohythane production in MEC is still limited. Here, we report biohythane production from waste sludge in biocathode microbial electrolysis cells and reveal syntrophic interactions in microbial communities based on high-throughput sequencing and quantitative PCR targeting 16S rRNA gene. The alkali-pretreated sludge fed MECs (AS-MEC) showed the highest biohythane production rate of 0.148 L·L(-1)-reactor·day(-1), which is 40 and 80 % higher than raw sludge fed MECs (RS-MEC) and anaerobic digestion (open circuit MEC, RS-OCMEC). Current density, metabolite profiles, and hydrogen-methane ratio results all confirm that alkali-pretreatment and microbial electrolysis greatly enhanced sludge hydrolysis and biohythane production. Illumina Miseq sequencing of 16S rRNA gene amplicons indicates that anode biofilm was dominated by exoelectrogenic Geobacter, fermentative bacteria and hydrogen-producing bacteria in the AS-MEC. The cathode biofilm was dominated by fermentative Clostridium. The dominant archaeal populations on the cathodes of AS-MEC and RS-MEC were affiliated with hydrogenotrophic Methanobacterium (98 %, relative abundance) and Methanocorpusculum (77 %), respectively. Multiple pathways of gas production were observed in the same MEC reactor, including fermentative and electrolytic H2 production, as well as hydrogenotrophic methanogenesis and electromethanogenesis. Real-time quantitative PCR analyses showed that higher amount of methanogens were enriched in AS-MEC than that in RS-MEC and RS-OCMEC, suggesting that alkali-pretreated sludge and MEC facilitated hydrogenotrophic methanogen enrichment. This study proves for the first time that biohythane could be produced directly in biocathode MECs using waste sludge. MEC and alkali-pretreatment accelerated enrichment of hydrogenotrophic methanogen and hydrolysis of waste sludge. The results indicate syntrophic interactions among fermentative bacteria, exoelectrogenic bacteria and methanogenic archaea in MECs are critical for highly efficient conversion of complex organics into biohythane, demonstrating that MECs can be more competitive than conventional anaerobic digestion for biohythane production using carbohydrate-deficient substrates. Biohythane production from waste sludge by MEC provides a promising new way for practical application of microbial electrochemical technology.
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McManus, Brenda A; Coleman, David C; Deasy, Emily C; Brennan, Gráinne I; O' Connell, Brian; Monecke, Stefan; Ehricht, Ralf; Leggett, Bernadette; Leonard, Nola; Shore, Anna C
2015-01-01
This study compares the characteristics of Staphylococcus epidermidis (SE) and Staphylococcus haemolyticus (SH) isolates from epidemiologically unrelated infections in humans (Hu) (28 SE-Hu; 8 SH-Hu) and companion animals (CpA) (12 SE-CpA; 13 SH-CpA). All isolates underwent antimicrobial susceptibility testing, multilocus sequence typing and DNA microarray profiling to detect antimicrobial resistance and SCCmec-associated genes. All methicillin-resistant (MR) isolates (33/40 SE, 20/21 SH) underwent dru and mecA allele typing. Isolates were predominantly assigned to sequence types (STs) within a single clonal complex (CC2, SE, 84.8%; CC1, SH, 95.2%). SCCmec IV predominated among MRSE with ST2-MRSE-IVc common to both Hu (40.9%) and CpA (54.5%). Identical mecA alleles and nontypeable dru types (dts) were identified in one ST2-MRSE-IVc Hu and CpA isolate, however, all mecA alleles and 2/4 dts detected among 18 ST2-MRSE-IVc isolates were closely related, sharing >96.5% DNA sequence homology. Although only one ST-SCCmec type combination (ST1 with a non-typeable [NT] SCCmec NT9 [class C mec and ccrB4]) was common to four MRSH-Hu and one MRSH-CpA, all MRSH isolates were closely related based on similar STs, SCCmec genes (V/VT or components thereof), mecA alleles and dts. Overall, 39.6% of MR isolates harbored NT SCCmec elements, and ACME was more common amongst MRSE and CpA isolates. Multidrug resistance (MDR) was detected among 96.7% of isolates but they differed in the prevalence of specific macrolide, aminoglycoside and trimethoprim resistance genes amongst SE and SH isolates. Ciprofloxacin, rifampicin, chloramphenicol [fexA, cat-pC221], tetracycline [tet(K)], aminoglycosides [aadD, aphA3] and fusidic acid [fusB] resistance was significantly more common amongst CpA isolates. SE and SH isolates causing infections in Hu and CpA hosts belong predominantly to STs within a single lineage, harboring similar but variable SCCmec genes, mecA alleles and dts. Host and staphylococcal species-specific characteristics were identified in relation to antimicrobial resistance genes and phenotypes, SCCmec and ACME.
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McManus, Brenda A.; Coleman, David C.; Deasy, Emily C.; Brennan, Gráinne I.; O’ Connell, Brian; Monecke, Stefan; Ehricht, Ralf; Leggett, Bernadette; Leonard, Nola; Shore, Anna C.
2015-01-01
This study compares the characteristics of Staphylococcus epidermidis (SE) and Staphylococcus haemolyticus (SH) isolates from epidemiologically unrelated infections in humans (Hu) (28 SE-Hu; 8 SH-Hu) and companion animals (CpA) (12 SE-CpA; 13 SH-CpA). All isolates underwent antimicrobial susceptibility testing, multilocus sequence typing and DNA microarray profiling to detect antimicrobial resistance and SCCmec-associated genes. All methicillin-resistant (MR) isolates (33/40 SE, 20/21 SH) underwent dru and mecA allele typing. Isolates were predominantly assigned to sequence types (STs) within a single clonal complex (CC2, SE, 84.8%; CC1, SH, 95.2%). SCCmec IV predominated among MRSE with ST2-MRSE-IVc common to both Hu (40.9%) and CpA (54.5%). Identical mecA alleles and nontypeable dru types (dts) were identified in one ST2-MRSE-IVc Hu and CpA isolate, however, all mecA alleles and 2/4 dts detected among 18 ST2-MRSE-IVc isolates were closely related, sharing >96.5% DNA sequence homology. Although only one ST-SCCmec type combination (ST1 with a non-typeable [NT] SCCmec NT9 [class C mec and ccrB4]) was common to four MRSH-Hu and one MRSH-CpA, all MRSH isolates were closely related based on similar STs, SCCmec genes (V/VT or components thereof), mecA alleles and dts. Overall, 39.6% of MR isolates harbored NT SCCmec elements, and ACME was more common amongst MRSE and CpA isolates. Multidrug resistance (MDR) was detected among 96.7% of isolates but they differed in the prevalence of specific macrolide, aminoglycoside and trimethoprim resistance genes amongst SE and SH isolates. Ciprofloxacin, rifampicin, chloramphenicol [fexA, cat-pC221], tetracycline [tet(K)], aminoglycosides [aadD, aphA3] and fusidic acid [fusB] resistance was significantly more common amongst CpA isolates. SE and SH isolates causing infections in Hu and CpA hosts belong predominantly to STs within a single lineage, harboring similar but variable SCCmec genes, mecA alleles and dts. Host and staphylococcal species-specific characteristics were identified in relation to antimicrobial resistance genes and phenotypes, SCCmec and ACME. PMID:26379051
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Plasmid-Encoded Transferable mecB-Mediated Methicillin Resistance in Staphylococcus aureus
van Alen, Sarah; Idelevich, Evgeny A.; Schleimer, Nina; Seggewiß, Jochen; Mellmann, Alexander; Kaspar, Ursula; Peters, Georg
2018-01-01
During cefoxitin-based nasal screening, phenotypically categorized methicillin-resistant Staphylococcus aureus (MRSA) was isolated and tested negative for the presence of the mecA and mecC genes as well as for the SCCmec-orfX junction region. The isolate was found to carry a mecB gene previously described for Macrococcus caseolyticus but not for staphylococcal species. The gene is flanked by β-lactam regulatory genes similar to mecR, mecI, and blaZ and is part of an 84.6-kb multidrug-resistance plasmid that harbors genes encoding additional resistances to aminoglycosides (aacA-aphD, aphA, and aadK) as well as macrolides (ermB) and tetracyclines (tetS). This further plasmidborne β-lactam resistance mechanism harbors the putative risk of acceleration or reacceleration of MRSA spread, resulting in broad ineffectiveness of β-lactams as a main therapeutic application against staphylococcal infections. PMID:29350135
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Ouchenane, Z; Agabou, A; Smati, F; Rolain, J-M; Raoult, D
2013-12-01
Staphylococcal cassette chromosome mec is a genetic mobile element that carries the gene mecA mediating the methicillin resistance in staphylococci. The aim of this study is to type the Staphylococcal cassette chromosome mec (SCCmec) in 64 non-redundant methicillin-resistant Staphylococcus aureus (MRSA) strains recovered at the military hospital of Constantine (Algeria) between 2005 and 2007. Methicillin resistance was detected by oxacillin and cefoxitin discs and PBP2a test, and then confirmed by mecA PCR. The SCCmec complex types were determined by real time PCR. The analysis showed that 50 isolates were hospital acquired (HA-MRSA) and 14 were community-acquired (CA-MRSA). SCCmec type IV and V (traditionally attributed to CA-MRSA) were harbored by both HA-MRSA and CA-MRSA, while SCCmec type I, II and III were not recorded. These findings motivate more investigations to be carried on HA-MRSA in our hospital and other national health care centers. Copyright © 2013. Published by Elsevier SAS.
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Smith, Cody J.; O’Brien, Timothy; Chatzigeorgiou, Marios; Spencer, W. Clay; Feingold-Link, Elana; Husson, Steven J.; Hori, Sayaka; Mitani, Shohei; Gottschalk, Alexander; Schafer, William R.; Miller, David M.
2013-01-01
SUMMARY Sensory neurons adopt distinct morphologies and functional modalities to mediate responses to specific stimuli. Transcription factors and their downstream effectors orchestrate this outcome but are incompletely defined. Here, we show that different classes of mechanosensory neurons in C. elegans are distinguished by the combined action of the transcription factors MEC-3, AHR-1, and ZAG-1. Low levels of MEC-3 specify the elaborate branching pattern of PVD nociceptors, whereas high MEC-3 is correlated with the simple morphology of AVM and PVM touch neurons. AHR-1 specifies AVM touch neuron fate by elevating MEC-3 while simultaneously blocking expression of nociceptive genes such as the MEC-3 target, the claudin-like membrane protein HPO-30, that promotes the complex dendritic branching pattern of PVD. ZAG-1 exercises a parallel role to prevent PVM from adopting the PVD fate. The conserved dendritic branching function of the Drosophila AHR-1 homolog, Spineless, argues for similar pathways in mammals. PMID:23889932
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Yu, Dongliang; Pi, Borui; Chen, Yan; Wang, Yanfei; Ruan, Zhi; Otto, Michael; Yu, Yunsong
2014-01-01
Staphylococcal cassette chromosome (SCC) elements contribute considerably to virulence and resistance to antibiotic agents in staphylococci. SCC elements in coagulase-negative staphylococci (CoNS) are highly diverse and there is evidence suggesting that they serve as a reservoir for antibiotic resistance genes in methicillin-resistant Staphylococcus aureus (MRSA). However, only a small number of SCC elements have been characterized in CoNS and their exact roles in the emergence and evolution of MRSA remain to be demonstrated. Here, we determined the structure of an SCC composite island (CISH32) found in the clinical Staphylococcus haemolyticus isolate SH32 by whole-genome DNA sequencing. CISH32 was 48 kb in length and mainly composed of two imperfect SCC elements, namely (i) a ΨSCCmec(SH32) part containing a class C1 mec gene complex but lacking ccr genes and (ii) a SCCSH32 part with a ccrA5B3 gene complex but lacking mec genes. In addition, CISH32 contained a type III restriction-modification system and several resistance loci, for example genes conferring resistance to cadmium and arsenic. ΨSCCmec(SH32) is almost entirely identical to a pseudo SCCmec element found in S. haemolyticus WCH1 and shares pronounced sequence similarity to a ΨSCCmec element of S. haemolyticus JCSC1435. However, staphylococci other than S. haemolyticus, including S. aureus and S. epidermidis, contain homologs of SCCSH32 that are more similar to SCCSH32 than those elements found in S. haemolyticus, suggesting that CISH32 of S. haemolyticus SH32 was assembled in recent evolutionary events. Moreover, the composite structure of CISH32 indicates that the detection of class C1 mec and ccrA5B3 gene complexes in S. haemolyticus does not always indicate the existence of a UT9-type SCCmec element, which has remained questionable.
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Yu, Dongliang; Pi, Borui; Chen, Yan; Wang, Yanfei; Ruan, Zhi; Otto, Michael; Yu, Yunsong
2014-01-01
Staphylococcal cassette chromosome (SCC) elements contribute considerably to virulence and resistance to antibiotic agents in staphylococci. SCC elements in coagulase-negative staphylococci (CoNS) are highly diverse and there is evidence suggesting that they serve as a reservoir for antibiotic resistance genes in methicillin-resistant Staphylococcus aureus (MRSA). However, only a small number of SCC elements have been characterized in CoNS and their exact roles in the emergence and evolution of MRSA remain to be demonstrated. Here, we determined the structure of an SCC composite island (CISH32) found in the clinical Staphylococcus haemolyticus isolate SH32 by whole-genome DNA sequencing. CISH32 was 48 kb in length and mainly composed of two imperfect SCC elements, namely (i) a ΨSCCmec(SH32) part containing a class C1 mec gene complex but lacking ccr genes and (ii) a SCCSH32 part with a ccrA5B3 gene complex but lacking mec genes. In addition, CISH32 contained a type III restriction-modification system and several resistance loci, for example genes conferring resistance to cadmium and arsenic. ΨSCCmec(SH32) is almost entirely identical to a pseudo SCCmec element found in S. haemolyticus WCH1 and shares pronounced sequence similarity to a ΨSCCmec element of S. haemolyticus JCSC1435. However, staphylococci other than S. haemolyticus, including S. aureus and S. epidermidis, contain homologs of SCCSH32 that are more similar to SCCSH32 than those elements found in S. haemolyticus, suggesting that CISH32 of S. haemolyticus SH32 was assembled in recent evolutionary events. Moreover, the composite structure of CISH32 indicates that the detection of class C1 mec and ccrA5B3 gene complexes in S. haemolyticus does not always indicate the existence of a UT9-type SCCmec element, which has remained questionable. PMID:24466348
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Genetic analysis of mecA gene and detection of homologue pbpD in Stahylococcus sciuri group
Calazans-Silva, Amanda C.; Medeiros, Pedro T.C.; Araujo, Dayanne M.; Carvalho, Bruno O.; Coelho, Irene S.; Coelho, Shana M.O.; Souza, Miliane M.S.
2014-01-01
Oxacillin/methicillin-resistance is related to the mecA and its regulatory genes mecR1 and mecI. Its origin is still unknown, although evidences support that it is related to CNS, once mecA and a homologue gene, pbpD, were both detected in Staphylococcus sciuri species group. The present work evaluated 210 samples of skin and ear swabs from rodents and 60 nasal swabs from equines of Army Biologic Institute, Rio de Janeiro. Pheno- and genotypic characterization provided 59.52% (25/42) and 78.57% (11/14) S. lentus and S. sciuri, respectively. It was observed that although all S. sciuri isolates tested positive for pbpD, there was no correlation with oxacillin-resistance. On the other hand, isolates tested positive for mecA gene also presented phenotypic oxacillin-resistance in at least one assay. The alignment of the mecA gene showed that the nucleotide sequences were sorted into 2 different groups, one comprising the bovine strains and the other containing human and equine strains. PMID:25242954
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Chovanová, Romana; Mikulášová, Mária; Vaverková, Štefánia
2016-01-01
The essential oil (EO) from Salvia sclarea was shown to increase the susceptibility of methicillin resistant Staphylococcus epidermidis (MRSE) isolates to oxacillin. The purpose of this study was to investigate the effect of EO from S. sclarea on expression of mecA gene of MRSE carrying different types of staphylococcal chromosomal cassette (SCCmec) and to evaluate potential synergistic effect of EO with oxacillin. Using real-time PCR we found that EO alone inhibited the expression of the resistant genes mecA, mecR1, and mecI and blaZ, blaR1, and blaI. The use of the combination of EO with oxacillin resulted in significantly inhibited expression of mecA gene in all tested strains with different types of SCCmec. Using time-kill assay and checkerboard assay we confirmed synergistic effect of EO from S. sclarea and oxacillin in MRSE. PMID:26880926
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Ma, Xiao Xue; Sun, Dan Dan; Hu, Jian; Wang, En Hua; Luo, En Jie
2011-06-01
In the present study, we report on the reduced susceptibility to teicoplanin among clinical isolates of Staphylococcus haemolyticus in a hematology ward of a teaching hospital. The molecular characterization of 17 S. haemolyticus strains was performed using mec gene complex classification, pulsed-field gel electrophoresis analysis, and minimum inhibitory concentration examination. Pulsotype A strains carrying a class C2 mec gene complex were the most prevalent strains, at 64.7%. In vivo selection of stepwise increase in resistance to vancomycin and teicoplanin was observed in three S. haemolyticus strains serially isolated from a case patient. The results of the present study suggest the regional spread of certain S. haemolyticus clones with diminished susceptibility to glycopeptides, emphasizing the need for continuous monitoring of minimum inhibitory concentration levels of vancomycin and teicoplanin in S. haemolyticus strains, and the importance of infection control practices to prevent its transmission.
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Wan, Min Tao; Chou, Chin Cheng
2014-11-01
Methicillin-resistant Staphylococcus aureus (MRSA) is a potential zoonotic agent. Municipal wastewater treatment plants (WWTPs) can be reservoirs for MRSA dissemination. It is unclear, however, whether MRSA and its β-lactam resistance gene (mecA) can be spread from WWTPs that treat the wastewater of swine auction markets. The aims of the study were to compare (1) the abundance of the mecA gene in one municipal (M-) and one swine (S-) WWTP and (2) the genotypic and phenotypic characteristics of MRSA isolates from these two types of WWTPs. The concentrations of mecA gene from 96 wastewater samples were quantified using real-time quantitative polymerase chain reaction (real-time qPCR). One hundred and thirteen MRSA isolates were recovered and were characterized by antimicrobial susceptibility testing, minimum inhibitory concentrations (MICs), and staphylococcal cassette chromosome mec (SCCmec) typing. The mecA gene could be detected in all the wastewater samples. A high abundance of recovered mecA gene (2.6 × 10(1) to 1.9 × 10(4) gene copies μg(-1) of total DNA) in swine slaughterhouse wastewater implied a correspondingly high transferring/receiving potential. All MRSA isolates were multidrug resistant (MDR) and showed high MICs to different antimicrobials. The M-WWTP MRSA isolates harbored SCCmec II-IV and VII, whereas those from the S-WWTP harbored SCCmec V and IX. In conclusion, wastewater from swine slaughterhouses can make these slaughterhouses potential hotspots for the dissemination of mecA gene and MRSA, and the high MICs of MRSA from both WWTP origins may pose a health risk not only to workers but also to the general public. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Requirement of the Mre11 complex and exonuclease 1 for activation of the Mec1 signaling pathway.
Nakada, Daisuke; Hirano, Yukinori; Sugimoto, Katsunori
2004-11-01
The large protein kinases, ataxia-telangiectasia mutated (ATM) and ATM-Rad3-related (ATR), orchestrate DNA damage checkpoint pathways. In budding yeast, ATM and ATR homologs are encoded by TEL1 and MEC1, respectively. The Mre11 complex consists of two highly related proteins, Mre11 and Rad50, and a third protein, Xrs2 in budding yeast or Nbs1 in mammals. The Mre11 complex controls the ATM/Tel1 signaling pathway in response to double-strand break (DSB) induction. We show here that the Mre11 complex functions together with exonuclease 1 (Exo1) in activation of the Mec1 signaling pathway after DNA damage and replication block. Mec1 controls the checkpoint responses following UV irradiation as well as DSB induction. Correspondingly, the Mre11 complex and Exo1 play an overlapping role in activation of DSB- and UV-induced checkpoints. The Mre11 complex and Exo1 collaborate in producing long single-stranded DNA (ssDNA) tails at DSB ends and promote Mec1 association with the DSBs. The Ddc1-Mec3-Rad17 complex associates with sites of DNA damage and modulates the Mec1 signaling pathway. However, Ddc1 association with DSBs does not require the function of the Mre11 complex and Exo1. Mec1 controls checkpoint responses to stalled DNA replication as well. Accordingly, the Mre11 complex and Exo1 contribute to activation of the replication checkpoint pathway. Our results provide a model in which the Mre11 complex and Exo1 cooperate in generating long ssDNA tracts and thereby facilitate Mec1 association with sites of DNA damage or replication block.
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Ramsden, Helen L; Sürmeli, Gülşen; McDonagh, Steven G; Nolan, Matthew F
2015-01-01
Neural circuits in the medial entorhinal cortex (MEC) encode an animal's position and orientation in space. Within the MEC spatial representations, including grid and directional firing fields, have a laminar and dorsoventral organization that corresponds to a similar topography of neuronal connectivity and cellular properties. Yet, in part due to the challenges of integrating anatomical data at the resolution of cortical layers and borders, we know little about the molecular components underlying this organization. To address this we develop a new computational pipeline for high-throughput analysis and comparison of in situ hybridization (ISH) images at laminar resolution. We apply this pipeline to ISH data for over 16,000 genes in the Allen Brain Atlas and validate our analysis with RNA sequencing of MEC tissue from adult mice. We find that differential gene expression delineates the borders of the MEC with neighboring brain structures and reveals its laminar and dorsoventral organization. We propose a new molecular basis for distinguishing the deep layers of the MEC and show that their similarity to corresponding layers of neocortex is greater than that of superficial layers. Our analysis identifies ion channel-, cell adhesion- and synapse-related genes as candidates for functional differentiation of MEC layers and for encoding of spatial information at different scales along the dorsoventral axis of the MEC. We also reveal laminar organization of genes related to disease pathology and suggest that a high metabolic demand predisposes layer II to neurodegenerative pathology. In principle, our computational pipeline can be applied to high-throughput analysis of many forms of neuroanatomical data. Our results support the hypothesis that differences in gene expression contribute to functional specialization of superficial layers of the MEC and dorsoventral organization of the scale of spatial representations.
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Ramsden, Helen L.; Sürmeli, Gülşen; McDonagh, Steven G.; Nolan, Matthew F.
2015-01-01
Neural circuits in the medial entorhinal cortex (MEC) encode an animal’s position and orientation in space. Within the MEC spatial representations, including grid and directional firing fields, have a laminar and dorsoventral organization that corresponds to a similar topography of neuronal connectivity and cellular properties. Yet, in part due to the challenges of integrating anatomical data at the resolution of cortical layers and borders, we know little about the molecular components underlying this organization. To address this we develop a new computational pipeline for high-throughput analysis and comparison of in situ hybridization (ISH) images at laminar resolution. We apply this pipeline to ISH data for over 16,000 genes in the Allen Brain Atlas and validate our analysis with RNA sequencing of MEC tissue from adult mice. We find that differential gene expression delineates the borders of the MEC with neighboring brain structures and reveals its laminar and dorsoventral organization. We propose a new molecular basis for distinguishing the deep layers of the MEC and show that their similarity to corresponding layers of neocortex is greater than that of superficial layers. Our analysis identifies ion channel-, cell adhesion- and synapse-related genes as candidates for functional differentiation of MEC layers and for encoding of spatial information at different scales along the dorsoventral axis of the MEC. We also reveal laminar organization of genes related to disease pathology and suggest that a high metabolic demand predisposes layer II to neurodegenerative pathology. In principle, our computational pipeline can be applied to high-throughput analysis of many forms of neuroanatomical data. Our results support the hypothesis that differences in gene expression contribute to functional specialization of superficial layers of the MEC and dorsoventral organization of the scale of spatial representations. PMID:25615592
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Wei, Zhengkai; Xiao, Chong; Guo, Changming; Zhang, Xu; Wang, Yanan; Wang, Jingjing; Yang, Zhengtao; Fu, Yunhe
2017-06-01
Bovine mastitis is one of the most costly and prevalent disease affecting dairy cows worldwide. It was reported that Staphylococcus aureus could internalize into bovine mammary epithelial cells (bMEC) and induce mastitis. Some short chain fatty acids (SCFA) have shown to suppress S. aureus invasion into bMEC and regulate antimicrobial peptides expression. But it has not been evaluated that sodium acetate has the similar effect. The aim of this study was to investigate the effect of sodium acetate on the invasion of bovine mammary epithelial cells (bMEC) by S. aureus. Gentamicin protection assay showed that the invasion of S. aureus into bMEC was inhibited by sodium acetate in a dose-dependent manner. Sodium acetate (0.25-5 mM) did not affect S. aureus growth and bMEC viability. The TAP gene level was decreased, while the BNBD5 mRNA level was enhanced in sodium acetate treated bMEC. In sodium acetate treated and S. aureus challenged bMEC, the TAP gene expression was increased and BNBD5 gene expression was not modified at low concentrations, but decreased at high concentrations. The Nitric oxide (NO) production of bMEC after S. aureus stimulation was decreased by sodium acetate treatment. Furthermore, sodium acetate treatment suppressed S. aureus-induced NF-κB activation in bMEC in a dose manner. In conclusion, our results suggested that sodium acetate exerts an inhibitory property on S. aureus internalization and modulates antimicrobial peptides gene expression. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Pasko, Chris; Dunn, John; Jaeckel, Heidi; Nieuwlandt, Dan; Weed, Diane; Woodruff, Evelyn; Zheng, Xiaotian
2012-01-01
Rapid diagnosis of staphylococcal bacteremia directs appropriate antimicrobial therapy, leading to improved patient outcome. We describe herein a rapid test (<75 min) that can identify the major pathogenic strains of Staphylococcus to the species level as well as the presence or absence of the methicillin resistance determinant gene, mecA. The test, Staph ID/R, combines a rapid isothermal nucleic acid amplification method, helicase-dependent amplification (HDA), with a chip-based array that produces unambiguous visible results. The analytic sensitivity was 1 CFU per reaction for the mecA gene and was 1 to 250 CFU per reaction depending on the staphylococcal species present in the positive blood culture. Staph ID/R has excellent specificity as well, with no cross-reactivity observed. We validated the performance of Staph ID/R by testing 104 frozen clinical positive blood cultures and comparing the results with rpoB gene or 16S rRNA gene sequencing for species identity determinations and mecA gene PCR to confirm mecA gene results. Staph ID/R agreed with mecA gene PCR for all samples and agreed with rpoB/16S rRNA gene sequencing in all cases except for one sample that contained a mixture of two staphylococcal species, one of which Staph ID/R correctly identified, for an overall agreement of 99.0% (P < 0.01). Staph ID/R could potentially be used to positively affect patient management for Staphylococcus-mediated bacteremia. PMID:22170912
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Kaya, Hülya; Hasman, Henrik; Larsen, Jesper; Stegger, Marc; Johannesen, Thor Bech; Allesøe, Rosa Lundbye; Lemvigh, Camilla Koldbæk; Aarestrup, Frank Møller; Lund, Ole; Larsen, Anders Rhod
2018-01-01
Typing of methicillin-resistant Staphylococcus aureus (MRSA) is important in infection control and surveillance. The current nomenclature of MRSA includes the genetic background of the S. aureus strain determined by multilocus sequence typing (MLST) or equivalent methods like spa typing and typing of the mobile genetic element staphylococcal cassette chromosome mec (SCC mec ), which carries the mecA or mecC gene. Whereas MLST and spa typing are relatively simple, typing of SCC mec is less trivial because of its heterogeneity. Whole-genome sequencing (WGS) provides the essential data for typing of the genetic background and SCC mec , but so far, no bioinformatic tools for SCC mec typing have been available. Here, we report the development and evaluation of SCC mec Finder for characterization of the SCC mec element from S. aureus WGS data. SCC mec Finder is able to identify all SCC mec element types, designated I to XIII, with subtyping of SCC mec types IV (2B) and V (5C2). SCC mec elements are characterized by two different gene prediction approaches to achieve correct annotation, a Basic Local Alignment Search Tool (BLAST)-based approach and a k -mer-based approach. Evaluation of SCC mec Finder by using a diverse collection of clinical isolates ( n = 93) showed a high typeability level of 96.7%, which increased to 98.9% upon modification of the default settings. In conclusion, SCC mec Finder can be an alternative to more laborious SCC mec typing methods and is freely available at https://cge.cbs.dtu.dk/services/SCCmecFinder. IMPORTANCE SCC mec in MRSA is acknowledged to be of importance not only because it contains the mecA or mecC gene but also for staphylococcal adaptation to different environments, e.g., in hospitals, the community, and livestock. Typing of SCC mec by PCR techniques has, because of its heterogeneity, been challenging, and whole-genome sequencing has only partially solved this since no good bioinformatic tools have been available. In this article, we describe the development of a new bioinformatic tool, SCC mec Finder, that includes most of the needs for infection control professionals and researchers regarding the interpretation of SCC mec elements. The software detects all of the SCC mec elements accepted by the International Working Group on the Classification of Staphylococcal Cassette Chromosome Elements, and users will be prompted if diverging and potential new elements are uploaded. Furthermore, SCC mec Finder will be curated and updated as new elements are found and it is easy to use and freely accessible.
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Xu, Zhen; Mkrtchyan, Hermine V.; Cutler, Ronald R.
2015-01-01
Antibiotic resistance in bacteria isolated from non-healthcare environments, is a potential problem to public health. In our survey a total of 71 coagulase negative staphylococci (CNS) belonging to 11 different species were isolated from three large hotels in London, UK. The most prevalent species was Staphylococcus haemolyticus, with S. hominis, S. warneri, S. cohnii, and Staphylococcus epidermidis commonly detected. Antimicrobial susceptibilities and carriage of the mecA gene were determined for all of these isolates. Most (85.9%) staphylococci were resistant to multiple antibiotics with all displaying increased susceptibility toward penicillin, fusidic acid, erythromycin, and cefepime. Twenty-one (29.5%) of the isolates were mecA positive, however MIC values to oxacillin, normally associated with the carriage of mecA, varied widely in this group (from 0.06 to 256 mg/L). Fifteen of the twenty-one mecA positive isolates carried SCCmec of these seven were type V, one type I, one type II, and one type IV. Additionally, five of these 15 isolates carried a previously unreported type, 1A, which involves an association between class A mec complex and ccr type 1. The remaining six of the 21 isolates were non-typeable and carried a combination of class A mec complex and ccrC. In addition to this, we also report on new MLST types which were assigned for five S. epidermidis isolates. Four out of these five isolates had MICs between 0.06 and 256 mg/L to oxacillin and would be regarded as clinically susceptible but one isolate had a high oxacillin MIC of 256 mg/L. We demonstrated widespread multiple drug resistance among different staphylococcal species isolated from non-healthcare environments highlighting the potential for these species to act as a reservoir for methicillin and other forms of drug resistance. PMID:26441881
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Xu, Zhen; Mkrtchyan, Hermine V; Cutler, Ronald R
2015-01-01
Antibiotic resistance in bacteria isolated from non-healthcare environments, is a potential problem to public health. In our survey a total of 71 coagulase negative staphylococci (CNS) belonging to 11 different species were isolated from three large hotels in London, UK. The most prevalent species was Staphylococcus haemolyticus, with S. hominis, S. warneri, S. cohnii, and Staphylococcus epidermidis commonly detected. Antimicrobial susceptibilities and carriage of the mecA gene were determined for all of these isolates. Most (85.9%) staphylococci were resistant to multiple antibiotics with all displaying increased susceptibility toward penicillin, fusidic acid, erythromycin, and cefepime. Twenty-one (29.5%) of the isolates were mecA positive, however MIC values to oxacillin, normally associated with the carriage of mecA, varied widely in this group (from 0.06 to 256 mg/L). Fifteen of the twenty-one mecA positive isolates carried SCCmec of these seven were type V, one type I, one type II, and one type IV. Additionally, five of these 15 isolates carried a previously unreported type, 1A, which involves an association between class A mec complex and ccr type 1. The remaining six of the 21 isolates were non-typeable and carried a combination of class A mec complex and ccrC. In addition to this, we also report on new MLST types which were assigned for five S. epidermidis isolates. Four out of these five isolates had MICs between 0.06 and 256 mg/L to oxacillin and would be regarded as clinically susceptible but one isolate had a high oxacillin MIC of 256 mg/L. We demonstrated widespread multiple drug resistance among different staphylococcal species isolated from non-healthcare environments highlighting the potential for these species to act as a reservoir for methicillin and other forms of drug resistance.
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Pi, Borui; Yu, Meihong; Chen, Yagang; Yu, Yunsong; Li, Lanjuan
2009-06-01
The aim of this study was to investigate the prevalence and characteristics of ACME (arginine catabolic mobile element)-arcA-positive isolates among meticillin-resistant Staphylococcus haemolyticus (MRSH). ACME-arcA, native arcA and SCCmec elements were detected by PCR. Susceptibilities to 10 antimicrobial agents were compared between ACME-arcA-positive and -negative isolates by chi-square test. PFGE was used to investigate the clonal relatedness of ACME-arcA-positive isolates. The phylogenetic relationships of ACME-arcA and native arcA were analysed using the neighbour-joining methods of mega software. A total of 42 (47.7 %) of 88 isolates distributed in 13 PFGE types were positive for the ACME-arcA gene. There were no significant differences in antimicrobial susceptibility between ACME-arcA-positive and -negative isolates. A novel ccr allotype (ccrAB(SHP)) was identified in ACME-arcA-positive isolates. Among 42 ACME-arcA-positive isolates: 8 isolates harboured SCCmec V, 8 isolates harboured class C1 mec complex and ccrAB(SHP); 22 isolates harbouring class C1 mec complex and 4 isolates harbouring class C2 mec complex were negative for all known ccr allotypes. The ACME-arcA-positive isolates were first found in MRSH with high prevalence and clonal diversity, which suggests a mobility of ACME within MRSH. The results from this study revealed that MRSH is likely to be one of the potential reservoirs of ACME for Staphylococcus aureus.
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RNA-processing proteins regulate Mec1/ATR activation by promoting generation of RPA-coated ssDNA.
Manfrini, Nicola; Trovesi, Camilla; Wery, Maxime; Martina, Marina; Cesena, Daniele; Descrimes, Marc; Morillon, Antonin; d'Adda di Fagagna, Fabrizio; Longhese, Maria Pia
2015-02-01
Eukaryotic cells respond to DNA double-strand breaks (DSBs) by activating a checkpoint that depends on the protein kinases Tel1/ATM and Mec1/ATR. Mec1/ATR is activated by RPA-coated single-stranded DNA (ssDNA), which arises upon nucleolytic degradation (resection) of the DSB. Emerging evidences indicate that RNA-processing factors play critical, yet poorly understood, roles in genomic stability. Here, we provide evidence that the Saccharomyces cerevisiae RNA decay factors Xrn1, Rrp6 and Trf4 regulate Mec1/ATR activation by promoting generation of RPA-coated ssDNA. The lack of Xrn1 inhibits ssDNA generation at the DSB by preventing the loading of the MRX complex. By contrast, DSB resection is not affected in the absence of Rrp6 or Trf4, but their lack impairs the recruitment of RPA, and therefore of Mec1, to the DSB. Rrp6 and Trf4 inactivation affects neither Rad51/Rad52 association nor DSB repair by homologous recombination (HR), suggesting that full Mec1 activation requires higher amount of RPA-coated ssDNA than HR-mediated repair. Noteworthy, deep transcriptome analyses do not identify common misregulated gene expression that could explain the observed phenotypes. Our results provide a novel link between RNA processing and genome stability. © 2014 The Authors.
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Chen, Z; Chen, J; Gu, Y; Hu, C; Li, J-L; Lin, S; Shen, H; Cao, C; Gao, R; Li, J; Ha, P K; Kaye, F J; Griffin, J D; Wu, L
2014-07-17
Salivary gland tumors (SGT) are a group of highly heterogeneous head and neck malignancies with widely varied clinical outcomes and no standard effective treatments. The CRTC1-MAML2 fusion oncogene, encoded by a recurring chromosomal translocation t(11;19)(q14-21;p12-13), is a frequent genetic alteration found in >50% of mucoepidermoid carcinomas (MEC), the most common malignant SGT. In this study, we aimed to define the role of the CRTC1-MAML2 oncogene in the maintenance of MEC tumor growth and to investigate critical downstream target genes and pathways for therapeutic targeting of MEC. By performing gene expression analyses and functional studies via RNA interference and pharmacological modulation, we determined the importance of the CRTC1-MAML2 fusion gene and its downstream AREG-EGFR signaling in human MEC cancer cell growth and survival in vitro and in vivo using human MEC xenograft models. We found that CRTC1-MAML2 fusion oncogene was required for the growth and survival of fusion-positive human MEC cancer cells in vitro and in vivo. The CRTC1-MAML2 oncoprotein induced the upregulation of the epidermal growth factor receptor (EGFR) ligand Amphiregulin (AREG) by co-activating the transcription factor CREB, and AREG subsequently activated EGFR signaling in an autocrine manner that promoted MEC cell growth and survival. Importantly, CRTC1-MAML2-positive MEC cells were highly sensitive to EGFR signaling inhibition. Therefore, our study revealed that aberrantly activated AREG-EGFR signaling is required for CRTC1-MAML2-positive MEC cell growth and survival, suggesting that EGFR-targeted therapies will benefit patients with advanced, unresectable CRTC1-MAML2-positive MEC.
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Schurek, Eva-Maria; Völker, Linus A.; Tax, Judit; Lamkemeyer, Tobias; Rinschen, Markus M.; Ungrue, Denise; Kratz, John E.; Sirianant, Lalida; Kunzelmann, Karl; Chalfie, Martin; Schermer, Bernhard; Benzing, Thomas; Höhne, Martin
2014-01-01
Mutations in the NPHS2 gene are a major cause of steroid-resistant nephrotic syndrome, a severe human kidney disorder. The NPHS2 gene product podocin is a key component of the slit diaphragm cell junction at the kidney filtration barrier and part of a multiprotein-lipid supercomplex. A similar complex with the podocin ortholog MEC-2 is required for touch sensation in Caenorhabditis elegans. Although podocin and MEC-2 are membrane-associated proteins with a predicted hairpin-like structure and amino and carboxyl termini facing the cytoplasm, this membrane topology has not been convincingly confirmed. One particular mutation that causes kidney disease in humans (podocinP118L) has also been identified in C. elegans in genetic screens for touch insensitivity (MEC-2P134S). Here we show that both mutant proteins, in contrast to the wild-type variants, are N-glycosylated because of the fact that the mutant C termini project extracellularly. PodocinP118L and MEC-2P134S did not fractionate in detergent-resistant membrane domains. Moreover, mutant podocin failed to activate the ion channel TRPC6, which is part of the multiprotein-lipid supercomplex, indicative of the fact that cholesterol recruitment to the ion channels, an intrinsic function of both proteins, requires C termini facing the cytoplasmic leaflet of the plasma membrane. Taken together, this study demonstrates that the carboxyl terminus of podocin/MEC-2 has to be placed at the inner leaflet of the plasma membrane to mediate cholesterol binding and contribute to ion channel activity, a prerequisite for mechanosensation and the integrity of the kidney filtration barrier. PMID:24596097
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de Lencastre, Hermínia; Tomasz, Alexander
2017-01-01
ABSTRACT Most methicillin-resistant Staphylococcus aureus (MRSA) strains are resistant to beta-lactam antibiotics due to the presence of the mecA gene, encoding an extra penicillin-binding protein (PBP2A) that has low affinity for virtually all beta-lactam antibiotics. Recently, a new resistance determinant—the mecC gene—was identified in S. aureus isolates recovered from humans and dairy cattle. Although having typically low MICs to beta-lactam antibiotics, MRSA strains with the mecC determinant are also capable of expressing high levels of oxacillin resistance when in an optimal genetic background. In order to test the impact of extensive beta-lactam selection on the emergence of mecC-carrying strains with high levels of antibiotic resistance, we exposed the prototype mecC-carrying MRSA strain, LGA251, to increasing concentrations of oxacillin. LGA251 was able to rapidly adapt to high concentrations of oxacillin in growth medium. In such laboratory mutants with increased levels of oxacillin resistance, we identified mutations in genes with no relationship to the mecC regulatory system, indicating that the genetic background plays an important role in the establishment of the levels of oxacillin resistance. Our data also indicate that the stringent stress response plays a critical role in the beta-lactam antibiotic resistance phenotype of MRSA strains carrying the mecC determinant. PMID:28069659
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Velasco, Valeria; Sherwood, Julie S.; Rojas-García, Pedro P.; Logue, Catherine M.
2014-01-01
The aim of this study was to compare a real-time PCR assay, with a conventional culture/PCR method, to detect S. aureus, mecA and Panton-Valentine Leukocidin (PVL) genes in animals and retail meat, using a two-step selective enrichment protocol. A total of 234 samples were examined (77 animal nasal swabs, 112 retail raw meat, and 45 deli meat). The multiplex real-time PCR targeted the genes: nuc (identification of S. aureus), mecA (associated with methicillin resistance) and PVL (virulence factor), and the primary and secondary enrichment samples were assessed. The conventional culture/PCR method included the two-step selective enrichment, selective plating, biochemical testing, and multiplex PCR for confirmation. The conventional culture/PCR method recovered 95/234 positive S. aureus samples. Application of real-time PCR on samples following primary and secondary enrichment detected S. aureus in 111/234 and 120/234 samples respectively. For detection of S. aureus, the kappa statistic was 0.68–0.88 (from substantial to almost perfect agreement) and 0.29–0.77 (from fair to substantial agreement) for primary and secondary enrichments, using real-time PCR. For detection of mecA gene, the kappa statistic was 0–0.49 (from no agreement beyond that expected by chance to moderate agreement) for primary and secondary enrichment samples. Two pork samples were mecA gene positive by all methods. The real-time PCR assay detected the mecA gene in samples that were negative for S. aureus, but positive for Staphylococcus spp. The PVL gene was not detected in any sample by the conventional culture/PCR method or the real-time PCR assay. Among S. aureus isolated by conventional culture/PCR method, the sequence type ST398, and multi-drug resistant strains were found in animals and raw meat samples. The real-time PCR assay may be recommended as a rapid method for detection of S. aureus and the mecA gene, with further confirmation of methicillin-resistant S. aureus (MRSA) using the standard culture method. PMID:24849624
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Velasco, Valeria; Sherwood, Julie S; Rojas-García, Pedro P; Logue, Catherine M
2014-01-01
The aim of this study was to compare a real-time PCR assay, with a conventional culture/PCR method, to detect S. aureus, mecA and Panton-Valentine Leukocidin (PVL) genes in animals and retail meat, using a two-step selective enrichment protocol. A total of 234 samples were examined (77 animal nasal swabs, 112 retail raw meat, and 45 deli meat). The multiplex real-time PCR targeted the genes: nuc (identification of S. aureus), mecA (associated with methicillin resistance) and PVL (virulence factor), and the primary and secondary enrichment samples were assessed. The conventional culture/PCR method included the two-step selective enrichment, selective plating, biochemical testing, and multiplex PCR for confirmation. The conventional culture/PCR method recovered 95/234 positive S. aureus samples. Application of real-time PCR on samples following primary and secondary enrichment detected S. aureus in 111/234 and 120/234 samples respectively. For detection of S. aureus, the kappa statistic was 0.68-0.88 (from substantial to almost perfect agreement) and 0.29-0.77 (from fair to substantial agreement) for primary and secondary enrichments, using real-time PCR. For detection of mecA gene, the kappa statistic was 0-0.49 (from no agreement beyond that expected by chance to moderate agreement) for primary and secondary enrichment samples. Two pork samples were mecA gene positive by all methods. The real-time PCR assay detected the mecA gene in samples that were negative for S. aureus, but positive for Staphylococcus spp. The PVL gene was not detected in any sample by the conventional culture/PCR method or the real-time PCR assay. Among S. aureus isolated by conventional culture/PCR method, the sequence type ST398, and multi-drug resistant strains were found in animals and raw meat samples. The real-time PCR assay may be recommended as a rapid method for detection of S. aureus and the mecA gene, with further confirmation of methicillin-resistant S. aureus (MRSA) using the standard culture method.
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Osman, Kamelia; Alvarez-Ordóñez, Avelino; Ruiz, Lorena; Badr, Jihan; ElHofy, Fatma; Al-Maary, Khalid S; Moussa, Ihab M I; Hessain, Ashgan M; Orabi, Ahmed; Saad, Alaa; Elhadidy, Mohamed
2017-05-10
The objectives of this study were to characterize the diversity and magnitude of antimicrobial resistance among Staphylococcus species recovered from imported beef meat sold in the Egyptian market and the potential mechanisms underlying the antimicrobial resistance phenotypes including harboring of resistance genes (mecA, cfr, gyrA, gyrB, and grlA) and biofilm formation. The resistance gene mecA was detected in 50% of methicillin-resistant non-Staphylococcus aureus isolates (4/8). Interestingly, our results showed that: (i) resistance genes mecA, gyrA, gyrB, grlA, and cfr were absent in Staphylococcus hominis and Staphylococcus hemolyticus isolates, although S. hominis was phenotypically resistant to methicillin (MR-non-S. aureus) while S. hemolyticus was resistant to vancomycin only; (ii) S. aureus isolates did not carry the mecA gene (100%) and were phenotypically characterized as methicillin- susceptible S. aureus (MSS); and (iii) the resistance gene mecA was present in one isolate (1/3) of Staphylococcus lugdunensis that was phenotypically characterized as methicillin-susceptible non-S. aureus (MSNSA). Our findings highlight the potential risk for consumers, in the absence of actionable risk management information systems, of imported foods and advice a strict implementation of international standards by different venues such as CODEX to avoid the increase in prevalence of coagulase positive and coagulase negative Staphylococcus isolates and their antibiotic resistance genes in imported beef meat at the Egyptian market.
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Berglund, C; Söderquist, B
2008-11-01
The first methicillin-resistant Staphylococcus aureus (MRSA) strain originated when a staphylococcal cassette chromosome mec (SCCmec) with the gene mecA was integrated into the chromosome of a susceptible S. aureus cell. The SCCmec elements are common among the coagulase-negative staphylococci, e.g. Staphylococcus haemolyticus, and these are considered to be potential SCCmec donors when new clones of MRSA arise. An outbreak of MRSA occurred at a neonatal intensive-care unit, and the isolates were all of sequence type (ST) 45, as characterized by multilocus sequence typing, but were not typeable with respect to SCCmec types I, II, III or IV. During the same time period, methicillin-resistant S. haemolyticus (MRSH) isolates identified in blood cultures at the same ward were found to be genotypically homogenous by pulsed-field gel electrophoresis, and did not carry a type I, II, III or IV SCCmec either. Thus, the hypothesis was raised that an SCCmec of MRSH had been transferred to a methicillin-susceptible S. aureus strain and thereby created a new clone of MRSA that caused the outbreak. This study showed that MRSA from the outbreak carried a ccrC and a class C mec complex that was also found among MRSH isolates. Partial sequencing of the mec complexes showed more than 99% homology, indicative of a common type V SCCmec. This finding may provide evidence for a recent horizontal transfer of an SCCmec from MRSH to an identified potential recipient, an ST45 methicillin-susceptible S. aureus strain, thereby creating a new clone of MRSA that caused the outbreak.
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USDA-ARS?s Scientific Manuscript database
Zinc resistance in livestock-associated methicillin resistant Staphylococcus aureus (LA-MRSA) is mediated by the czrC gene co-located with the mecA gene, encoding methicillin resistance, on the type V SCCmec element. Since the czrC gene and the mecA gene are co-located on the SCCmec element, it has ...
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Boyle-Vavra, Susan; Ereshefsky, Ben; Wang, Chih-Chien; Daum, Robert S.
2005-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) isolates carry the methicillin resistance gene (mecA) on a horizontally transferred genetic element called the staphylococcal chromosome cassette mec (SCCmec). Community-acquired MRSA (CAMRSA) isolates usually carry SCCmec type IV. We previously reported that 76% of 17 CAMRSA isolates (multilocus sequence type 59) obtained from pediatric patients with skin and soft tissue infections (SSTI) from Taipei did not carry SCCmec types I to IV. We used DNA sequence analysis to determine that the element harbored by these nontypeable isolates is a novel subtype of SCCmec V called SCCmec VT. It contains a ccrC recombinase gene variant (ccrC2) and mec complex C2. One SSTI isolate contained molecular features of SCCmec IV but also contained ccrC2 (a feature of SCCmec VT), suggesting that it may harbor a composite SCCmec element. The genes lukS-PV and lukF-PV encoding the Panton-Valentine leukocidin (PVL) were present in all CAMRSA SSTI isolates whether they contained SCCmec type IV or VT. SCCmec VT was also present in 5 of 34 (14.7%) CAMRSA colonization isolates collected from healthy children from Taipei who lacked MRSA risk factors. Four (80%) of the these isolates contained lukS-PV and lukF-PV, as did 1 of 27 (3.7%) SCCmec IV-containing colonization isolates. A total of 63% (10 of 16) of the SSTI isolates and 61.7% (21 of 34) of the colonization isolates tested were resistant to at least four classes of non-β-lactam antimicrobials. SCCmec VT is a novel SCCmec variant that is found in multiply resistant CAMRSA strains with sequence type 59 in Taipei in association with the PVL leukotoxin genes. PMID:16145133
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Bhatt, Puneet; Tandel, Kundan; Singh, Alina; Kumar, M; Grover, Naveen; Sahni, A K
2016-12-01
Methicillin-resistant Coagulase-negative Staphylococci (MR-CoNS) have emerged as an important cause of nosocomial infections especially in patients with prosthetic devices and implants. This study was conducted with an aim to determine the prevalence of methicillin resistance among CoNS isolates at a tertiary care center by both phenotypic and genotypic methods. This cross sectional study was carried out from September 2011 to February 2014 in which 150 non-repetitive clinical isolates of CoNS were identified at the species level by conventional phenotypic methods. Cefoxitin disk (30 μg) diffusion testing was used to determine methicillin resistance and confirmed by detection of mec A gene by polymerase chain reaction (PCR). Out of 150 CoNS isolates, 51 were methicillin resistant by cefoxitin disk diffusion method. Out of these 51 isolates, mec A gene was detected only in 45 isolates. Moreover, mec A gene was also detected in 4 isolates, which were cefoxitin sensitive. Thus, the prevalence of methicillin resistance among CoNS was found to be 32.7% by PCR. The prevalence of methicillin resistance among Coagulase-negative Staphylococci (CoNS) was 32.7% by PCR detection of mec A gene. The sensitivity and specificity of cefoxitin disk diffusion method against mec A gene detection by PCR were found to be more than 90%. It can be concluded from this study that cefoxitin disk diffusion test can be used as a useful screening method to detect methicillin resistance among CoNS isolates. However, detection of mec A gene by PCR remains a more accurate method of detecting methicillin resistance among CoNS.
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Gong, Zongrong; Shu, Min; Xia, Qing; Tan, Shan; Zhou, Wei; Zhu, Yu; Wan, Chaomin
2017-06-01
To describe the epidemiological profile of nasal carriage of Staphylococcus aureus (S. aureus) strains, its antibiotic resistance and mecA and Panton Valentine leukocidin (PVL) genes presence, in school children residing in high altitude areas of Southwestern China. The cross sectional study screened nasal swabs taken from students for S.aureus. PCR was performed to identify mecA and PVL genes. Of the total 314 children 5.10% (16/314) was detected S.aureus. The resistance of isolated strains to penicillin, erythromycin, clindamycin, rifampicin and cefoxitin was 100%, 81.3%, 81.3%, 0.0%, and 6.3% respectively. No strains demonstrated resistance to vancomycin; expression of mecA gene was detected in 3 isolates and 10 isolates were PVL-positive. S. aureus was detected in 5.10% (16/314) of the study population; 0.96% (3/314) had methicillin resistant S.aureus (MRSA); expression of the mecA and PVL genes were detected in 3 and 10 isolates respectively.
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Bouchami, O; Ben Hassen, A; de Lencastre, H; Miragaia, M
2012-04-01
Staphylococcus haemolyticus is one of the most clinically relevant coagulase-negative staphylococci (CoNS), particularly in immunocompromised patients; however, little is known regarding its molecular epidemiology. In this work, we characterized the genetic background and the SCCmec region of 36 methicillin-resistant S. haemolyticus (MRSHae) and 10 methicillin-susceptible S. haemolyticus (MSSHae) collected from neutropenic patients in Tunisia between 2002 and 2004. The molecular characterization of MRSHae by pulsed-field gel electrophoresis (PFGE) showed that the great majority of the isolates (77.8%) belonged to only four types. SCCmec typing by polymerase chain reaction (PCR) and Southern hybridization showed that isolates belonging to each PFGE type could carry either one or two SCCmec types. SCCmec V was the most common, but mec complex C was frequently associated to ccr allotypes other than ccrC. The mec complex class C was predominant in MRSHae (47%) and ccrC was predominant among both methicillin-resistant and -susceptible isolates (31 and 50%, respectively). Interestingly, one half (50%) of the MRSHae isolates analyzed lacked the known ccr complexes (ccrAB and ccrC), although they carried the mecA. Conversely, all MSSHae carrying a ccrC complex were multidrug-resistant, although they lack the mecA. The results suggest that ccrC and mec complex C are frequent and may exist autonomously and independently of SCCmec type V in S. haemolyticus. Moreover, the data obtained suggest that small chromosomal rearrangements promoting the loss or structural variation of mec and ccr complex appear to occur frequently, which probably provide S. haemolyticus with a specialized means for SCCmec trapping and/or diversification.
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Zhang, Kunyan; McClure, Jo-Ann; Elsayed, Sameer; Louie, Thomas; Conly, John M.
2005-01-01
Staphylococcal cassette chromosome mec (SCCmec) typing is essential for understanding the molecular epidemiology of methicillin-resistant Staphylococcus aureus (MRSA). SCCmec elements are currently classified into types I to V based on the nature of the mec and ccr gene complexes, and are further classified into subtypes according to their junkyard region DNA segments. Previously described traditional SCCmec PCR typing schemes require multiple primer sets and PCR experiments, while a previously published multiplex PCR assay is limited in its ability to detect recently discovered types and subtypes such as SCCmec type V and subtypes IVa, b, c, and d. We designed new sets of SCCmec type- and subtype-unique and specific primers and developed a novel multiplex PCR assay allowing for concomitant detection of the methicillin resistance (mecA gene) (also serving as an internal control) to facilitate detection and classification of all currently described SCCmec types and subtypes I, II, III, IVa, b, c, d, and V. Our assay demonstrated 100% sensitivity and specificity in accurately characterizing 54 MRSA strains belonging to the various known SCCmec types and subtypes, when compared with previously described typing methods. Further application of our assay in 453 randomly selected local clinical isolates confirmed its feasibility and practicality. This novel assay offers a rapid, simple, and feasible method for SCCmec typing of MRSA, and may serve as a useful tool for clinicians and epidemiologists in their efforts to prevent and control infections caused by this organism. PMID:16207957
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Cheung, Gordon Y C; Villaruz, Amer E; Joo, Hwang-Soo; Duong, Anthony C; Yeh, Anthony J; Nguyen, Thuan H; Sturdevant, Daniel E; Queck, S Y; Otto, M
2014-07-01
Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM)-mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance. Published by Elsevier GmbH.
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Kapila, Neha; Sharma, Ankita; Kishore, Amit; Sodhi, Monika; Tripathi, Pawan K.; Mohanty, Ashok K.
2016-01-01
The present study aims to identify the heat responsive genes and biological pathways in heat stressed buffalo mammary epithelial cells (MECs). The primary mammary epithelial cells of riverine buffalo were exposed to thermal stress at 42°C for one hour. The cells were subsequently allowed to recover at 37°C and harvested at different time intervals (30 min to 48 h) along with control samples (un-stressed). In order to assess the impact of heat stress in buffalo MECs, several in-vitro cellular parameters (lactate dehydrogenase activity, cell proliferation assay, cellular viability, cell death and apoptosis) and transcriptional studies were conducted. The heat stress resulted in overall decrease in cell viability and cell proliferation of MECs while induction of cellular apoptosis and necrosis. The transcriptomic profile of heat stressed MECs was generated using Agilent 44 K bovine oligonucleotide array and at cutoff criteria of ≥3-or ≤3 fold change, a total of 153 genes were observed to be upregulated while 8 genes were down regulated across all time points post heat stress. The genes that were specifically up-regulated or down-regulated were identified as heat responsive genes. The upregulated genes in heat stressed MECs belonged to heat shock family viz., HSPA6, HSPB8, DNAJB2, HSPA1A. Along with HSPs, genes like BOLA, MRPL55, PFKFB3, PSMC2, ENDODD1, ARID5A, and SENP3 were also upregulated. Microarray data revealed that the heat responsive genes belonged to different functional classes viz., chaperons; immune responsive; cell proliferation and metabolism related. Gene ontology analysis revealed enrichment of several biological processes like; cellular process, metabolic process, response to stimulus, biological regulation, immune system processes and signaling. The transcriptome analysis data was further validated by RT-qPCR studies. Several HSP (HSP40, HSP60, HSP70, HSP90, and HSPB1), apoptotic (Bax and Bcl2), immune (IL6, TNFα and NF-kβ) and oxidative stress (GPX1 and DUSP1) related genes showed differential expression profile at different time points post heat stress. The transcriptional data strongly indicated the induction of survival/apoptotic mechanism in heat stressed buffalo MECs. The overrepresented pathways across all time points were; electron transport chain, cytochrome P450, apoptosis, MAPK, FAS and stress induction of HSP regulation, delta Notch signaling, apoptosis modulation by HSP70, EGFR1 signaling, cytokines and inflammatory response, oxidative stress, TNF-alpha and NF- kB signaling pathway. The study thus identified several genes from different functional classes and biological pathways that could be termed as heat responsive in buffalo MEC. The responsiveness of buffalo MECs to heat stress in the present study clearly suggested its suitability as a model to understand the modulation of buffalo mammary gland expression signature in response to environmental heat load. PMID:27682256
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The Andes Virus Nucleocapsid Protein Directs Basal Endothelial Cell Permeability by Activating RhoA
Gorbunova, Elena E.; Simons, Matthew J.; Gavrilovskaya, Irina N.
2016-01-01
ABSTRACT Andes virus (ANDV) predominantly infects microvascular endothelial cells (MECs) and nonlytically causes an acute pulmonary edema termed hantavirus pulmonary syndrome (HPS). In HPS patients, virtually every pulmonary MEC is infected, MECs are enlarged, and infection results in vascular leakage and highly lethal pulmonary edema. We observed that MECs infected with the ANDV hantavirus or expressing the ANDV nucleocapsid (N) protein showed increased size and permeability by activating the Rheb and RhoA GTPases. Expression of ANDV N in MECs increased cell size by preventing tuberous sclerosis complex (TSC) repression of Rheb-mTOR-pS6K. N selectively bound the TSC2 N terminus (1 to 1403) within a complex containing TSC2/TSC1/TBC1D7, and endogenous TSC2 reciprocally coprecipitated N protein from ANDV-infected MECs. TSCs normally restrict RhoA-induced MEC permeability, and we found that ANDV infection or N protein expression constitutively activated RhoA. This suggests that the ANDV N protein alone is sufficient to activate signaling pathways that control MEC size and permeability. Further, RhoA small interfering RNA, dominant-negative RhoA(N19), and the RhoA/Rho kinase inhibitors fasudil and Y27632 dramatically reduced the permeability of ANDV-infected MECs by 80 to 90%. Fasudil also reduced the bradykinin-directed permeability of ANDV and Hantaan virus-infected MECs to control levels. These findings demonstrate that ANDV activation of RhoA causes MEC permeability and reveal a potential edemagenic mechanism for ANDV to constitutively inhibit the basal barrier integrity of infected MECs. The central importance of RhoA activation in MEC permeability further suggests therapeutically targeting RhoA, TSCs, and Rac1 as potential means of resolving capillary leakage during hantavirus infections. PMID:27795403
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Aedo, Sandra
2016-01-01
Resistance to beta-lactam antibiotics in methicillin-resistant Staphylococcus aureus (MRSA) requires the presence of an acquired genetic determinant, mecA or mecC, which encode penicillin-binding protein PBP2A or PBP2A′, respectively. Although all MRSA strains share a mechanism of resistance, the phenotypic expression of beta-lactam resistance shows considerable strain-to-strain variation. The stringent stress response, a stress response that results from nutrient limitation, was shown to play a key role in determining the resistance level of an MRSA strain. In the present study, we validated the impact of the stringent stress response on transcription and translation of mecA in the MRSA clinical isolate strain N315, which also carries known regulatory genes (mecI/mecR1/mecR2 and blaI/blaR1) for mecA transcription. We showed that the impact of the stringent stress response on the resistance level may be restricted to beta-lactam resistance based on a “foreign” determinant such as mecA, as opposed to resistance based on mutations in the native S. aureus determinant pbpB (encoding PBP2). Our observations demonstrate that high-level resistance mediated by the stringent stress response follows the current model of beta-lactam resistance in which the native PBP2 protein is also essential for expression of the resistance phenotype. We also show that the Staphylococcus sciuri pbpD gene (also called mecAI), the putative evolutionary precursor of mecA, confers oxacillin resistance in an S. aureus strain, generating a heterogeneous phenotype that can be converted to high and homogenous resistance by induction of the stringent stress response in the bacteria. PMID:26833147
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NASA Astrophysics Data System (ADS)
Zulkifli, Aisya; Ahmad, Asmat
2015-09-01
Water samples were collected in triplicates from three different locations choosen from the recreational beach of Teluk Kemang, Port Dickson as sampling station including main area of recreation activity for the public. Bacteria were isolated from the water and cultured. Out of 286 presumptive Staphylococcus aureus enumerated by using culture method, only 4 (1.4 %) confirmed as Meticillin Resistant S. aureus (MRSA) based on PCR detection of mecA gene. Interestingly, all of MRSA detections were found at the main area of recreational activity. Our results suggested that public beaches may be reservoir for transmission of MRSA to beach visitors and PCR using the mecA gene is the fastest way to detect this pathogenic bacteria.
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Saito, Yuki; Mao, Han; Sekimizu, Kazuhisa; Kaito, Chikara
2014-01-01
Staphylococcal species acquire antibiotic resistance by incorporating the mobile-genetic element SCCmec. We previously found that SCCmec-encoded psm-mec RNA suppresses exotoxin production as a regulatory RNA, and the psm-mec translation product increases biofilm formation in Staphylococcus aureus. Here, we examined whether the regulatory role of psm-mec on host bacterial virulence properties is conserved among other staphylococcal species, S. epidermidis and S. haemolyticus, both of which are important causes of nosocomial infections. In S. epidermidis, introduction of psm-mec decreased the production of cytolytic toxins called phenol-soluble modulins (PSMs) and increased biofilm formation. Introduction of psm-mec with a stop-codon mutation that did not express PSM-mec protein but did express psm-mec RNA also decreased PSM production, but did not increase biofilm formation. Thus, the psm-mec RNA inhibits PSM production, whereas the PSM-mec protein increases biofilm formation in S. epidermidis. In S. haemolyticus, introduction of psm-mec decreased PSM production, but did not affect biofilm formation. The mutated psm-mec with a stop-codon also caused the same effect. Thus, the psm-mec RNA also inhibits PSM production in S. haemolyticus. These findings suggest that the inhibitory role of psm-mec RNA on exotoxin production is conserved among staphylococcal species, although the stimulating effect of the psm-mec gene on biofilm formation is not conserved. PMID:24926994
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Rodo, Christophe; Sargolini, Francesca; Save, Etienne
2017-03-01
The entorhinal-hippocampal circuitry has been suggested to play an important role in episodic memory but the contribution of the entorhinal cortex remains elusive. Predominant theories propose that the medial entorhinal cortex (MEC) processes spatial information whereas the lateral entorhinal cortex (LEC) processes non spatial information. A recent study using an object exploration task has suggested that the involvement of the MEC and LEC spatial and non-spatial information processing could be modulated by the amount of information to be processed, i.e. environmental complexity. To address this hypothesis we used an object exploration task in which rats with excitotoxic lesions of the MEC and LEC had to detect spatial and non-spatial novelty among a set of objects and we varied environmental complexity by decreasing the number of objects or amount of object diversity. Reducing diversity resulted in restored ability to process spatial and non-spatial information in MEC and LEC groups, respectively. Reducing the number of objects yielded restored ability to process non-spatial information in the LEC group but not the ability to process spatial information in the MEC group. The findings indicate that the MEC and LEC are not strictly necessary for spatial and non-spatial processing but that their involvement depends on the complexity of the information to be processed. Copyright © 2016 Elsevier B.V. All rights reserved.
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Ba, Xiaoliang; Lovering, Andrew L.; Gleadall, Nicholas; Zadoks, Ruth; Peacock, Sharon J.; Holden, Matthew T. G.; Paterson, Gavin K.; Holmes, Mark A.
2015-01-01
β-Lactam resistance in methicillin-resistant Staphylococcus aureus (MRSA) is mediated by the expression of an alternative penicillin-binding protein 2a (PBP2a) (encoded by mecA) with a low affinity for β-lactam antibiotics. Recently, a novel variant of mecA, known as mecC, was identified in MRSA isolates from both humans and animals. In this study, we demonstrate that mecC-encoded PBP2c does not mediate resistance to penicillin. Rather, broad-spectrum β-lactam resistance in MRSA strains carrying mecC (mecC-MRSA strains) is mediated by a combination of both PBP2c and the distinct β-lactamase encoded by the blaZ gene of strain LGA251 (blaZLGA251), which is part of mecC-encoding staphylococcal cassette chromosome mec (SCCmec) type XI. We further demonstrate that mecC-MRSA strains are susceptible to the combination of penicillin and the β-lactam inhibitor clavulanic acid in vitro and that the same combination is effective in vivo for the treatment of experimental mecC-MRSA infection in wax moth larvae. Thus, we demonstrate how the distinct biological differences between mecA- and mecC-encoded PBP2a and PBP2c have the potential to be exploited as a novel approach for the treatment of mecC-MRSA infections. PMID:26392513
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Bele, Aditya; Mirza, Sameer; Zhang, Ying; Ahmad Mir, Riyaz; Lin, Simon; Kim, Jun Hyun; Gurumurthy, Channabasavaiah Basavaraju; West, William; Qiu, Fang; Band, Hamid; Band, Vimla
2015-01-01
The mammalian ortholog of Drosophila ecdysoneless (Ecd) gene product regulates Rb-E2F interaction and is required for cell cycle progression. Ecd is overexpressed in breast cancer and its overexpression predicts shorter survival in patients with ErbB2-positive tumors. Here, we demonstrate Ecd knock down (KD) in human mammary epithelial cells (hMECs) induces growth arrest, similar to the impact of Ecd Knock out (KO) in mouse embryonic fibroblasts. Furthermore, whole-genome mRNA expression analysis of control vs. Ecd KD in hMECs demonstrated that several of the top 40 genes that were down-regulated were E2F target genes. To address the role of Ecd in mammary oncogenesis, we overexpressed Ecd and/or mutant H-Ras in hTERT-immortalized hMECs. Cell cycle analyses revealed hMECs overexpressing Ecd+Ras showed incomplete arrest in G1 phase upon growth factor deprivation, and more rapid cell cycle progression in growth factor-containing medium. Analyses of cell migration, invasion, acinar structures in 3-D Matrigel and anchorage-independent growth demonstrated that Ecd+Ras-overexpressing cells exhibit substantially more dramatic transformed phenotype as compared to cells expressing vector, Ras or Ecd. Under conditions of nutrient deprivation, Ecd+Ras-overexpressing hMECs exhibited better survival, with substantial upregulation of the autophagy marker LC3 both at the mRNA and protein levels. Significantly, while hMECs expressing Ecd or mutant Ras alone did not form tumors in NOD/SCID mice, Ecd+Ras-overexpressing hMECs formed tumors, clearly demonstrating oncogenic cooperation between Ecd and mutant Ras. Collectively, we demonstrate an important co-oncogenic role of Ecd in the progression of mammary oncogenesis through promoting cell survival.
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Bele, Aditya; Mirza, Sameer; Zhang, Ying; Ahmad Mir, Riyaz; Lin, Simon; Kim, Jun Hyun; Gurumurthy, Channabasavaiah Basavaraju; West, William; Qiu, Fang; Band, Hamid; Band, Vimla
2015-01-01
The mammalian ortholog of Drosophila ecdysoneless (Ecd) gene product regulates Rb-E2F interaction and is required for cell cycle progression. Ecd is overexpressed in breast cancer and its overexpression predicts shorter survival in patients with ErbB2-positive tumors. Here, we demonstrate Ecd knock down (KD) in human mammary epithelial cells (hMECs) induces growth arrest, similar to the impact of Ecd Knock out (KO) in mouse embryonic fibroblasts. Furthermore, whole-genome mRNA expression analysis of control vs. Ecd KD in hMECs demonstrated that several of the top 40 genes that were down-regulated were E2F target genes. To address the role of Ecd in mammary oncogenesis, we overexpressed Ecd and/or mutant H-Ras in hTERT-immortalized hMECs. Cell cycle analyses revealed hMECs overexpressing Ecd+Ras showed incomplete arrest in G1 phase upon growth factor deprivation, and more rapid cell cycle progression in growth factor-containing medium. Analyses of cell migration, invasion, acinar structures in 3-D Matrigel and anchorage-independent growth demonstrated that Ecd+Ras-overexpressing cells exhibit substantially more dramatic transformed phenotype as compared to cells expressing vector, Ras or Ecd. Under conditions of nutrient deprivation, Ecd+Ras-overexpressing hMECs exhibited better survival, with substantial upregulation of the autophagy marker LC3 both at the mRNA and protein levels. Significantly, while hMECs expressing Ecd or mutant Ras alone did not form tumors in NOD/SCID mice, Ecd+Ras-overexpressing hMECs formed tumors, clearly demonstrating oncogenic cooperation between Ecd and mutant Ras. Collectively, we demonstrate an important co-oncogenic role of Ecd in the progression of mammary oncogenesis through promoting cell survival. PMID:25616580
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Frey, Yvonne; Rodriguez, Joan Peña; Thomann, Andreas; Schwendener, Sybille; Perreten, Vincent
2013-04-01
Coagulase-negative staphylococci (CNS; n=417) were isolated from bovine milk and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Nineteen different species were identified, and Staphylococcus xylosus, Staphylococcus chromogenes, Staphylococcus haemolyticus, and Staphylococcus sciuri were the most prevalent species. Resistance to oxacillin (47.0% of the isolates), fusidic acid (33.8%), tiamulin (31.9%), penicillin (23.3%), tetracycline (15.8%), streptomycin (9.6%), erythromycin (7.0%), sulfonamides (5%), trimethoprim (4.3%), clindamycin (3.4%), kanamycin (2.4%), and gentamicin (2.4%) was detected. Resistance to oxacillin was attributed to the mecA gene in 9.7% of the oxacillin-resistant isolates. The remaining oxacillin-resistant CNS did not contain the mecC gene or mecA1 promoter mutations. The mecA gene was detected in Staphylococcus fleurettii, Staphylococcus epidermidis, Staph. haemolyticus, and Staph. xylosus. Resistance to tetracycline was attributed to the presence of tet(K) and tet(L), penicillin resistance to blaZ, streptomycin resistance to str and ant(6)-Ia, and erythromycin resistance to erm(C), erm(B), and msr. Resistance to tiamulin and fusidic acid could not be attributed to an acquired resistance gene. In total, 15.1% of the CNS isolates were multidrug resistant (i.e., resistant to 2 or more antimicrobials). The remaining CNS isolates were susceptible to antimicrobials commonly used in mastitis treatment. Methicillin-resistant CNS isolates were diverse, as determined by mecA gene sequence analysis, staphylococcal cassette chromosome mec typing, and pulsed-field gel electrophoresis. Arginine catabolic mobile element types 1 and 3 were detected in both methicillin-resistant and methicillin-susceptible Staph. epidermidis and were associated with sequence types ST59 and ST111. Because this study revealed the presence of multidrug-resistant CNS in a heterogeneous CNS population, we recommend antibiogram analysis of CNS in persistent infections before treatment with antimicrobials. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
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Kumar, V Anil; Steffy, Katherin; Chatterjee, Maitrayee; Sugumar, Madhan; Dinesh, Kavitha R; Manoharan, Anand; Karim, Shamsul; Biswas, Raja
2013-01-01
Reports of oxacillin-susceptible mecA-positive Staphylococcus aureus strains are on the rise. Because of their susceptibility to oxacillin and cefoxitin, it is very difficult to detect them by using routine phenotypic methods. We describe two such isolates that were detected by chromogenic medium and confirmed by characterization of the mecA gene element.
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Mayer, Boris F; Ali-Benali, Mohamed Ali; Demone, Jordan; Bertrand, Annick; Charron, Jean-Benoit
2015-11-01
Little is known about the capacity of Cannabis sativa to cold-acclimate and develop freezing tolerance. This study investigates the cold acclimation (CA) capacity of nine C. sativa varieties and the underlying genetic and epigenetic responses. The varieties were divided into three groups based on their contrasting CA capacities by comparing the survival of non-acclimated and cold-acclimated plants in whole-plant freeze tests. In response to the CA treatment, all varieties accumulated soluble sugars but only the varieties with superior capacity for CA could maintain higher levels throughout the treatment. In addition, the varieties that acclimated most efficiently accumulated higher transcript levels of cold-regulated (COR) genes and genes involved in de novo DNA methylation while displaying locus- and variety-specific changes in the levels of H3K9ac, H3K27me3 and methylcytosine (MeC) during CA. Furthermore, these hardy C. sativa varieties displayed significant increases in MeC levels at COR gene loci when deacclimated, suggesting a role for locus-specific DNA methylation in deacclimation. This study uncovers the molecular mechanisms underlying CA in C. sativa and reveals higher levels of complexity regarding how genetic, epigenetic and environmental factors intertwine. © 2014 Scandinavian Plant Physiology Society.
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NASA Astrophysics Data System (ADS)
Azwar; Hussain, M. A.; Abdul-Wahab, A. K.; Zanil, M. F.; Mukhlishien
2018-03-01
One of major challenge in bio-hydrogen production process by using MEC process is nonlinear and highly complex system. This is mainly due to the presence of microbial interactions and highly complex phenomena in the system. Its complexity makes MEC system difficult to operate and control under optimal conditions. Thus, precise control is required for the MEC reactor, so that the amount of current required to produce hydrogen gas can be controlled according to the composition of the substrate in the reactor. In this work, two schemes for controlling the current and voltage of MEC were evaluated. The controllers evaluated are PID and Inverse neural network (NN) controller. The comparative study has been carried out under optimal condition for the production of bio-hydrogen gas wherein the controller output is based on the correlation of optimal current and voltage to the MEC. Various simulation tests involving multiple set-point changes and disturbances rejection have been evaluated and the performances of both controllers are discussed. The neural network-based controller results in fast response time and less overshoots while the offset effects are minimal. In conclusion, the Inverse neural network (NN)-based controllers provide better control performance for the MEC system compared to the PID controller.
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Erdelyi, Peter; Wang, Xing; Suleski, Marina; Wicky, Chantal
2016-01-01
Mi2 proteins are evolutionarily conserved, ATP-dependent chromatin remodelers of the CHD family that play key roles in stem cell differentiation and reprogramming. In Caenorhabditis elegans, the let-418 gene encodes one of the two Mi2 homologs, which is part of at least two chromatin complexes, namely the Nucleosome Remodeling and histone Deacetylase (NuRD) complex and the MEC complex, and functions in larval development, vulval morphogenesis, lifespan regulation, and cell fate determination. To explore the mechanisms involved in the action of LET-418/Mi2, we performed a genome-wide RNA interference (RNAi) screen for suppressors of early larval arrest associated with let-418 mutations. We identified 29 suppressor genes, of which 24 encode chromatin regulators, mostly orthologs of proteins present in transcriptional activator complexes. The remaining five genes vary broadly in their predicted functions. All suppressor genes could suppress multiple aspects of the let-418 phenotype, including developmental arrest and ectopic expression of germline genes in the soma. Analysis of available transcriptomic data and quantitative PCR revealed that LET-418 and the suppressors of early larval arrest are regulating common target genes. These suppressors might represent direct competitors of LET-418 complexes for chromatin regulation of crucial genes involved in the transition to postembryonic development. PMID:28007841
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Erdelyi, Peter; Wang, Xing; Suleski, Marina; Wicky, Chantal
2017-02-09
Mi2 proteins are evolutionarily conserved, ATP-dependent chromatin remodelers of the CHD family that play key roles in stem cell differentiation and reprogramming. In Caenorhabditis elegans , the let-418 gene encodes one of the two Mi2 homologs, which is part of at least two chromatin complexes, namely the Nucleosome Remodeling and histone Deacetylase (NuRD) complex and the MEC complex, and functions in larval development, vulval morphogenesis, lifespan regulation, and cell fate determination. To explore the mechanisms involved in the action of LET-418/Mi2, we performed a genome-wide RNA interference (RNAi) screen for suppressors of early larval arrest associated with let-418 mutations. We identified 29 suppressor genes, of which 24 encode chromatin regulators, mostly orthologs of proteins present in transcriptional activator complexes. The remaining five genes vary broadly in their predicted functions. All suppressor genes could suppress multiple aspects of the let-418 phenotype, including developmental arrest and ectopic expression of germline genes in the soma. Analysis of available transcriptomic data and quantitative PCR revealed that LET-418 and the suppressors of early larval arrest are regulating common target genes. These suppressors might represent direct competitors of LET-418 complexes for chromatin regulation of crucial genes involved in the transition to postembryonic development. Copyright © 2017 Erdelyi et al.
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Assembly of Slx4 signaling complexes behind DNA replication forks.
Balint, Attila; Kim, TaeHyung; Gallo, David; Cussiol, Jose Renato; Bastos de Oliveira, Francisco M; Yimit, Askar; Ou, Jiongwen; Nakato, Ryuichiro; Gurevich, Alexey; Shirahige, Katsuhiko; Smolka, Marcus B; Zhang, Zhaolei; Brown, Grant W
2015-08-13
Obstructions to replication fork progression, referred to collectively as DNA replication stress, challenge genome stability. In Saccharomyces cerevisiae, cells lacking RTT107 or SLX4 show genome instability and sensitivity to DNA replication stress and are defective in the completion of DNA replication during recovery from replication stress. We demonstrate that Slx4 is recruited to chromatin behind stressed replication forks, in a region that is spatially distinct from that occupied by the replication machinery. Slx4 complex formation is nucleated by Mec1 phosphorylation of histone H2A, which is recognized by the constitutive Slx4 binding partner Rtt107. Slx4 is essential for recruiting the Mec1 activator Dpb11 behind stressed replication forks, and Slx4 complexes are important for full activity of Mec1. We propose that Slx4 complexes promote robust checkpoint signaling by Mec1 by stably recruiting Dpb11 within a discrete domain behind the replication fork, during DNA replication stress. © 2015 The Authors.
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Bell, Diana; Holsinger, Christopher F; El-Naggar, Adel K
2010-12-01
Intraosseous salivary gland carcinomas are extremely rare, comprising only 2% to 3% of all mucoepidermoid carcinomas (MECs) reported. The t(11;19) translocation and its CRTC1/MAML1 fusion transcript have been identified in MEC at different sites and are believed to be associated with the development of a subset of these tumors. However, the status of the fusion transcript has not been reported in intraosseous MEC. Here, we report 3 examples of central MEC of the mandible, including a case with a history of primary retromolar MEC. Reverse transcriptase-polymerase chain reaction and DNA sequencing analyses of the microdissected components of these tumors were used for the detection and verification of the fusion transcript. We identified, for the first time, the t(11;19) fusion gene transcript in central MEC, including in the previous primary retromolar MEC. No fusion transcript was detected in the second primary noncentral MEC or in another central MEC. The results indicate that central MEC can manifest the fusion transcript. This finding may have diagnostic and histogenetic roles in the future analysis of this entity. Published by Elsevier Inc.
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Barros, E M; Ceotto, H; Bastos, M C F; Dos Santos, K R N; Giambiagi-Demarval, M
2012-01-01
Phenotypic and molecular methods were used to characterize the antibiotic resistance of 64 clinical isolates of Staphylococcus haemolyticus. By PCR of the mecA gene, 87% were found to be methicillin resistant. Approximately 55% harbored staphylococcal cassette chromosome mec element (SCCmec) type V, and only one SCCmec type IV. Many isolates (75%) displayed multiresistance, and pulsotype analysis showed a high diversity.
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Ba, Xiaoliang; Harrison, Ewan M; Lovering, Andrew L; Gleadall, Nicholas; Zadoks, Ruth; Parkhill, Julian; Peacock, Sharon J; Holden, Matthew T G; Paterson, Gavin K; Holmes, Mark A
2015-12-01
β-Lactam resistance in methicillin-resistant Staphylococcus aureus (MRSA) is mediated by the expression of an alternative penicillin-binding protein 2a (PBP2a) (encoded by mecA) with a low affinity for β-lactam antibiotics. Recently, a novel variant of mecA, known as mecC, was identified in MRSA isolates from both humans and animals. In this study, we demonstrate that mecC-encoded PBP2c does not mediate resistance to penicillin. Rather, broad-spectrum β-lactam resistance in MRSA strains carrying mecC (mecC-MRSA strains) is mediated by a combination of both PBP2c and the distinct β-lactamase encoded by the blaZ gene of strain LGA251 (blaZLGA251), which is part of mecC-encoding staphylococcal cassette chromosome mec (SCCmec) type XI. We further demonstrate that mecC-MRSA strains are susceptible to the combination of penicillin and the β-lactam inhibitor clavulanic acid in vitro and that the same combination is effective in vivo for the treatment of experimental mecC-MRSA infection in wax moth larvae. Thus, we demonstrate how the distinct biological differences between mecA- and mecC-encoded PBP2a and PBP2c have the potential to be exploited as a novel approach for the treatment of mecC-MRSA infections. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
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Gómez, Paula; Lozano, Carmen; González-Barrio, David; Zarazaga, Myriam; Ruiz-Fons, Francisco; Torres, Carmen
2015-06-12
The objective was to determine the prevalence of Staphylococcus aureus nasal carriage in red deer of a semi-extensive farm and in humans in contact with the estate animals, and to characterize obtained isolates. Nasal swabs of 65 deer and 15 humans were seeded on mannitol-salt-agar and oxacillin-resistance-screening-agar-base. Isolates were identified by microbiological and molecular methods. Antimicrobial susceptibility profile was determined for 16 antibiotics by disk-diffusion and the presence of eight antibiotic resistance genes, seven virulence genes and genes of immune-evasion-cluster (IEC) was analyzed by PCR. S. aureus was typed by PFGE-SmaI, spa, agr, SCCmec and MLST. Isolates were detected in 16 deer (24.6%). Eleven S. aureus isolates were methicillin-resistant (MRSA), and five were methicillin-susceptible (MSSA). All MRSA harbored mecC gene and were agr-III/SCCmecXI/ST1945 (four spa-t843 and seven spa-t1535). All mecC-MRSA carried blaZ-SCCmecXI and etd2, were IEC-type-E, and belonged to the same PFGE pattern. The five MSSA were typed as spa-t2420/agr-I/ST133. Regarding humans, S. aureus was recovered from six samples (40%). The isolates were MSSA and were typed as spa-t002/agr-II, spa-t012/agr-III or spa-t822/agr-III and showed different IEC types (A, B, D and F). blaZ and erm(A) genes were detected, as well as cna and tst genes. As conclusion, red deer analyzed in this study are frequent carriers of mecC-MRSA CC130 (16.9%), they are characterized by few resistance and virulence determinants, and by the presence of IEC type-E. Deer could be a source of mecC-MRSA which could potentially be transmitted to other animals, or even to humans. Copyright © 2015 Elsevier B.V. All rights reserved.
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Ohshiro, Takeya; Miyagi, Chihiro; Tamaki, Yoshikazu; Mizuno, Takuya; Ezaki, Takayuki
2016-06-01
Blood culturing and the rapid reporting of results are essential for infectious disease clinics to obtain bacterial information that can affect patient prognosis. When gram-positive coccoid cells are observed in blood culture bottles, it is important to determine whether the strain is Staphylococcus aureus and whether the strain has resistance genes, such as mecA and blaZ, for proper antibiotic selection. Previous work led to the development of a PCR method that is useful for rapid identification of bacterial species and antimicrobial susceptibility. However, that method has not yet been adopted in community hospitals due to the high cost and methodological complexity. We report here the development of a quick PCR and DNA-chromatography test, based on single-tag hybridization chromatography, that permits detection of S. aureus and the mecA and blaZ genes; results can be obtained within 1 h for positive blood culture bottles. We evaluated this method using 42 clinical isolates. Detection of S. aureus and the resistance genes by the PCR-DNA-chromatography method was compared with that obtained via the conventional identification method and actual antimicrobial susceptibility testing. Our method had a sensitivity of 97.0% and a specificity of 100% for the identification of the bacterial species. For the detection of the mecA gene of S. aureus, the sensitivity was 100% and the specificity was 95.2%. For the detection of the blaZ gene of S. aureus, the sensitivity was 100% and the specificity was 88.9%. The speed and simplicity of this PCR-DNA-chromatography method suggest that our method will facilitate rapid diagnoses. Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.
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Barros, E. M.; Ceotto, H.; Bastos, M. C. F.; dos Santos, K. R. N.
2012-01-01
Phenotypic and molecular methods were used to characterize the antibiotic resistance of 64 clinical isolates of Staphylococcus haemolyticus. By PCR of the mecA gene, 87% were found to be methicillin resistant. Approximately 55% harbored staphylococcal cassette chromosome mec element (SCCmec) type V, and only one SCCmec type IV. Many isolates (75%) displayed multiresistance, and pulsotype analysis showed a high diversity. PMID:21976766
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Mobile genetic element-encoded cytolysin connects virulence to methicillin resistance in MRSA.
Queck, Shu Y; Khan, Burhan A; Wang, Rong; Bach, Thanh-Huy L; Kretschmer, Dorothee; Chen, Liang; Kreiswirth, Barry N; Peschel, Andreas; Deleo, Frank R; Otto, Michael
2009-07-01
Bacterial virulence and antibiotic resistance have a significant influence on disease severity and treatment options during bacterial infections. Frequently, the underlying genetic determinants are encoded on mobile genetic elements (MGEs). In the leading human pathogen Staphylococcus aureus, MGEs that contain antibiotic resistance genes commonly do not contain genes for virulence determinants. The phenol-soluble modulins (PSMs) are staphylococcal cytolytic toxins with a crucial role in immune evasion. While all known PSMs are core genome-encoded, we here describe a previously unidentified psm gene, psm-mec, within the staphylococcal methicillin resistance-encoding MGE SCCmec. PSM-mec was strongly expressed in many strains and showed the physico-chemical, pro-inflammatory, and cytolytic characteristics typical of PSMs. Notably, in an S. aureus strain with low production of core genome-encoded PSMs, expression of PSM-mec had a significant impact on immune evasion and disease. In addition to providing high-level resistance to methicillin, acquisition of SCCmec elements encoding PSM-mec by horizontal gene transfer may therefore contribute to staphylococcal virulence by substituting for the lack of expression of core genome-encoded PSMs. Thus, our study reveals a previously unknown role of methicillin resistance clusters in staphylococcal pathogenesis and shows that important virulence and antibiotic resistance determinants may be combined in staphylococcal MGEs.
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Magnetic Field Analysis of Lorentz Motors Using a Novel Segmented Magnetic Equivalent Circuit Method
Qian, Junbing; Chen, Xuedong; Chen, Han; Zeng, Lizhan; Li, Xiaoqing
2013-01-01
A simple and accurate method based on the magnetic equivalent circuit (MEC) model is proposed in this paper to predict magnetic flux density (MFD) distribution of the air-gap in a Lorentz motor (LM). In conventional MEC methods, the permanent magnet (PM) is treated as one common source and all branches of MEC are coupled together to become a MEC network. In our proposed method, every PM flux source is divided into three sub-sections (the outer, the middle and the inner). Thus, the MEC of LM is divided correspondingly into three independent sub-loops. As the size of the middle sub-MEC is small enough, it can be treated as an ideal MEC and solved accurately. Combining with decoupled analysis of outer and inner MECs, MFD distribution in the air-gap can be approximated by a quadratic curve, and the complex calculation of reluctances in MECs can be avoided. The segmented magnetic equivalent circuit (SMEC) method is used to analyze a LM, and its effectiveness is demonstrated by comparison with FEA, conventional MEC and experimental results. PMID:23358368
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Legrand, Melanie; Chan, Christine L.; Jauert, Peter A.; Kirkpatrick, David T.
2011-01-01
Genome rearrangements, a common feature of Candida albicans isolates, are often associated with the acquisition of antifungal drug resistance. In Saccharomyces cerevisiae, perturbations in the S-phase checkpoints result in the same sort of Gross Chromosomal Rearrangements (GCRs) observed in C. albicans. Several proteins are involved in the S. cerevisiae cell cycle checkpoints, including Mec1p, a protein kinase of the PIKK (phosphatidyl inositol 3-kinase-like kinase) family and the central player in the DNA damage checkpoint. Sgs1p, the ortholog of BLM, the Bloom’s syndrome gene, is a RecQ-related DNA helicase; cells from BLM patients are characterized by an increase in genome instability. Yeast strains bearing deletions in MEC1 or SGS1 are viable (in contrast to the inviability seen with loss of MEC1 in S. cerevisiae) but the different deletion mutants have significantly different phenotypes. The mec1Δ/Δ colonies have a wild-type colony morphology, while the sgs1Δ/Δ mutants are slow-growing, producing wrinkled colonies with pseudohyphal-like cells. The mec1Δ/Δ mutants are only sensitive to ethylmethane sulfonate (EMS), methylmethane sulfonate (MMS), and hydroxyurea (HU) but the sgs1Δ/Δ mutants exhibit a high sensitivity to all DNA-damaging agents tested. In an assay for chromosome 1 integrity, the mec1Δ/Δ mutants exhibit an increase in genome instability; no change was observed in the sgs1Δ/Δ mutants. Finally, loss of MEC1 does not affect sensitivity to the antifungal drug fluconazole, while loss of SGS1 leads to an increased susceptibility to fluconazole. Neither deletion elevated the level of antifungal drug resistance acquisition. PMID:21511048
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Antibiotic resistance of canine Staphylococcus intermedius group (SIG)--practical implications.
Chrobak, D; Kizerwetter-Swida, M; Rzewuska, M; Binek, M
2011-01-01
A total of 221 SIG strains were isolated from clinical samples of canine origin submitted to the Diagnostic Laboratory of the Division of Bacteriology and Molecular Biology at the Warsaw University of Life Sciences in Warsaw during the period 2006-2010. The aim of the study was to investigate the frequency of prevalence of methicillin-resistant SIG strains and to determine the MIC values of cephalotin, amoxicillin/clavulanic acid, ciprofloxacin, clindamycin, gentamicin, chloramphenicol, mupirocin for a collection of randomly selected 79 strains belonging to Staphylococcus intermedius group (SIG), including 23 mecA-positive and 56 mecA-negative strains. All isolates were identified as belonging to SIG based on their phenotypic properties and PCR amplification of S. intermedius-specific fragment of the 16S rRNA gene. The mecA gene was detected in 26 (12%) of 221 SIG strains. All tested mecA-negative SIG strains were susceptible to amoxicillin/clavulanic acid and cephalotin. One of the 56 mecA-negative SIG strains was resistant to ciprofloxacin, six (11%) to gentamicin. It was found that sixteen (29%) of 56 mecA-negative SIG strains were resistant to clindamycin. Most of the mecA-positive SIG strains were resistant to ciprofloxacin (96%), clindamycin (96%), and gentamicin (96%). Only one MRSIG strain was resistant to chloramphenicol. All examined mecA-positive SIG strains were found to be susceptible to mupirocin. Our results imply that staphylococcal multidrug resistance has become more prevalent, which could lead to difficulties in effective treatment. With some resistant strains the only therapeutic possibility are antimicrobial agents important in human medicine. New regulations for veterinary medicine concerning appropriate therapy of infections caused by multidrug-resistat staphylococci are needed.
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Saber, Huda; Jasni, Azmiza Syawani; Jamaluddin, Tengku Zetty Maztura Tengku; Ibrahim, Rosni
2017-10-01
Coagulase-negative staphylococci (CoNS) are considered low pathogenic organisms. However, they are progressively causing more serious infections with time because they have adapted well to various antibiotics owing to their ability to form biofilms. Few studies have been conducted on CoNS in both, hospital and community-acquired settings, especially in Malaysia. Thus, it is important to study their species and gene distributions. A mobile genetic element, staphylococcal cassette chromosome mec (SCC mec ), plays an important role in staphylococci pathogenesis. Among CoNS, SCC mec has been studied less frequently than Staphylococcus aureus (coagulase-positive staphylococci). A recent study (8) conducted in Malaysia successfully detected SCC mec type I to VIII as well as several new combination patterns in CoNS species, particularly Staphylococcus epidermidis . However, data are still limited, and further research is warranted. This paper provides a review on SCC mec types among CoNS species.
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Jena, Smrutiti; Panda, Sasmita; Nayak, Kinshuk C.; Singh, Durg V.
2017-01-01
We examined the presence of virulence and antibiotic resistance genes, SCCmec types and determined the genomic diversity among ocular S. epidermidis isolates (patients-23, healthy controls-29). PCR determined the presence of antibiotic resistance genes, virulence genes and SCCmec types among all isolates. MLST and PFGE determined the genomic relatedness among them. All isolates of S. epidermidis showed resistance to at least one class of antibiotics of which 48 isolates were multidrug resistant and carried ARGs. Thirty-five isolates were methicillin resistant and carried mecA gene. Majority of the isolates were resistant to fluoroquinolones and showed mutation in gyrA, parC, and parE genes, however, few isolates showed additional novel mutations in parC gene. Of the MRSE strains, 17 strains carried SCCmec type IV, four type V, two type II, and two UT4. Seven strains carried novel combination of ccr complex and SCCmercury element, not reported earlier. All the S. epidermidis strains harbored icaA and icaD genes, 47 carried ACME operon, and 50 contained IS256. A noteworthy finding was the presence of ST179 among 43% of infected eye isolates an observation rarely reported among S. epidermidis. PFGE and MLST analysis showed genomic diversity among them. Statistical analysis suggests that few healthy conjunctiva isolates had characteristics similar to infected eye isolates. S. epidermidis strains carrying mecA gene are multidrug resistant, virulent and diverse irrespective of sources of isolation. IS256 cannot be used as marker to differentiate isolates of infected eye from healthy conjunctiva. PMID:28824564
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Structural dynamics of the MecA-ClpC complex: a type II AAA+ protein unfolding machine.
Liu, Jing; Mei, Ziqing; Li, Ningning; Qi, Yutao; Xu, Yanji; Shi, Yigong; Wang, Feng; Lei, Jianlin; Gao, Ning
2013-06-14
The MecA-ClpC complex is a bacterial type II AAA(+) molecular machine responsible for regulated unfolding of substrates, such as transcription factors ComK and ComS, and targeting them to ClpP for degradation. The six subunits of the MecA-ClpC complex form a closed barrel-like structure, featured with three stacked rings and a hollow passage, where substrates are threaded and translocated through successive pores. Although the general concepts of how polypeptides are unfolded and translocated by internal pore loops of AAA(+) proteins have long been conceived, the detailed mechanistic model remains elusive. With cryoelectron microscopy, we captured four different structures of the MecA-ClpC complexes. These complexes differ in the nucleotide binding states of the two AAA(+) rings and therefore might presumably reflect distinctive, representative snapshots from a dynamic unfolding cycle of this hexameric complex. Structural analysis reveals that nucleotide binding and hydrolysis modulate the hexameric complex in a number of ways, including the opening of the N-terminal ring, the axial and radial positions of pore loops, the compactness of the C-terminal ring, as well as the relative rotation between the two nucleotide-binding domain rings. More importantly, our structural and biochemical data indicate there is an active allosteric communication between the two AAA(+) rings and suggest that concerted actions of the two AAA(+) rings are required for the efficiency of the substrate unfolding and translocation. These findings provide important mechanistic insights into the dynamic cycle of the MecA-ClpC unfoldase and especially lay a foundation toward the complete understanding of the structural dynamics of the general type II AAA(+) hexamers.
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Silva, Patricia Vollú; Cruz, Raquel Souza; Keim, Luiz Sérgio; de Paula, Geraldo Renato; Carvalho, Bernadete Teixeira Ferreira; Coelho, Leonardo Rocchetto; Carvalho, Maria Cícera da Silva; da Rosa, Joel Mauricio Corrêa; Figueiredo, Agnes Marie Sá; Teixeira, Lenise Arneiro
2013-01-01
We analysed the antimicrobial susceptibility, biofilm formation and genotypic profiles of 27 isolates of Staphylococcus haemolyticus obtained from the blood of 19 patients admitted to a hospital in Rio de Janeiro, Brazil. Our analysis revealed a clinical significance of 36.8% and a multi-resistance rate of 92.6% among these isolates. All but one isolate carried the mecA gene. The staphylococcal cassette chromosome mec type I was the most prevalent mec element detected (67%). Nevertheless, the isolates showed clonal diversity based on pulsed-field gel electrophoresis analysis. The ability to form biofilms was detected in 66% of the isolates studied. Surprisingly, no icaAD genes were found among the biofilm-producing isolates. PMID:24037208
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Silva, Patricia Vollú; Cruz, Raquel Souza; Keim, Luiz Sérgio; Paula, Geraldo Renato de; Carvalho, Bernadete Teixeira Ferreira; Coelho, Leonardo Rocchetto; Carvalho, Maria Cícera da Silva; Rosa, Joel Mauricio Corrêa da; Figueiredo, Agnes Marie Sá; Teixeira, Lenise Arneiro
2013-09-01
We analysed the antimicrobial susceptibility, biofilm formation and genotypic profiles of 27 isolates of Staphylococcus haemolyticus obtained from the blood of 19 patients admitted to a hospital in Rio de Janeiro, Brazil. Our analysis revealed a clinical significance of 36.8% and a multi-resistance rate of 92.6% among these isolates. All but one isolate carried the mecA gene. The staphylococcal cassette chromosome mec type I was the most prevalent mec element detected (67%). Nevertheless, the isolates showed clonal diversity based on pulsed-field gel electrophoresis analysis. The ability to form biofilms was detected in 66% of the isolates studied. Surprisingly, no icaAD genes were found among the biofilm-producing isolates.
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Kizerwetter-Swida, M; Chrobak, D; Rzewuska, M; Binek, M
2009-01-01
We have evaluated 102 Staphylococcus intermedius isolates of canine origin for susceptibility to antimicrobial primary agents, i.e. penicillin, amoxicillin, amoxicillin with clavulanic acid, cefuroxime, trimethoprim/sulfonamides, neomycin, streptomycin, gentamicin, norfloxacin, tetracycline, vancomycin, erythromycin and secondary agents, i.e., chloramphenicol, ciprofloxacin, lincomycin, teicoplanin, rifampicin, imipenem, mupirocin. Antimicrobial sensitivity was examined using the disk diffusion method and performed according to NCCLS quidelines. Methicillin resistance was detected using the disk diffusion method with oxacillin, and the occurrence of mecA gene was detected by PCR. Resistance to streptomycin, penicillin, amoxicillin, neomycin, followed by tetracycline was predominant. From 14 mecA-positive strains, 12 were multidrug-resitant, and the remaining two showed atypical susceptibility. One strain resistant to oxacillin in the disc diffusion method was mecA-negative, suggesting a different mechanism of resistance. Our results indicate that the emergence of S. intermedius resistance to methicillin may be underestimated. In case of clinical multidrug-resitant S. intermedius isolates, resistance to methicillin should be considered.
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Human Staphylococcus aureus lineages among Zoological Park residents in Greece
Drougka, E.; Foka, A.; Posantzis, D.; Giormezis, N.; Anastassiou, E.D.; Petinaki, E.; Spiliopoulou, I.
2015-01-01
Staphylococcus aureus is a part of the microbiota flora in many animal species. The clonal spread of S. aureus among animals and personnel in a Zoological Park was investigated. Samples were collected from colonized and infected sites among 32 mammals, 11 birds and eight humans. The genes mecA, mecC, lukF/lukS-PV (encoding Panton-Valentine leukocidin, PVL) and tst (toxic shock syndrome toxin-1) were investigated by PCR. Clones were defined by Multilocus Sequence Typing (MLST), spa type and Pulsed-Field Gel Electrophoresis (PFGE). Seven S. aureus isolates were recovered from four animals and one from an employee. All were mecA, mecC and tst–negative, whereas, one carried the PVL genes and was isolated from an infected Squirrel monkey. Clonal analysis revealed the occurrence of seven STs, eight PFGE and five spa types including ones of human origin. Even though a variety of genotypes were identified among S. aureus strains colonizing zoo park residents, our results indicate that colonization with human lineages has indeed occurred. PMID:26623381
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Luini, M; Cremonesi, P; Magro, G; Bianchini, V; Minozzi, G; Castiglioni, B; Piccinini, R
2015-08-05
Staphylococcus aureus is one of the most common mastitis-causing pathogens worldwide. In the last decade, livestock-associated methicillin-resistant S. aureus (LA-MRSA) infections have been described in several species, included the bovines. Hence, this paper investigates the diffusion of MRSA within Italian dairy herds; the strains were further characterized using a DNA microarray, which detects 330 different sequences, including the methicillin-resistance genes mecA and mecC and SCCmec typing. The analysis of overall patterns allows the assignment to Clonal Complexes (CC). Overall 163 S. aureus isolates, collected from quarter milk samples in 61 herds, were tested. MRSA strains were further processed using spa typing. Fifteen strains (9.2%), isolated in 9 herds (14.75%), carried mecA, but none harboured mecC. MRSA detection was significantly associated (P<0.011) with a within-herd prevalence of S. aureus intra-mammary infections (IMI) ≤5%. Ten MRSA strains were assigned to CC398, the remaining ones to CC97 (n=2), CC1 (n=2) or CC8 (n=1). In 3 herds, MRSA and MSSA co-existed: CC97-MRSA with CC398-MSSA, CC1-MRSA with CC8-MSSA and CC398-MRSA with CC126-MSSA. The results of spa typing showed an overall similar profile of the strains belonging to the same CC: t127-CC1, t1730-CC97, t899 in 8 out of 10 CC398. In the remaining 2 isolates a new spa type, t14644, was identified. The single CC8 was a t3092. The SCCmec cassettes were classified as type IV, type V or type IV/V composite. All or most strains harboured the genes encoding the β-lactamase operon and the tetracycline resistance. Streptogramin resistance gene was related to CC398. Enterotoxin and leukocidin genes were carried only by CC1, CC8 and CC97-MRSA. The persistence of MRSA clones characterized by broader host range, in epidemiologically unrelated areas and in dairy herds with low prevalence of S. aureus IMI, might enhance the risk for adaptation to human species. Copyright © 2015 Elsevier B.V. All rights reserved.
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Lemos, Mark; Xiao, Yanmei; Bjornson, Marta; Wang, Jin-zheng; Hicks, Derrick; de Souza, Amancio; Wang, Chang-Quan; Yang, Panyu; Ma, Shisong; Dinesh-Kumar, Savithramma; Dehesh, Katayoon
2016-01-01
The exquisite harmony between hormones and their corresponding signaling pathways is central to prioritizing plant responses to simultaneous and/or successive environmental trepidations. The crosstalk between jasmonic acid (JA) and salicylic acid (SA) is an established effective mechanism that optimizes and tailors plant adaptive responses. However, the underlying regulatory modules of this crosstalk are largely unknown. Global transcriptomic analyses of mutant plants (ceh1) with elevated levels of the stress-induced plastidial retrograde signaling metabolite 2-C-methyl-D-erythritol cyclopyrophosphate (MEcPP) revealed robustly induced JA marker genes, expected to be suppressed by the presence of constitutively high SA levels in the mutant background. Analyses of a range of genotypes with varying SA and MEcPP levels established the selective role of MEcPP-mediated signal(s) in induction of JA-responsive genes in the presence of elevated SA. Metabolic profiling revealed the presence of high levels of the JA precursor 12-oxo-phytodienoic acid (OPDA), but near wild type levels of JA in the ceh1 mutant plants. Analyses of coronatine-insensitive 1 (coi1)/ceh1 double mutant plants confirmed that the MEcPP-mediated induction is JA receptor COI1 dependent, potentially through elevated OPDA. These findings identify MEcPP as a previously unrecognized central regulatory module that induces JA-responsive genes in the presence of high SA, thereby staging a multifaceted plant response within the environmental context. PMID:26733689
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Bacteriophages carrying antibiotic resistance genes in fecal waste from cattle, pigs, and poultry.
Colomer-Lluch, Marta; Imamovic, Lejla; Jofre, Juan; Muniesa, Maite
2011-10-01
This study evaluates the occurrence of bacteriophages carrying antibiotic resistance genes in animal environments. bla(TEM), bla(CTX-M) (clusters 1 and 9), and mecA were quantified by quantitative PCR in 71 phage DNA samples from pigs, poultry, and cattle fecal wastes. Densities of 3 to 4 log(10) gene copies (GC) of bla(TEM), 2 to 3 log(10) GC of bla(CTX-M), and 1 to 3 log(10) GC of mecA per milliliter or gram of sample were detected, suggesting that bacteriophages can be environmental vectors for the horizontal transfer of antibiotic resistance genes.
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Chen, Changguo; Zhao, Qiangyuan; Guo, Jianwei; Li, Yanjun; Chen, Qiuyuan
2017-08-01
The aim of this study was to develop a rapid detection assay to identify methicillin-resistant Staphylococcus aureus by simultaneous testing for the mecA, nuc, and femB genes using the loop-mediated isothermal amplification (LAMP) method. LAMP primers were designed using online bio-software ( http://primerexplorer.jp/e/ ), and amplification reactions were performed in an isothermal temperature bath. The products were then examined using 2% agarose gel electrophoresis. MecA, nuc, and femB were confirmed by triplex TaqMan real-time PCR. For better naked-eye inspection of the reaction result, hydroxy naphthol blue (HNB) was added to the amplification system. Within 60 min, LAMP successfully amplified the genes of interest under isothermal conditions at 63 °C. The results of 2% gel electrophoresis indicated that when the Mg 2+ concentration in the reaction system was 6 μmol, the amplification of the mecA gene was relatively good, while the amplification of the nuc and femB genes was better at an Mg 2+ concentration of 8 μmol. Obvious color differences were observed by adding 1 μL (3.75 mM) of HNB into 25 μL reaction system. The LAMP assay was applied to 128 isolates cases of methicillin-resistant Staphylococcus aureus, which were separated from the daily specimens and identified by Vitek microbial identification instruments. The results were identical for both LAMP and PCR. LAMP offers an alternative detection assay for mecA, nuc, and femB and is faster than other methods.
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Rapid detection of mecA and spa by the loop-mediated isothermal amplification (LAMP) method.
Koide, Y; Maeda, H; Yamabe, K; Naruishi, K; Yamamoto, T; Kokeguchi, S; Takashiba, S
2010-04-01
To develop a detection assay for staphylococcal mecA and spa by using loop-mediated isothermal amplification (LAMP) method. Staphylococcus aureus and other related species were subjected to the detection of mecA and spa by both PCR and LAMP methods. The LAMP successfully amplified the genes under isothermal conditions at 64 degrees C within 60 min, and demonstrated identical results with the conventional PCR methods. The detection limits of the LAMP for mecA and spa, by gel electrophoresis, were 10(2) and 10 cells per tube, respectively. The naked-eye inspections were possible with 10(3) and 10 cells for detection of mecA and spa, respectively. The LAMP method was then applied to sputum and dental plaque samples. The LAMP and PCR demonstrated identical results for the plaque samples, although frequency in detection of mecA and spa by the LAMP was relatively lower for the sputum samples when compared to the PCR methods. Application of the LAMP enabled a rapid detection assay for mecA and spa. The assay may be applicable to clinical plaque samples. The LAMP offers an alternative detection assay for mecA and spa with a great advantage of the rapidity.
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Vialard, J E; Gilbert, C S; Green, C M; Lowndes, N F
1998-10-01
The Saccharomyces cerevisiae RAD9 checkpoint gene is required for transient cell-cycle arrests and transcriptional induction of DNA repair genes in response to DNA damage. Polyclonal antibodies raised against the Rad9 protein recognized several polypeptides in asynchronous cultures, and in cells arrested in S or G2/M phases while a single form was observed in G1-arrested cells. Treatment with various DNA damaging agents, i.e. UV, ionizing radiation or methyl methane sulfonate, resulted in the appearance of hypermodified forms of the protein. All modifications detected during a normal cell cycle and after DNA damage were sensitive to phosphatase treatment, indicating that they resulted from phosphorylation. Damage-induced hyperphosphorylation of Rad9 correlated with checkpoint functions (cell-cycle arrest and transcriptional induction) and was cell-cycle stage- and progression-independent. In asynchronous cultures, Rad9 hyperphosphorylation was dependent on MEC1 and TEL1, homologues of the ATR and ATM genes. In G1-arrested cells, damage-dependent hyperphosphorylation required functional MEC1 in addition to RAD17, RAD24, MEC3 and DDC1, demonstrating cell-cycle stage specificity of the checkpoint genes in this response to DNA damage. Analysis of checkpoint protein interactions after DNA damage revealed that Rad9 physically associates with Rad53.
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Skálová, Alena; Vanecek, Tomas; Simpson, Roderick H W; Vazmitsel, Marina A; Majewska, Hanna; Mukensnabl, Petr; Hauer, Lukas; Andrle, Pavel; Hosticka, Lubor; Grossmann, Petr; Michal, Michal
2013-11-01
The recurrent translocations t(11;19) and t(11;15) resulting in CRTC1-MAML2 or CRTC3-MAML2 fusion oncogenes, respectively, are identified in a large proportion of mucoepidermoid carcinomas (MECs) of the salivary gland and have impact on prognosis. However, there are conflicting data on the specificity of this translocation, in particular, on its putative occurrence in Warthin tumor (WT) of the parotid gland as reported in few previous cases. It was speculated that extensive squamous metaplasia could explain the presence of t(11;19) translocation in a subset of WTs. We evaluated 76 salivary gland tumors, including 16 cases of metaplastic WT and 8 cases of pleomorphic adenoma (PA) with squamous and/or mucinous metaplasia, extensive enough morphologically to mimic MEC. Detection of CRTC1-MAML2 and CRTC3-MAML2 fusion transcripts and MAML2 gene break was performed using nested reverse transcription-polymerase chain reaction and fluorescence in situ hybridization (FISH), respectively. None of 16 analyzed metaplastic WTs showed positivity for fusion transcripts CRTC1-MAML2 or CRTC3-MAML2, and none showed rearrangement of the MAML2 gene by FISH. Similarly, we did not detect these transcripts or break of MAML2 gene in any case of PA with extensive squamous/mucinous metaplasia. For comparison, 40 cases of low-grade MEC were also evaluated. CRTC1-MAML2 and CRTC3-MAML2 fusion transcripts were detected in 17 and 5 cases, respectively. The FISH method using break-apart probe demonstrated the MAML2 gene rearrangement in 25 cases of low-grade MEC. In contrast to low-grade MEC, neither metaplastic WTs nor metaplastic PAs harbored translocations t(11;19) and anticipated t(11;15) resulting in CRTC1-MAML2 and CRTC3-MAML2 fusion transcripts, respectively, and/or MAML2 gene rearrangement.
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2001-02-01
a novel method for the detection of the mecA gene that confers the principle mechanism of methicillin resistance in Staphylococcus aureus . CPT is a...with the mecA gene was used to develop a culture confirmation F-33 assay for methicillin resistant Staphylococcus aureus . The CPT assay was used to...compounds. Examples of signatures would include peptides, aptamers and phage. These are being sought to serve as adjuncts and/or replacements for antibody
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Molecular mechanisms of methicillin resistance in Staphylococcus aureus.
Domínguez, M A; Liñares, J; Martín, R
1997-09-01
Methicillin-resistant Staphylococcus aureus (MRSA) strains are among the most common nosocomial pathogens. The most significant mechanism of resistance to methicillin in this-species is the acquisition of a genetic determinant (mecA gene). However, resistance seems to have a more complex molecular basis, since additional chromosomal material is involved in such resistance. Besides, overproduction of penicillinase and/or alterations in the PBPs can contribute to the formation of resistance phenotypes. Genetic and environmental factors leading to MRSA are reviewed.
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Guza, Rebecca; Kotandeniya, Delshanee; Murphy, Kristopher; Dissanayake, Thakshila; Lin, Chen; Giambasu, George Madalin; Lad, Rahul R; Wojciechowski, Filip; Amin, Shantu; Sturla, Shana J; Hudson, Robert H E; York, Darrin M; Jankowiak, Ryszard; Jones, Roger; Tretyakova, Natalia Y
2011-05-01
Endogenous 5-methylcytosine ((Me)C) residues are found at all CG dinucleotides of the p53 tumor suppressor gene, including the mutational 'hotspots' for smoking induced lung cancer. (Me)C enhances the reactivity of its base paired guanine towards carcinogenic diolepoxide metabolites of polycyclic aromatic hydrocarbons (PAH) present in cigarette smoke. In the present study, the structural basis for these effects was investigated using a series of unnatural nucleoside analogs and a representative PAH diolepoxide, benzo[a]pyrene diolepoxide (BPDE). Synthetic DNA duplexes derived from a frequently mutated region of the p53 gene (5'-CCCGGCACCC GC[(15)N(3),(13)C(1)-G]TCCGCG-3', + strand) were prepared containing [(15)N(3), (13)C(1)]-guanine opposite unsubstituted cytosine, (Me)C, abasic site, or unnatural nucleobase analogs. Following BPDE treatment and hydrolysis of the modified DNA to 2'-deoxynucleosides, N(2)-BPDE-dG adducts formed at the [(15)N(3), (13)C(1)]-labeled guanine and elsewhere in the sequence were quantified by mass spectrometry. We found that C-5 alkylcytosines and related structural analogs specifically enhance the reactivity of the base paired guanine towards BPDE and modify the diastereomeric composition of N(2)-BPDE-dG adducts. Fluorescence and molecular docking studies revealed that 5-alkylcytosines and unnatural nucleobase analogs with extended aromatic systems facilitate the formation of intercalative BPDE-DNA complexes, placing BPDE in a favorable orientation for nucleophilic attack by the N(2) position of guanine. © The Author(s) 2011. Published by Oxford University Press.
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Guza, Rebecca; Kotandeniya, Delshanee; Murphy, Kristopher; Dissanayake, Thakshila; Lin, Chen; Giambasu, George Madalin; Lad, Rahul R.; Wojciechowski, Filip; Amin, Shantu; Sturla, Shana J.; Hudson, Robert H.E.; York, Darrin M.; Jankowiak, Ryszard; Jones, Roger; Tretyakova, Natalia Y.
2011-01-01
Endogenous 5-methylcytosine (MeC) residues are found at all CG dinucleotides of the p53 tumor suppressor gene, including the mutational ‘hotspots’ for smoking induced lung cancer. MeC enhances the reactivity of its base paired guanine towards carcinogenic diolepoxide metabolites of polycyclic aromatic hydrocarbons (PAH) present in cigarette smoke. In the present study, the structural basis for these effects was investigated using a series of unnatural nucleoside analogs and a representative PAH diolepoxide, benzo[a]pyrene diolepoxide (BPDE). Synthetic DNA duplexes derived from a frequently mutated region of the p53 gene (5′-CCCGGCACCC GC[15N3,13C1-G]TCCGCG-3′, + strand) were prepared containing [15N3, 13C1]-guanine opposite unsubstituted cytosine, MeC, abasic site, or unnatural nucleobase analogs. Following BPDE treatment and hydrolysis of the modified DNA to 2′-deoxynucleosides, N2-BPDE-dG adducts formed at the [15N3, 13C1]-labeled guanine and elsewhere in the sequence were quantified by mass spectrometry. We found that C-5 alkylcytosines and related structural analogs specifically enhance the reactivity of the base paired guanine towards BPDE and modify the diastereomeric composition of N2-BPDE-dG adducts. Fluorescence and molecular docking studies revealed that 5-alkylcytosines and unnatural nucleobase analogs with extended aromatic systems facilitate the formation of intercalative BPDE–DNA complexes, placing BPDE in a favorable orientation for nucleophilic attack by the N2 position of guanine. PMID:21245046
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Characterization of Staphylococcus aureus Isolated from Food Products in Western Algeria.
Chaalal, Wafaa; Chaalal, Nadia; Bourafa, Nadjette; Kihal, Mebrouk; Diene, Seydina M; Rolain, Jean-Marc
2018-03-15
The current study aimed to characterize Staphylococcus aureus isolates from foodstuffs collected from western Algeria. A total of 153 S. aureus isolates from various raw and processed foods were obtained and identified using matrix-assisted laser desorption and ionization time-of-flight mass spectrometry. Isolates were characterized by antimicrobial susceptibility testing and toxin gene detection. Methicillin-resistant Staphylococcus aureus (MRSA) isolates were identified by detection of the mecA gene and characterized by staphylococcal cassette chromosome mec (SCCmec) typing. We found that 30.9% (153/495) of food samples were contaminated with S. aureus. Thirty-three (21.5%) S. aureus isolates were identified as MRSA, and 16.9% (26/153) carried the mecA gene. Three SCCmec types were identified of which type IV was the most common (69.2%) followed by type V (15.3%) and type II (7.6%). Two MRSA isolates were not typable with SCCmec typing. None of the examined isolates harbored mecC. Furthermore, 14.3% (22/153) of the isolates were toxigenic S. aureus. The cytotoxin gene pvl was detected in 11.1% of the S. aureus isolates. This gene was more commonly detected (76.4%) in MRSA isolates than in methicillin-suceptible Staphylococcus aureus (MSSA) isolates. The tsst-1 gene coding for toxic shock syndrome toxin was isolated rarely (3.2%) and only in MSSA isolates. According to disk diffusion test results, 70 isolates were resistant to only one antimicrobial drug, and 51 (33.3%) isolates were multidrug resistant. Other 32 isolates were susceptible to all antibiotics. Our study highlights, for the first time, a high prevalence of multidrug-resistant S. aureus isolates carrying pvl or tsst-1 found in food products in Algeria. The risk of MRSA transmission through the food chain cannot be disregarded, particularly in uncooked foods.
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Lemos, Mark; Xiao, Yanmei; Bjornson, Marta; Wang, Jin-Zheng; Hicks, Derrick; Souza, Amancio de; Wang, Chang-Quan; Yang, Panyu; Ma, Shisong; Dinesh-Kumar, Savithramma; Dehesh, Katayoon
2016-03-01
The exquisite harmony between hormones and their corresponding signaling pathways is central to prioritizing plant responses to simultaneous and/or successive environmental trepidations. The crosstalk between jasmonic acid (JA) and salicylic acid (SA) is an established effective mechanism that optimizes and tailors plant adaptive responses. However, the underlying regulatory modules of this crosstalk are largely unknown. Global transcriptomic analyses of mutant plants (ceh1) with elevated levels of the stress-induced plastidial retrograde signaling metabolite 2-C-methyl-D-erythritol cyclopyrophosphate (MEcPP) revealed robustly induced JA marker genes, expected to be suppressed by the presence of constitutively high SA levels in the mutant background. Analyses of a range of genotypes with varying SA and MEcPP levels established the selective role of MEcPP-mediated signal(s) in induction of JA-responsive genes in the presence of elevated SA. Metabolic profiling revealed the presence of high levels of the JA precursor 12-oxo-phytodienoic acid (OPDA), but near wild type levels of JA in the ceh1 mutant plants. Analyses of coronatine-insensitive 1 (coi1)/ceh1 double mutant plants confirmed that the MEcPP-mediated induction is JA receptor COI1 dependent, potentially through elevated OPDA. These findings identify MEcPP as a previously unrecognized central regulatory module that induces JA-responsive genes in the presence of high SA, thereby staging a multifaceted plant response within the environmental context. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.
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Téllez-Pérez, Ana Dolores; Alva-Murillo, Nayeli; Ochoa-Zarzosa, Alejandra; López-Meza, Joel E
2012-11-09
Vitamin D has immunomodulatory functions regulating the expression of host defense genes. The aim of this study was to determine the effect of cholecalciferol (vitamin D3) on S. aureus internalization into bovine mammary epithelial cells (bMEC) and antimicrobial peptide (AP) mRNA expression. Cholecalciferol (1-200 nM) did not affect S. aureus growth and bMEC viability; but it reduced bacterial internalization into bMEC (15-74%). Also, bMEC showed a basal expression of all AP genes evaluated, which were induced by S. aureus. Cholecalciferol alone or together with bacteria diminished tracheal antimicrobial peptide (TAP) and bovine neutrophil β-defensin (BNBD) 5 mRNA expression; while alone induced the expression of lingual antimicrobial peptide (LAP), bovine β-defensin 1 (DEFB1) and bovine psoriasin (S100A7), which was inhibited in the presence of S. aureus. This compound (50 nM) increased BNBD10 mRNA expression coinciding with the greatest reduction in S. aureus internalization. Genes of vitamin D pathway (25-hydroxylase and 1 α-hydroxylase) show basal expression, which was induced by cholecalciferol or bacteria. S. aureus induced vitamin D receptor (VDR) mRNA expression, but not in the presence of cholecalciferol. In conclusion, cholecalciferol can reduce S. aureus internalization and differentially regulates AP expression in bMEC. Thus, vitamin D could be an effective innate immunity modulator in mammary gland, which leads to a better defense against bacterial infection. Copyright © 2012 Elsevier B.V. All rights reserved.
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Bitrus, Asinamai Athliamai; Zunita, Zakaria; Bejo, Siti Khairani; Othman, Sarah; Nadzir, Nur Adilah Ahmad
2017-04-04
Staphylococcus aureus more than any other human pathogen is a better model for the study of the adaptive evolution of bacterial resistance to antibiotics, as it has demonstrated a remarkable ability in its response to new antibiotics. This study was designed to investigate the in vitro transfer of mecA gene from methicillin resistant S. aureus to methicillin susceptible S. aureus. The recipient transconjugants were resistant to erythromycin, cefpodoxime and were mecA positive. PCR amplification of mecA after mix culture plating on Luria Bertani agar containing 100 μg/mL showed that 75% of the donor and 58.3% of the recipient transconjugants were mecA positive. Additionally, 61.5% of both the donor cells and recipient transconjugants were mecA positive, while 46.2% and 41.75% of both donor and recipient transconjugants were mecA positive on LB agar containing 50 μg/mL and 30 μg/mL respectively. In this study, the direction of transfer of phenotypic resistance as well as mecA was observed to have occurred from the donor to the recipient strains. This study affirmed the importance of horizontal transfer events in the dissemination of antibiotics resistance among different strains of MRSA.
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Bishop, Justin A; Cowan, Morgan L; Shum, Chung H; Westra, William H
2018-01-01
Mucoepidermoid carcinoma (MEC) is the most common salivary gland malignancy. Recent studies have shown that most MECs harbor gene fusions involving MAML2-an alteration that appears to be specific for MEC, a finding that could be diagnostically useful. While most cases of MEC are histologically straightforward, uncommon variants can cause considerable diagnostic difficulty. We present 2 variants of MEC for which MAML2 studies were crucial in establishing a diagnosis: a previously undescribed ciliated variant, and the recently described Warthin-like variant. All cases of ciliated and Warthin-like MEC were retrieved from the archives of The Johns Hopkins Hospital. Break-apart fluorescence in situ hybridization for MAML2 was performed on all cases. One ciliated MEC and 6 Warthin-like MECs were identified. The ciliated MEC presented as a 4.6 cm cystic lymph node metastasis originating from the tongue base in a 47-year-old woman. The Warthin-like MECs presented as parotid masses ranging in size from 1.2 to 3.3 (mean, 2.7 cm) in 4 women and 2 men. The ciliated MEC consisted of macrocystic spaces punctuated by tubulopapillary proliferations of squamoid cells and ciliated columnar cells. The Warthin-like MECs were comprised of cystic spaces lined by multilayered oncocytic to squamoid cells surrounded by a circumscribed cuff of lymphoid tissue with germinal centers. In these cases, the Warthin-like areas dominated the histologic picture. Conventional MEC, when present, represented a minor tumor component. MAML2 rearrangements were identified in all cases. Warthin-like MEC, and now a ciliated form of MEC, are newly described variants of a common salivary gland carcinoma. Unfamiliarity with these novel forms, unanticipated cellular features (eg, cilia), and morphologic overlap with mundane benign processes (eg, developmental ciliated cysts, Warthin tumor) or other carcinomas (eg, ciliated human papillomavirus-related carcinoma) may render these variants susceptible to misdiagnosis. These unusual variants appear to consistently harbor MAML2 fusions-a finding that establishes a clear link to conventional MEC and provides a valuable adjunct in establishing the diagnosis.
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Couto, Natacha; Monchique, Cláudia; Belas, Adriana; Marques, Cátia; Gama, Luís T; Pomba, Constança
2016-06-01
The objective of this study was to investigate the evolution of resistance to antimicrobials, corresponding mechanisms and molecular characteristics of Staphylococcus spp., between 1999 and 2014. Susceptibility to 38 antimicrobials was determined for 632 clinical staphylococcal isolates obtained from companion animals (dogs, cats, horses and other animals). Twenty antimicrobial resistance genes, including mecA and mecC, were screened by PCR. Methicillin-resistant staphylococci were characterized by spa (Staphylococcus aureus), SCCmec, MLST and PFGE typing. Statistical analyses were performed using SAS v9.3 and differences were considered relevant if P ≤ 0.05. The mecA gene was identified in 74 staphylococcal isolates (11.6%): 11 MRSA (40.7%), 40 methicillin-resistant Staphylococcus pseudintermedius (MRSP; 8.7%) and 23 methicillin-resistant CoNS (26.7%). Resistance to the majority of antimicrobials and the number of mecA-positive isolates increased significantly over time. Eighteen spa types were identified, including two new ones. MRSA isolates were divided into three PFGE clusters that included ST22-IV, ST105-II, ST398-V and ST5-VI. Most methicillin-resistant Staphylococcus epidermidis isolates were of clonal complex (CC) 5, including a new ST, and clustered in eight PFGE clusters. MRSP were grouped into five PFGE clusters and included ST45-NT, ST71-II-III, ST195-III, ST196-V, ST339-NT, ST342-IV and the new ST400-III. Methicillin-resistant Staphylococcus haemolyticus clustered in two PFGE clusters. The significant increase in antimicrobial-resistant and mecA-positive isolates in recent years is worrying. Furthermore, several isolates are MDR, which complicates antimicrobial treatment and increases the risk of transfer to humans or human isolates. Several clonal lineages of MRSA and methicillin-resistant S. epidermidis circulating in human hospitals and the community were found, suggesting that companion animals can become infected with and contribute to the dissemination of highly successful human clones. Urgent measures, such as determination of clinical breakpoints and guidelines for antimicrobial use, are needed. © The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.
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POB3 is required for both transcription and replication in the yeast Saccharomyces cerevisiae.
Schlesinger, M B; Formosa, T
2000-01-01
Spt16 and Pob3 form stable heterodimers in Saccharomyces cerevisiae, and homologous proteins have also been purified as complexes from diverse eukaryotes. This conserved factor has been implicated in both transcription and replication and may affect both by altering the characteristics of chromatin. Here we describe the isolation and properties of a set of pob3 mutants and confirm that they have defects in both replication and transcription. Mutation of POB3 caused the Spt(-) phenotype, spt16 and pob3 alleles displayed severe synthetic defects, and elevated levels of Pob3 suppressed some spt16 phenotypes. These results are consistent with previous reports that Spt16 and Pob3 act in a complex that modulates transcription. Additional genetic interactions were observed between pob3 mutations and the genes encoding several DNA replication factors, including POL1, CTF4, DNA2, and CHL12. pob3 alleles caused sensitivity to the ribonucleotide reductase inhibitor hydroxyurea, indicating a defect in a process requiring rapid dNTP synthesis. Mutation of the S phase checkpoint gene MEC1 caused pob3 mutants to lose viability rapidly under restrictive conditions, revealing defects in a process monitored by Mec1. Direct examination of DNA contents by flow cytometry showed that S phase onset and progression were delayed when POB3 was mutated. We conclude that Pob3 is required for normal replication as well as for transcription. PMID:10924459
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Sawicka, Marta; Wanrooij, Paulina H; Darbari, Vidya C; Tannous, Elias; Hailemariam, Sarem; Bose, Daniel; Makarova, Alena V; Burgers, Peter M; Zhang, Xiaodong
2016-06-24
The phosphatidylinositol 3-kinase-related protein kinases are key regulators controlling a wide range of cellular events. The yeast Tel1 and Mec1·Ddc2 complex (ATM and ATR-ATRIP in humans) play pivotal roles in DNA replication, DNA damage signaling, and repair. Here, we present the first structural insight for dimers of Mec1·Ddc2 and Tel1 using single-particle electron microscopy. Both kinases reveal a head to head dimer with one major dimeric interface through the N-terminal HEAT (named after Huntingtin, elongation factor 3, protein phosphatase 2A, and yeast kinase TOR1) repeat. Their dimeric interface is significantly distinct from the interface of mTOR complex 1 dimer, which oligomerizes through two spatially separate interfaces. We also observe different structural organizations of kinase domains of Mec1 and Tel1. The kinase domains in the Mec1·Ddc2 dimer are located in close proximity to each other. However, in the Tel1 dimer they are fully separated, providing potential access of substrates to this kinase, even in its dimeric form. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.
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Mello, Priscila Luiza; Pinheiro, Luiza; Martins, Lisiane de Almeida; Brito, Maria Aparecida Vasconcelos Paiva; Ribeiro de Souza da Cunha, Maria de Lourdes
2017-08-01
The use of antimicrobial agents has led to the emergence of resistant bacterial strains over a relatively short period. Furthermore, Staphylococcus spp. can produce β-lactamase, which explains the survival of these strains in a focus of infection despite the use of a β-lactam antibiotic. The aim of this study was to evaluate the resistance of Staphylococcus spp. isolated from bovine subclinical mastitis to oxacillin and vancomycin (by minimum inhibitory concentration) and to detect vancomycin heteroresistance by a screening method. We also evaluated β-lactamase production and resistance due to hyperproduction of this enzyme and investigated the mecA and mecC genes and performed staphylococcal cassette chromosome mec typing. For this purpose, 181 Staphylococcus spp. isolated from mastitis subclinical bovine were analyzed. Using the phenotypic method, 33 (18.2%) of Staphylococcus spp. were resistant to oxacillin. In contrast, all isolates were susceptible to vancomycin, and heteroresistance was detected by the screening method in 13 isolates. Production of β-lactamase was observed in 174 (96%) of the Staphylococcus spp. isolates. The mecA gene was detected in 8 isolates, all of them belonging to the species Staphylococcus epidermidis, and staphylococcal cassette chromosome mec typing revealed the presence of type I and type IV isolates. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
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Panda, Sasmita; Kar, Sarita; Sharma, Savitri; Singh, Durg V
2016-09-01
This study aimed to determine the presence of antibiotic resistance genes (ARGs), SCCmec elements and genetic relatedness among Staphylococcus haemolyticus isolated from patients with a variety of eye infections (n=11) and from healthy conjunctiva (n=7). Minimum inhibitory concentrations were determined for 14 antimicrobials according to BSAC guidelines. PCR was used to identify the presence of mecA, mecC, SCCmec type and ARGs. Sequencing was used to determine mutations in gyrA, gyrB, topoisomerase IVA and IVB genes. Genetic relatedness was determined by PFGE. Of the 18 isolates, 17 showed resistance to at least one antibiotic, but none showed resistance to vancomycin or rifampicin. Ten isolates were oxacillin-resistant and carried the mecA gene, eight of which belonged to SCCmec type V. The presence of non-mec SCC elements in two meticillin-susceptible isolates and untypeable SCC elements in meticillin-resistant isolates suggests the involvement of S. haemolyticus in the diversification of SCC elements. Sequence analysis revealed point mutations in gyrA (Ser-84→Leu) and topoisomerase IVA genes (Ser-80→Leu) in 13 isolates, and additional variation in the QRDR (Asp-84→Asn) of two isolates, showing good correlation between mutations in gyrA and topoisomerase IV genes and the level of resistance to fluoroquinolones. PFGE analysis showed distinct pulsotypes forming two major clusters, indicating the existence of diversity among isolates, irrespective of the source of isolation. This study suggests that S. haemolyticus isolates from infected eyes and healthy conjunctivae invariably carried ARGs and SCCmec elements and showed diversity in their genomic content, irrespective of the source of isolation. Copyright © 2016 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.
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Wang, Hye-young; Kim, Sunghyun; Kim, Jungho; Park, Soon-Deok
2014-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) is the most prevalent cause of bloodstream infections (BSIs) and is recognized as a major nosocomial pathogen. This study aimed to evaluate a newly designed multiplex real-time PCR assay capable of the simultaneous detection of mecA, S. aureus, and coagulase-negative staphylococci (CoNS) in blood culture specimens. The Real-MRSA and Real-MRCoNS multiplex real-time PCR assays (M&D, Republic of Korea) use the TaqMan probes 16S rRNA for Staphylococcus spp., the nuc gene for S. aureus, and the mecA gene for methicillin resistance. The detection limit of the multiplex real-time PCR assay was 103 CFU/ml per PCR for each gene target. The multiplex real-time PCR assay was evaluated using 118 clinical isolates from various specimen types and a total of 350 positive blood cultures from a continuous monitoring blood culture system. The results obtained with the multiplex real-time PCR assay for the three targets were in agreement with those of conventional identification and susceptibility testing methods except for one organism. Of 350 positive bottle cultures, the sensitivities of the multiplex real-time PCR kit were 100% (166/166 cultures), 97.2% (35/36 cultures), and 99.2% (117/118 cultures) for the 16S rRNA, nuc, and mecA genes, respectively, and the specificities for all three targets were 100%. The Real-MRSA and Real-MRCoNS multiplex real-time PCR assays are very useful for the rapid accurate diagnosis of staphylococcal BSIs. In addition, the Real-MRSA and Real-MRCoNS multiplex real-time PCR assays could have an important impact on the choice of appropriate antimicrobial therapy, based on detection of the mecA gene. PMID:24648566
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Ebinuma, Takayuki; Soga, Yoshihiko; Sato, Takamaro; Matsunaga, Kazuyuki; Kudo, Chieko; Maeda, Hiroshi; Maeda, Yoshinobu; Tanimoto, Mitsune; Takashiba, Shogo
2014-06-01
We recently reported frequent detection of antibiotic-resistant bacteria on the oral mucosa during the period of hematopoietic cell transplantation (HCT) and suggested an association between oral mucositis and antibiotic-resistant bacterial infection. Methicillin-resistant Staphylococcus spp. were frequently detected, and the oral cavity may be a reservoir of the gene mediating methicillin resistance, mecA. Here, we examined the frequency of mecA carriers in patients undergoing HCT. Fifty-nine patients (male (M) = 37, female (F) = 22, 47.3 ± 11.0 years) receiving HCT were enrolled in this study. Buccal swab samples were obtained four times from day -7 to day +20 (once/week), and mecA was detected by PCR. Fifty-two subjects without systemic disease, who completed dental treatment, especially periodontal treatment (M = 21, F = 31, 55.4 ± 14.2 years), were also enrolled as controls and checked for mecA on the oral mucosa. Seventy-six percent (45/59) of the HCT patients carried mecA at least once in the study period (days -7 to +20), while no control subjects had mecA. The frequency of mecA carriers was 19.2 % from days -7 to -1, while it was significantly increased on days +7 to +13 and +14 to +20, with frequencies of 60.9 and 63.2 %, respectively (P < 0.01, ANOVA). mecA was detected in oral mucosa of patients undergoing HCT. The high detection frequency of staphylococci resistant to penicillin and beta-lactams in our recent report was supported.
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Tajima, Shogo; Namiki, Ichiro; Koda, Kenji
2017-06-01
The predominance of clear cells in mucoepidermoid carcinomas (MEC) is rare, and cases in which this occurs are termed clear cell variants of MEC. We present a case of a 70-year-old woman complaining of a right buccal mucosal mass, which had increased in size over 1 year. Histological examination revealed the mass to be composed predominantly of clear tumor cells, with mucin-containing cells and intermediate cell-like cells. Immunohistochemistry indicated that the tumor was positive for CK5/6 and p63, but negative for myoepithelial markers such as S-100 protein, αSMA, and calponin. These findings ruled out the possibility of a clear cell myoepithelial carcinoma, which is the most frequently observed type of salivary carcinoma composed predominantly of clear cells. However, it is difficult to distinguish between clear cell variants of MEC and hyalinizing clear cell carcinoma. Therefore, we performed fluorescence in situ hybridization to determine whether MAML2 rearrangement had occurred in this mass. Direct sequencing of the RT-PCR product demonstrated CRTC1-MAML2 fusion between exon 1 of CRTC1 and exon 2 of MAML2. Thus, the diagnosis of clear cell variant of MEC was confirmed. This is the first report of CRTC1-MAML2 fusion gene detection in a clear cell variant of MEC.
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Conceição, Teresa; de Lencastre, Hermínia; Aires-de-Sousa, Marta
2017-01-01
Although livestock-associated ST398 methicillin-resistant Staphylococcus aureus (MRSA) has been widely reported in different geographic regions, MRSA carriage studies among healthy pigs in Portugal are very limited. In total, 101 swine nasal samples from two Portuguese farms were screened for MRSA. In addition five swine workers (including one veterinary and one engineer) and four household members were nasally screened. The isolates were characterized by spa typing, SCCmec typing and MLST. All isolates were tested for antimicrobial susceptibility, presence of mecA and mecC genes, and virulence determinants. MRSA prevalence in swine was 99% (100/101), 80% (4/5) in swine workers and 25% (1/4) in household members. All isolates belonged to ST398 distributed over two spa types-t011 (57%) and t108 (42%). SCCmec type V was present in most of the isolates (n = 95; 82%) while 21 isolates amplified the mecA gene only and were classified as nontypeable. The majority of the isolates were resistant to tetracycline (100%), clindamycin (97%), erythromycin (96%), chloramphenicol (84%) and gentamycin (69%). Notably, 12% showed resistance to quinupristin-dalfopristin (MICs 3-8 μg/mL). Beta-hemolysin (81%) and gamma-hemolysin (74%) were the unique virulence determinants detected. None of the isolates harboured PVL or mecC gene. This study showed a massive occurrence of ST398-MRSA in two independent swine farms, highlighting its establishment among healthy pigs in Portugal.
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Xue, Huping; Wu, Zhaowei; Li, Longping; Li, Fan; Wang, Yiqing
2015-01-01
The structure of a composite staphylococcal cassette chromosome (SCC) carried by a methicillin-resistant Staphylococcus haemolyticus (NW19A) isolated from a bovine milk sample was analyzed. The formation of the circular forms of both single SCC elements and composite SCC elements was detected in NW19A. Twenty heavy metal and antibiotic resistance-related genes coexisted in this composite SCC, suggesting that these genes might be coselected under environmental pressure. The mec gene complex in NW19A, designated type C3, is different from classic C1 or C2 gene complexes structurally and likely evolves differently. Furthermore, results from alignment of the SCC composite island of NW19A with 50 related sequences from different staphylococcal strains provided additional evidence to support the notion that coagulase-negative staphylococci (CoNS) are the original host of heavy metal resistance genes among staphylococci. Given that a SCC composite island could transfer freely among different staphylococcal species from different hosts, more attention should be paid to contamination with heavy metals and antibiotics in dairy farming environments, including wastewater, soil, feces, and feed. PMID:26169408
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Xue, Huping; Wu, Zhaowei; Li, Longping; Li, Fan; Wang, Yiqing; Zhao, Xin
2015-09-01
The structure of a composite staphylococcal cassette chromosome (SCC) carried by a methicillin-resistant Staphylococcus haemolyticus (NW19A) isolated from a bovine milk sample was analyzed. The formation of the circular forms of both single SCC elements and composite SCC elements was detected in NW19A. Twenty heavy metal and antibiotic resistance-related genes coexisted in this composite SCC, suggesting that these genes might be coselected under environmental pressure. The mec gene complex in NW19A, designated type C3, is different from classic C1 or C2 gene complexes structurally and likely evolves differently. Furthermore, results from alignment of the SCC composite island of NW19A with 50 related sequences from different staphylococcal strains provided additional evidence to support the notion that coagulase-negative staphylococci (CoNS) are the original host of heavy metal resistance genes among staphylococci. Given that a SCC composite island could transfer freely among different staphylococcal species from different hosts, more attention should be paid to contamination with heavy metals and antibiotics in dairy farming environments, including wastewater, soil, feces, and feed. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
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Grandin, Nathalie; Charbonneau, Michel
2007-01-01
Saccharomyces cerevisiae telomerase-negative cells undergo homologous recombination on subtelomeric or TG1–3 telomeric sequences, thus allowing Type I or Type II post-senescence survival, respectively. Here, we find that the DNA damage sensors, Mec1, Mec3 and Rad24 control Type II recombination, while the Rad9 adaptor protein and the Rad53 and Chk1 effector kinases have no effect on survivor type selection. Therefore, the Mec1 and Mec3 checkpoint complexes control telomeric recombination independently of their roles in generating and amplifying the Mec1-Rad53-Chk1 kinase cascade. rfa1-t11 mutant cells, bearing a mutation in Replication Protein A (RPA) conferring a defect in recruiting Mec1-Ddc2, were also deficient in both types of telomeric recombination. Importantly, expression of an Rfa1-t11-Ddc2 hybrid fusion protein restored checkpoint-dependent arrest, but did not rescue defective telomeric recombination. Therefore, the Rfa1-t11-associated defect in telomeric recombination is not solely due to its failure to recruit Mec1. We have also isolated novel alleles of RFA1 that were deficient in Type I but not in Type II recombination and proficient in checkpoint control. Therefore, the checkpoint and recombination functions of RPA can be genetically separated, as can the RPA-mediated control of the two types of telomeric recombination. PMID:17202155
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Detection of meca gene from methicillin resistant staphylococcus aureus isolates of north sumatera
NASA Astrophysics Data System (ADS)
Septiani Nasution, Gabriella; Suryanto, Dwi; Lia Kusumawati, R.
2018-03-01
Methicillin Resistant Staphylococcus aureus (MRSA) is a major pathogen associated with hospital-acquired infections (nosocomial infections). MRSA is a type of S. aureus resistant to the sub-group of beta-lactam antibiotics such as penicillin, cephalosporin, monobactam, and carbapenem. MRSA is resistant because of genetic changes caused by exposure to irrational antibiotic therapy. This study aimed to detect mecA gene in North Sumatra isolates of MRSA and to determine the pattern of antibiotic resistance in S.aureus isolates classified as MRSA by Vitek 2 Compact in the Central Public Hospital Haji Adam Malik, Medan. Samples were 40 isolates of S. aureus classified as MRSA obtained from clinical microbiology specimens. DNA isolation of the isolates was conducted by a method of freeze-thaw cycling. Amplification of mecA gene was done by PCR technique using specific primer for the gene. PCR products were visualized using mini-gel electrophoresis. The results showed that all MRSA isolates showed to have 533 bp band of mecA. Antibiotics test of Vitek 2 Compact showed that despite all isolates were resistant to beta-lactam antibiotics groups; the isolates showed multidrug resistant to other common antibiotics, such as aminoglycosides, macrolides, and fluoroquinolones. However, they were still sensitive to vancomycin (82.5% isolates), linezolid (97.5% isolates), and tigecycline (100% isolates).
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Antibiotic Resistance Genes in the Bacteriophage DNA Fraction of Environmental Samples
Colomer-Lluch, Marta; Jofre, Juan; Muniesa, Maite
2011-01-01
Antibiotic resistance is an increasing global problem resulting from the pressure of antibiotic usage, greater mobility of the population, and industrialization. Many antibiotic resistance genes are believed to have originated in microorganisms in the environment, and to have been transferred to other bacteria through mobile genetic elements. Among others, β-lactam antibiotics show clinical efficacy and low toxicity, and they are thus widely used as antimicrobials. Resistance to β-lactam antibiotics is conferred by β-lactamase genes and penicillin-binding proteins, which are chromosomal- or plasmid-encoded, although there is little information available on the contribution of other mobile genetic elements, such as phages. This study is focused on three genes that confer resistance to β-lactam antibiotics, namely two β-lactamase genes (blaTEM and blaCTX-M9) and one encoding a penicillin-binding protein (mecA) in bacteriophage DNA isolated from environmental water samples. The three genes were quantified in the DNA isolated from bacteriophages collected from 30 urban sewage and river water samples, using quantitative PCR amplification. All three genes were detected in the DNA of phages from all the samples tested, in some cases reaching 104 gene copies (GC) of blaTEM or 102 GC of blaCTX-M and mecA. These values are consistent with the amount of fecal pollution in the sample, except for mecA, which showed a higher number of copies in river water samples than in urban sewage. The bla genes from phage DNA were transferred by electroporation to sensitive host bacteria, which became resistant to ampicillin. blaTEM and blaCTX were detected in the DNA of the resistant clones after transfection. This study indicates that phages are reservoirs of resistance genes in the environment. PMID:21390233
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Theoretical study on the multi-channel reaction of OH radical with 5-methylcytosine
NASA Astrophysics Data System (ADS)
Wang, Wenliang; Jin, Lingxia; Wang, Weina; Lu, Jian; Yang, Jianming
2007-08-01
All the possible addition and hydrogen abstraction reactions of OH radical with 5-methylcytosine (5-MeC) have been investigated at B3LYP/6-31++G(d,p)//B3LYP/6-31G(d,p)+ZPE level. The results indicate that OH radical may form complexes with 5-MeC, and the reaction is assumed to occur from these complexes. The estimated activation energies corresponding to addition reactions at N3, C4, C5 and C6 sites of the 5-MeC are 80.96, 63.41, 0.00 and 0.30 kJ/mol, respectively. The order of stability of adducts is P4(C6) > P3(C5) > P2(C4) > P1(N3). The activation energies corresponding to the H9, H10, H11, and H14 abstraction reactions from the 5-MeC are all small, and the stabilization of the products is P8(H14) > P6(H10) > P5(H9) > P7(H11).
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Choi, Su Mi; Kim, Seung-Han; Kim, Hee-Jung; Lee, Dong-Gun; Choi, Jung-Hyun; Yoo, Jin-Hong; Kang, Jin-Han; Shin, Wan-Shik; Kang, Moon-Won
2003-01-01
We developed multiplex polymerase chain reaction (PCR) to detect aac(6 ')/aph(2 "), aph(3 ')-IIIa, and ant(4 ')-Ia, the genes encoding the most clinically relevant amino-glycoside modifying enzymes (AME), and simultaneously, the methicillin resistant gene, mecA, in Staphylococcus species. Clinical isolates of 45 S. aureus and 47 coagulase negative staphylococci (CNS) from tertiary university hospitals were tested by conventional susceptibility testing, using the agar dilution method and by multiplex PCR. Of a total of 92 isolates, 61 isolates were found to be methicillin-resistant. Of these, 54 isolates (89%) were found to be harboring mecA. Seventy-five percent of the 92 isolates demonstrated resistance to at least one of the aminoglycosides tested. Moreover, resistance to aminoglycosides was closely associated with methicillin-resistance (p<0.05). The most prevalent AME gene was aac(6 ')/aph(2 ") which was found in 65% of the isolates, and ant(4 ')-Ia and aph(3 ')-IIIa were present in 41% and 9% of the isolates, respectively. The concordance between methicillin-resistance and the presence of mecA gene was 98% in S. aureus and 81% in CNS. The concordance between gentamicin resistance and the presence of aac(6 ')/aph(2 ") gene was 100% in S. aureus and 85% in CNS. The multiplex PCR method that we developed appears to be both a more rapid and reliable than conventional method. PMID:14555812
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Soloaga, R; Corso, A; Gagetti, P; Faccone, D; Galas, M
2004-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) is a significant pathogen that has emerged over the last four decades, causing both nosocomial and community-acquired infections. Rapid and accurate detection of methicillin resistance in S. aureus is important for the use of appropriate antimicrobial therapy and for the control of nosocomial spread of MRSA strains. We evaluated the efficiency of conventional methods for detection of methicillin resistance such as the disk diffusion, agar dilution, oxacillin agar screen test, and the latex agglutination test MRSA-Screen latex, in 100 isolates of S. aureus, 79 mecA positive and 21 mecA negative. The MRSA-Screen latex (Denka Seiken, Niigata, Japón), is a latex agglutination method that detects the presence of PLP-2a, product of mecA gene in S. aureus. The PCR of the mecA gene was used as the "gold standard" for the evaluation of the different methods tested. The percentages of sensitivity and specificity were as follows: disk difusión 97 and 100%, agar dilution 97 and 95%, oxacillin agar screen test 100 and 100%, and MRSA-Screen latex, 100 and 100 %. All methods presented high sensitivity and specificity, but MRSA-Screen latex had the advantage of giving a reliable result, equivalent to PCR, in only 15 minutes.
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Frey, Amy B; Wilson, Deborah A; LaSalvia, Margaret M; Tan, Carmela D; Rodriguez, E Rene; Shrestha, Nabin K; Hall, Gerri S; Procop, Gary W
2011-11-01
We use the BD GeneOhm StaphSR Assay (BD Diagnostics, Oakville, Canada) to screen for Staphylococcus aureus nasal colonization and sought to evaluate this assay for the assessment of valve specimens from patients with endocarditis. We examined 23 paired fresh and formalin-fixed, paraffin-embedded cardiac valve tissue samples, 12 of which had S aureus endocarditis, using the BD GeneOhm StaphSR Assay for the detection and differentiation of methicillin-susceptible and methicillin-resistant S aureus. This assay appropriately characterized all specimens with respect to the presence or absence of S aureus. There was an 87.5% correlation between the presence or absence of the mecA gene and the oxacillin susceptibility results for the S aureus isolates studied. The GeneOhm StaphSR assay accurately detected S aureus in cardiac valve tissue samples. Rare discordances were observed between oxacillin susceptibility status and mecA gene detection by this assay.
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Genotypes, antibiotic resistance, and virulence factors of staphylococci from ready-to-eat food.
Podkowik, Magdalena; Bystroń, Jarosław; Bania, Jacek
2012-01-01
Sixty-seven staphylococcal isolates belonging to 12 species were obtained from 70 ready-to-eat food products. Staphylococcus aureus (n=25), and Staphylococcus epidermidis (n=13) were dominant. Susceptibility to penicillin, oxacillin, tetracycline, clindamycin, gentamicin, erythromycin, ciprofloxacin, and vancomycin was determined. All investigated S. aureus isolates were resistant to at least one antibiotic, and fifteen isolates were resistant to four and more antibiotics. Thirty-eight coagulase-negative staphylococci (CNS) isolates were resistant to at least one antibiotic, and seventeen to four and more antibiotics. Fifteen CNS isolates were mecA positive, and grew in the presence of 6 μg/mL oxacillin. All S. aureus isolates were mecA-negative. Arginine catabolic mobile element (ACME) was found in seven S. epidermidis isolates. Five S. epidermidis isolates harbored ica operon, ACME and were able to form biofilm. Three of them also possessed IS256 element and were mecA-positive. The expression of icaA gene was comparable in five ica-positive S. epidermidis isolates. One of six mecA positive S. epidermidis isolates was classified as sequence type (ST)155, one as ST110, and two as ST88. Two methicillin-resistant Staphylococcus epidermis (MRSE) belonged to new STs, that is, ST362, and ST363. Enterotoxin genes were found in 92% of S. aureus isolates. No enterotoxin gene was detected in analyzed CNS population. We show that ready-to-eat products are an important source of antibiotic-resistant CNS and potentially virulent strains of S. epidermidis, including genotypes undistinguishable from hospital-adapted clones.
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Samutela, Mulemba Tillika; Kalonda, Annie; Mwansa, James; Lukwesa-Musyani, Chileshe; Mwaba, John; Mumbula, Enoch Mulowa; Mwenya, Darlington; Simulundu, Edgar; Kwenda, Geoffrey
2017-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) is globally recognized as an important public health problem. Whereas comprehensive molecular typing data of MRSA strains is available, particularly in Europe, North America and Australia, similar information is very limited in sub-Saharan Africa including Zambia. In this study, thirty two clinical isolates of Staphylococcus aureus , collected at a large referral hospital in Lusaka, Zambia between June 2009 and December 2012 were analysed by Staphylococcal cassette chromosome mec (SCCmec), Staphylococcus protein A gene typing (spa) and detection of the Panton-Valentine Leukocidin genes (pvl) . Three SCC mec types were identified namely SCC mec type IV (65.6%), SCCmec type III (21.9%), SCC mec type I (3.1%). Nine point four percent (9.4%) of the isolates were untypable. Five spa types, which included a novel type, were detected and the most prevalent spa type was t064 (40.6%). Other spa types included spa types t2104 (31.3%), t355 (3.1%) and t1257 (21.9%). The pvl genes were detected in 3 out of 32 isolates. These molecular typing data indicated that the MRSA strains collected in Lusaka were diverse. Although the source of these MRSA was not established, these results stress the need for assessing infection prevention and control procedures at this health-care facility in order to curtail possible nosocomial infections. Furthermore, country-wide surveillance of MRSA in both the community and health-care facilities is recommended for infection prevention and control. To our knowledge, this represents the first study to characterise MRSA using molecular tools in Zambia.
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Bowrin, Valerie; Sutton, Fedora
2016-01-01
Cassava (M. esculenta) gives rise to unique underground stem tubers when stem cuttings are planted in an inverted orientation. The nutritional profile of the stem and root tubers were similar except for protein content which was higher in stem than in root tubers. RT-PCR revealed that several key genes (Mec1, RZF, SuSy1 and PIN2) involved in root tuberization were also expressed in these stem tubers. At five weeks post planting, these genes were expressed in roots and underground stems as in the mature tubers. However at 15 weeks post planting, they were expressed in both root and stem tubers but not in adventitious roots or in the non-tuberized stems. Expression of, the root auxin efflux carrier gene PIN2 in the stem tubers indicate a role for auxin in the stem tuberization process.
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NASA Astrophysics Data System (ADS)
Malla, Spundana; Kadimisetty, Karteek; Fu, You-Jun; Choudhary, Dharamainder; Schenkman, John B.; Rusling, James F.
2017-01-01
Methylation of cytosine (C) at C-phosphate-guanine (CpG) sites enhances reactivity of DNA towards electrophiles. Mutations at CpG sites on the p53 tumor suppressor gene that can result from these adductions are in turn correlated with specific cancers. Here we describe the first restriction-enzyme-assisted LC-MS/MS sequencing study of the influence of methyl cytosines (MeC) on kinetics of p53 gene adduction by model metabolite benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE), using methodology applicable to correlate gene damage sites for drug and pollutant metabolites with mutation sites. This method allows direct kinetic measurements by LC-MS/MS sequencing for oligonucleotides longer than 20 base pairs (bp). We used MeC and non-MeC (C) versions of a 32 bp exon 7 fragment of the p53 gene. Methylation of 19 cytosines increased the rate constant 3-fold for adduction on G at the major reactive CpG in codon 248 vs. the non-MeC fragment. Rate constants for non-CpG codons 244 and 243 were not influenced significantly by MeC. Conformational and hydrophobicity changes in the MeC-p53 exon 7 fragment revealed by CD spectra and molecular modeling increase the BPDE binding constant to G in codon 248 consistent with a pathway in which preceding reactant binding greatly facilitates the rate of covalent SN2 coupling.
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Aoyagi, T; Kaito, C; Sekimizu, K; Omae, Y; Saito, Y; Mao, H; Inomata, S; Hatta, M; Endo, S; Kanamori, H; Gu, Y; Tokuda, K; Yano, H; Kitagawa, M; Kaku, M
2014-09-01
Over-expression of alpha-phenol-soluble modulins (PSMs) results in high virulence of community-associated methicillin-resistant Staphylococcus aureus (MRSA). The psm-mec gene, located in the mobile genetic element SCCmec-II, suppresses PSMαs production. Fifty-two patients with MRSA bacteraemia were enrolled. MRSA isolates were evaluated with regard to the psm-mec gene sequence, bacterial virulence, and the minimum inhibitory concentration (MIC) of vancomycin and teicoplanin. Fifty-one MRSA isolates were classified as SCCmec-II, and 10 had one point mutation in the psm-mec promoter. We compared clinical characteristics and outcomes between mutant MRSA and wild-type MRSA. Production of PSMα3 in mutant MRSA was significantly increased, but biofilm formation was suppressed. Wild-type MRSA caused more catheter-related bloodstream infections (30/41 vs. 3/10, p 0.0028), whereas mutant MRSA formed more deep abscesses (4/10 vs. 3/41, p 0.035). Bacteraemia caused by mutant MRSA was associated with reduced 30-day mortality (1/10 vs. 13/41, p 0.25), although this difference was not significant. The MIC90 of teicoplanin was higher for wild-type MRSA (1.5 mg/L vs. 1 mg/L), but the MIC of vancomycin was not different between the two groups. The 30-day mortality of MRSA with a high MIC of teicoplanin (≥1.5 mg/L) was higher than that of strains with a lower MIC (≤0.75 mg/L) (6/10 vs. 6/33, p 0.017). Mutation of the psm-mec promoter contributes to virulence of SCCmec-II MRSA, and the product of psm-mec may determine the clinical characteristics of bacteraemia caused by SCCmec-II MRSA, but it does not affect mortality. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.
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Lozano, Carmen; Gharsa, Haythem; Ben Slama, Karim; Zarazaga, Myriam; Torres, Carmen
2016-02-04
The interest about Staphylococcus aureus (S. aureus) and methicillin resistant S. aureus (MRSA) in livestock, and domestic and wild animals has significantly increased. The spread of different clonal complexes related to livestock animals, mainly CC398, and the recent description of the new mecC gene, make it necessary to know more about the epidemiology and population structure of this microorganism all over the world. Nowadays, there are several descriptions about the presence of S. aureus and/or MRSA in different animal species (dogs, sheep, donkeys, bats, pigs, and monkeys), and in food of animal origin in African countries. In this continent, there is a high diversity of ethnicities, cultures or religions, as well as a high number of wild animal species and close contact between humans and animals, which can have a relevant impact in the epidemiology of this microorganism. This review shows that some clonal lineages associated with humans (CC1, CC15, CC72, CC80, CC101, and CC152) and animals (CC398, CC130 and CC133) are present in this continent in animal isolates, although the mecC gene has not been detected yet. However, available studies are limited to a few countries, very often with incomplete information, and many more studies are necessary to cover a larger number of African countries.
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Lozano, Carmen; Gharsa, Haythem; Ben Slama, Karim; Zarazaga, Myriam; Torres, Carmen
2016-01-01
The interest about Staphylococcus aureus (S. aureus) and methicillin resistant S. aureus (MRSA) in livestock, and domestic and wild animals has significantly increased. The spread of different clonal complexes related to livestock animals, mainly CC398, and the recent description of the new mecC gene, make it necessary to know more about the epidemiology and population structure of this microorganism all over the world. Nowadays, there are several descriptions about the presence of S. aureus and/or MRSA in different animal species (dogs, sheep, donkeys, bats, pigs, and monkeys), and in food of animal origin in African countries. In this continent, there is a high diversity of ethnicities, cultures or religions, as well as a high number of wild animal species and close contact between humans and animals, which can have a relevant impact in the epidemiology of this microorganism. This review shows that some clonal lineages associated with humans (CC1, CC15, CC72, CC80, CC101, and CC152) and animals (CC398, CC130 and CC133) are present in this continent in animal isolates, although the mecC gene has not been detected yet. However, available studies are limited to a few countries, very often with incomplete information, and many more studies are necessary to cover a larger number of African countries. PMID:27681906
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Imbert, S; Normand, A C; Ranque, S; Costa, J M; Guitard, J; Accoceberry, I; Bonnal, C; Fekkar, A; Bourgeois, N; Houzé, S; Hennequin, C; Piarroux, R; Dannaoui, E; Botterel, F
2018-05-01
Aspergillus section Terrei is a species complex currently comprised of 14 cryptic species whose prevalence in clinical samples as well as antifungal susceptibility are poorly known. The aims of this study were to investigate A. Terrei clinical isolates at the species level and to perform antifungal susceptibility analyses by reference and commercial methods. Eighty-two clinical A. Terrei isolates were collected from 8 French university hospitals. Molecular identification was performed by sequencing parts of beta-tubulin and calmodulin genes. MICs or minimum effective concentrations (MECs) were determined for 8 antifungal drugs using both EUCAST broth microdilution (BMD) methods and concentration gradient strips (CGS). Among the 79 A. Terrei isolates, A. terreus stricto sensu ( n = 61), A. citrinoterreus ( n = 13), A. hortai ( n = 3), and A. alabamensis ( n = 2) were identified. All strains had MICs of ≥1 mg/liter for amphotericin B, except for two isolates (both A. hortai ) that had MICs of 0.25 mg/liter. Four A. terreus isolates were resistant to at least one azole drug, including one with pan-azole resistance, yet no mutation in the CYP51A gene was found. All strains had low MECs for the three echinocandins. The essential agreements (EAs) between BMD and CGS were >90%, except for those of amphotericin B (79.7%) and itraconazole (73.4%). Isolates belonging to the A section Terrei identified in clinical samples show wider species diversity beyond the known A. terreus sensu stricto Azole resistance inside the section Terrei is uncommon and is not related to CYP51A mutations here. Finally, CGS is an interesting alternative for routine antifungal susceptibility testing. Copyright © 2018 American Society for Microbiology.
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García-Álvarez, Laura; Holden, Matthew TG; Lindsay, Heather; Webb, Cerian R; Brown, Derek FJ; Curran, Martin D; Walpole, Enid; Brooks, Karen; Pickard, Derek J; Teale, Christopher; Parkhill, Julian; Bentley, Stephen D; Edwards, Giles F; Girvan, E Kirsty; Kearns, Angela M; Pichon, Bruno; Hill, Robert LR; Larsen, Anders Rhod; Skov, Robert L; Peacock, Sharon J; Maskell, Duncan J; Holmes, Mark A
2011-01-01
Summary Background Animals can act as a reservoir and source for the emergence of novel meticillin-resistant Staphylococcus aureus (MRSA) clones in human beings. Here, we report the discovery of a strain of S aureus (LGA251) isolated from bulk milk that was phenotypically resistant to meticillin but tested negative for the mecA gene and a preliminary investigation of the extent to which such strains are present in bovine and human populations. Methods Isolates of bovine MRSA were obtained from the Veterinary Laboratories Agency in the UK, and isolates of human MRSA were obtained from diagnostic or reference laboratories (two in the UK and one in Denmark). From these collections, we searched for mecA PCR-negative bovine and human S aureus isolates showing phenotypic meticillin resistance. We used whole-genome sequencing to establish the genetic basis for the observed antibiotic resistance. Findings A divergent mecA homologue (mecALGA251) was discovered in the LGA251 genome located in a novel staphylococcal cassette chromosome mec element, designated type-XI SCCmec. The mecALGA251 was 70% identical to S aureus mecA homologues and was initially detected in 15 S aureus isolates from dairy cattle in England. These isolates were from three different multilocus sequence type lineages (CC130, CC705, and ST425); spa type t843 (associated with CC130) was identified in 60% of bovine isolates. When human mecA-negative MRSA isolates were tested, the mecALGA251 homologue was identified in 12 of 16 isolates from Scotland, 15 of 26 from England, and 24 of 32 from Denmark. As in cows, t843 was the most common spa type detected in human beings. Interpretation Although routine culture and antimicrobial susceptibility testing will identify S aureus isolates with this novel mecA homologue as meticillin resistant, present confirmatory methods will not identify them as MRSA. New diagnostic guidelines for the detection of MRSA should consider the inclusion of tests for mecALGA251. Funding Department for Environment, Food and Rural Affairs, Higher Education Funding Council for England, Isaac Newton Trust (University of Cambridge), and the Wellcome Trust. PMID:21641281
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Figueiredo, Agnes Marie Sá; Ferreira, Fabienne Antunes
2014-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important bacterial pathogens based on its incidence and the severity of its associated infections. In addition, severe MRSA infections can occur in hospitalised patients or healthy individuals from the community. Studies have shown the infiltration of MRSA isolates of community origin into hospitals and variants of hospital-associated MRSA have caused infections in the community. These rapid epidemiological changes represent a challenge for the molecular characterisation of such bacteria as a hospital or community-acquired pathogen. To efficiently control the spread of MRSA, it is important to promptly detect the mecA gene, which is the determinant of methicillin resistance, using a polymerase chain reaction-based test or other rapidly and accurate methods that detect the mecA product penicillin-binding protein (PBP)2a or PBP2’. The recent emergence of MRSA isolates that harbour a mecA allotype, i.e., the mecC gene, infecting animals and humans has raised an additional and significant issue regarding MRSA laboratory detection. Antimicrobial drugs for MRSA therapy are becoming depleted and vancomycin is still the main choice in many cases. In this review, we present an overview of MRSA infections in community and healthcare settings with focus on recent changes in the global epidemiology, with special reference to the MRSA picture in Brazil. PMID:24789555
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Brown-Jaque, Maryury; Rodriguez Oyarzun, Lirain; Cornejo-Sánchez, Thais; Martín-Gómez, Maria T; Gartner, Silvia; de Gracia, Javier; Rovira, Sandra; Alvarez, Antonio; Jofre, Joan; González-López, Juan J; Muniesa, Maite
2018-01-01
Cystic fibrosis (CF) is a chronic disease in which the bacterial colonization of the lung is linked to an excessive inflammatory response that leads to respiratory failure. The microbiology of CF is complex. Staphylococcus aureus is the first bacterium to colonize the lungs in 30% of pediatric CF patients, and 80% of adult patients develop a chronic Pseudomonas aeruginosa infection, but other microorganisms can also be found. The use of antibiotics is essential to treat the disease, but antibiotic performance is compromised by resistance mechanisms. Among various mechanisms of transfer of antibiotic resistance genes (ARGs), the recently been reported bacteriophages are the least explored in clinical settings. To determine the role of phages in CF as mobile genetic elements (MGEs) carrying ARGs, we evaluated their presence in 71 CF patients. 71 sputum samples taken from these patients were screened for eight ARGs ( bla TEM , bla CTX-M-1 -group, bla CTX-M-9 -group, bla OXA-48 , bla VIM , mecA , qnrA , and qnrS ) in the bacteriophage DNA fraction. The phages found were also purified and observed by electron microscopy. 32.4% of CF patients harbored ARGs in phage DNA. β-lactamase genes, particularly bla VIM and bla TEM , were the most prevalent and abundant, whereas mecA , qnrA , and qnrS were very rare. Siphoviridae phage particles capable of infecting P. aeruginosa and Klebsiella pneumoniae were detected in CF sputum. Phage particles harboring ARGs were found to be abundant in the lungs of both CF patients and healthy individuals and could contribute to the colonization of multiresistant strains.
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Sakoulas, George; Gold, Howard S.; Venkataraman, Lata; DeGirolami, Paola C.; Eliopoulos, George M.; Qian, Qinfang
2001-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) is responsible for an increasing number of serious nosocomial and community-acquired infections. Phenotypic heterogeneous drug resistance (heteroresistance) to antistaphylococcal beta-lactams affects the results of susceptibility testing. The present study compared the MRSA-Screen latex agglutination test (Denka Seiken Co., Ltd., Tokyo, Japan) for detection of PBP 2a with agar dilution, the VITEK-1 and VITEK-2 systems (bioMérieux, St. Louis, Mo.), and the oxacillin agar screen test for detection of MRSA, with PCR for the mecA gene used as the “gold standard” assay. Analysis of 107 methicillin-susceptible S. aureus (MSSA) isolates and 203 MRSA isolates revealed that the MRSA-Screen latex agglutination test is superior to any single phenotype-based susceptibility testing method, with a sensitivity of 100% and a specificity of 99.1%. Only one isolate that lacked mecA was weakly positive by the MRSA-Screen latex agglutination test. This isolate was phenotypically susceptible to oxacillin and did not contain the mecA gene by Southern blot hybridization. The oxacillin agar screen test, the VITEK-1 system, the VITEK-2 system, and agar dilution showed sensitivities of 99.0, 99.0, 99.5, and 99%, respectively, and specificities of 98.1, 100, 97.2, and 100%, respectively. The differences in sensitivity or specificity were not statistically significant. Oxacillin bactericidal assays showed that mecA- and PBP 2a-positive S. aureus isolates that are susceptible to antistaphylococcal beta-lactams by conventional methods are functionally resistant to oxacillin. We conclude that the accuracy of the MRSA-Screen latex agglutination method for detection of PBP 2a approaches the accuracy of PCR and is more accurate than any susceptibility testing method used alone for the detection of MRSA. PMID:11682512
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Coagulase-Negative Staphylococci in Human Milk From Mothers of Preterm Compared With Term Neonates.
Soeorg, Hiie; Metsvaht, Tuuli; Eelmäe, Imbi; Metsvaht, Hanna Kadri; Treumuth, Sirli; Merila, Mirjam; Ilmoja, Mari-Liis; Lutsar, Irja
2017-05-01
Human milk is the preferred nutrition for neonates and a source of bacteria. Research aim: The authors aimed to characterize the molecular epidemiology and genetic content of staphylococci in the human milk of mothers of preterm and term neonates. Staphylococci were isolated once per week in the 1st month postpartum from the human milk of mothers of 20 healthy term and 49 preterm neonates hospitalized in the neonatal intensive care unit. Multilocus variable-number tandem-repeats analysis and multilocus sequence typing were used. The presence of the mecA gene, icaA gene of the ica-operon, IS 256, and ACME genetic elements was determined by PCR. The human milk of mothers of preterm compared with term neonates had higher counts of staphylococci but lower species diversity. The human milk of mothers of preterm compared with term neonates more often contained Staphylococcus epidermidis mecA (32.7% vs. 2.6%), icaA (18.8% vs. 6%), IS 256 (7.9% vs. 0.9%), and ACME (15.4% vs. 5.1%), as well as Staphylococcus haemolyticus mecA (90.5% vs. 10%) and IS 256 (61.9% vs. 10%). The overall distribution of multilocus variable-number tandem-repeats analysis (MLVA) types and sequence types was similar between the human milk of mothers of preterm and term neonates, but a few mecA-IS 256-positive MLVA types colonized only mothers of preterm neonates. Maternal hospitalization within 1 month postpartum and the use of an arterial catheter or antibacterial treatment in the neonate increased the odds of harboring mecA-positive staphylococci in human milk. Limiting exposure of mothers of preterm neonates to the hospital could prevent human milk colonization with more pathogenic staphylococci.
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Weterings, Veronica; Bosch, Thijs; Witteveen, Sandra; Landman, Fabian; Schouls, Leo; Kluytmans, Jan
2017-09-01
Resistance to methicillin in Staphylococcus aureus is caused primarily by the mecA gene, which is carried on a mobile genetic element, the staphylococcal cassette chromosome mec (SCC mec ). Horizontal transfer of this element is supposed to be an important factor in the emergence of new clones of methicillin-resistant Staphylococcus aureus (MRSA) but has been rarely observed in real time. In 2012, an outbreak occurred involving a health care worker (HCW) and three patients, all carrying a fusidic acid-resistant MRSA strain. The husband of the HCW was screened for MRSA carriage, but only a methicillin-susceptible S. aureus (MSSA) strain, which was also resistant to fusidic acid, was detected. Multiple-locus variable-number tandem-repeat analysis (MLVA) typing showed that both the MSSA and MRSA isolates were MT4053-MC0005. This finding led to the hypothesis that the MSSA strain acquired the SCC mec and subsequently caused an outbreak. To support this hypothesis, next-generation sequencing of the MSSA and MRSA isolates was performed. This study showed that the MSSA isolate clustered closely with the outbreak isolates based on whole-genome multilocus sequence typing and single-nucleotide polymorphism (SNP) analysis, with a genetic distance of 17 genes and 44 SNPs, respectively. Remarkably, there were relatively large differences in the mobile genetic elements in strains within and between individuals. The limited genetic distance between the MSSA and MRSA isolates in combination with a clear epidemiologic link supports the hypothesis that the MSSA isolate acquired a SCC mec and that the resulting MRSA strain caused an outbreak. Copyright © 2017 American Society for Microbiology.
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Pardos de la Gandara, Maria; Borges, Vitor; Chung, Marilyn; Milheiriço, Catarina; Gomes, João Paulo; de Lencastre, Herminia; Tomasz, Alexander
2018-06-01
Methicillin-resistant Staphylococcus aureus (MRSA) strains carry either a mecA - or a mecC -mediated mechanism of resistance to beta-lactam antibiotics, and the phenotypic expression of resistance shows extensive strain-to-strain variation. In recent communications, we identified the genetic determinants associated with the stringent stress response that play a major role in the antibiotic resistant phenotype of the historically earliest "archaic" clone of MRSA and in the mecC -carrying MRSA strain LGA251. Here, we sought to test whether or not the same genetic determinants also contribute to the resistant phenotype of highly and homogeneously resistant (H*R) derivatives of a major contemporary MRSA clone, USA300. We found that the resistance phenotype was linked to six genes ( fruB , gmk , hpt , purB , prsA , and relA ), which were most frequently targeted among the analyzed 20 H*R strains (one mutation per clone in 19 of the 20 H*R strains). Besides the strong parallels with our previous findings (five of the six genes matched), all but one of the repeatedly targeted genes were found to be linked to guanine metabolism, pointing to the key role that this pathway plays in defining the level of antibiotic resistance independent of the clonal type of MRSA. Copyright © 2018 American Society for Microbiology.
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Sivasailam, Asok; Sasidharan, Suchithra; Kollannur, Justin Davis; Syam, Radhika
2017-01-01
Dairy cows affected with subclinical mastitis can be sources of virulent, antimicrobial-resistant Staphylococci to humans because of the excretion of the bacteria through their milk. This study focussed on the phenotypic and genotypic antibiotic resistance patterns of Staphylococci isolated from dairy cows in early dry period. Among 96 isolates of Gram positive cocci from 157 cows, 76 were identified as Coagulase Negative Staphylococci and the remaining 20 were Staphylococcus aureus. Typical amplicons of coagulase gene were obtained for all 20 samples of S. aureus with three major coagulase types being identified as giving 627 bp (40%), 910 bp (35%) and 710 bp (25%) long PCR products. The groEL gene was amplified in PCR of all 76 isolates of Coagulase Negative Staphylococci, and incubation of PCR products with restriction enzyme PvuII yielded three distinct PCR-RFLP fragment patterns bearing resemblance to S. chromogenes and S. hyicus. Highest sensitivity of Coagulase Negative Staphylococci was noted for Azithromycin (92.5%) and the least to Tetracyclines (76.3%), whereas for S. aureus, it was Cefoperazone (95%) and Azithromycin (72.2%) respectively. Phenotypic resistance to Oxacillin (25 isolates), and Cefoxitin (11 isolates) was detected by dilution method with a commercial strip (Ezy MICTM). Genotypic resistance to β-Lactam antibiotics was found in 65 (34 with mecA gene and 31 with blaZ gene) isolates. Eighteen isolates possessed both the genes, with the PVL gene for virulence being detected in five of them. Nine isolates which had mecA gene were phenotypically susceptible to oxacillin while phenotypic resistance to oxacillin was observed in seven isolates that did not have either mecA or blaZ gene. This is the first report of persistent Staphylococcal infections possessing PVL gene and high level of genotypic resistance to β-Lactam antibiotics in small- holder dairy cattle from India. PMID:29091956
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Siebel, Sabine; Dammann, Claudia; Sanz Miguel, Pablo J; Drewello, Thomas; Kampf, Gunnar; Teubner, Natascha; Bednarski, Patrick J; Freisinger, Eva; Lippert, Bernhard
2015-12-01
Mono(nucleobase) complexes of the general composition cis-[PtCl2 (NH3 )L] with L=1-methylcytosine, 1-MeC (1 a) and L=1-ethyl-5-methylcytosine, as well as trans-[PtX2 (NH3 )(1-MeC)] with X=I (5 a) and X=Br (5 b) have been isolated and were characterized by X-ray crystallography. The Pt coordination occurs through the N3 atom of the cytosine in all cases. The diaqua complexes of compounds 1 a and 5 a, cis-[Pt(H2 O)2 (NH3 )(1-MeC)](2+) and trans-[Pt(H2 O)2 (NH3 )(1-MeC)](2+) , display a rich chemistry in aqueous solution, which is dominated by extensive condensation reactions leading to μ-OH- and μ-(1-MeC(-) -N3,N4)-bridged species and ready oxidation of Pt to mixed-valence state complexes as well as diplatinum(III) compounds, one of which was characterized by X-ray crystallography: h,t-[{Pt(NH3 )2 (OH)(1-MeC(-) -N3,N4)}2 ](NO3 )2 ⋅2 [NH4 ](NO3 )⋅2 H2 O. A combination of (1) H NMR spectroscopy and ESI mass spectrometry was applied to identify some of the various species present in solution and the gas phase, respectively. As it turned out, mass spectrometry did not permit an unambiguous assignment of the structures of +1 cations due to the possibilities of realizing multiple bridging patterns in isomeric species, the occurrence of different tautomers, and uncertainties regarding the Pt oxidation states. Additionally, compound 1 a was found to have selective and moderate antiproliferative activity for a human cervix cancer line (SISO) compared to six other human cancer cell lines. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Kinnevey, Peter M.; Shore, Anna C.; Brennan, Grainne I.; Sullivan, Derek J.; Ehricht, Ralf; Monecke, Stefan; Slickers, Peter
2013-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) has been a major cause of nosocomial infection in Irish hospitals for 4 decades, and replacement of predominant MRSA clones has occurred several times. An MRSA isolate recovered in 2006 as part of a larger study of sporadic MRSA exhibited a rare spa (t878) and multilocus sequence (ST779) type and was nontypeable by PCR- and DNA microarray-based staphylococcal cassette chromosome mec (SCCmec) element typing. Whole-genome sequencing revealed the presence of a novel 51-kb composite island (CI) element with three distinct domains, each flanked by direct repeat and inverted repeat sequences, including (i) a pseudo SCCmec element (16.3 kb) carrying mecA with a novel mec class region, a fusidic acid resistance gene (fusC), and two copper resistance genes (copB and copC) but lacking ccr genes; (ii) an SCC element (17.5 kb) carrying a novel ccrAB4 allele; and (iii) an SCC element (17.4 kb) carrying a novel ccrC allele and a clustered regularly interspaced short palindromic repeat (CRISPR) region. The novel CI was subsequently identified by PCR in an additional 13 t878/ST779 MRSA isolates, six from bloodstream infections, recovered between 2006 and 2011 in 11 hospitals. Analysis of open reading frames (ORFs) carried by the CI showed amino acid sequence similarity of 44 to 100% to ORFs from S. aureus and coagulase-negative staphylococci (CoNS). These findings provide further evidence of genetic transfer between S. aureus and CoNS and show how this contributes to the emergence of novel SCCmec elements and MRSA strains. Ongoing surveillance of this MRSA strain is warranted and will require updating of currently used SCCmec typing methods. PMID:23147725
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Platinum Group Metal-free Catalysts for Hydrogen Evolution Reaction in Microbial Electrolysis Cells.
Yuan, Heyang; He, Zhen
2017-07-01
Hydrogen gas is a green energy carrier with great environmental benefits. Microbial electrolysis cells (MECs) can convert low-grade organic matter to hydrogen gas with low energy consumption and have gained a growing interest in the past decade. Cathode catalysts for the hydrogen evolution reaction (HER) present a major challenge for the development and future applications of MECs. An ideal cathode catalyst should be catalytically active, simple to synthesize, durable in a complex environment, and cost-effective. A variety of noble-metal free catalysts have been developed and investigated for HER in MECs, including Nickel and its alloys, MoS 2 , carbon-based catalysts and biocatalysts. MECs in turn can serve as a research platform to study the durability of the HER catalysts. This personal account has reviewed, analyzed, and discussed those catalysts with an emphasis on synthesis and modification, system performance and potential for practical applications. It is expected to provide insights into the development of HER catalysts towards MEC applications. © 2017 The Chemical Society of Japan & Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
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NASA Astrophysics Data System (ADS)
Xu, Shoutao
Microbial electrochemical systems (MESs) have attracted much research attention in recent years due to their promising applications in renewable energy generation, bioremediation, and wastewater treatment. In a MES, microorganisms interact with electrodes via electrons, catalyzing oxidation and reduction reactions at the anode and the cathode. The bacterial community of a high power mixed consortium MESs (maximum power density is 6.5W/m2) was analyzed by using denature gradient gel electrophoresis (DGGE) and 16S DNA clone library methods. The bacterial DGGE profiles were relatively complex (more than 10 bands) but only three brightly dominant bands in DGGE results. These results indicated there are three dominant bacterial species in mixed consortium MFCs. The 16S DNA clone library method results revealed that the predominant bacterial species in mixed culture is Geobacter sp (66%), Arcobacter sp and Citrobacter sp. These three bacterial species reached to 88% of total bacterial species. This result is consistent with the DGGE result which showed that three bright bands represented three dominant bacterial species. Exoelectrogenic bacterial strain SX-1 was isolated from a mediator-less microbial fuel cell by conventional plating techniques with ferric citrate as electron acceptor under anaerobic conditions. Phylogenetic analysis of the 16S rDNA sequence revealed that it was related to the members of Citrobacter genus with Citrobacter sp. sdy-48 being the most closely related species. The bacterial strain SX-1 produced electricity from citrate, acetate, glucose, sucrose, glycerol, and lactose in MFCs with the highest current density of 205 mA/m2 generated from citrate. Cyclic voltammetry analysis indicated that membrane associated proteins may play an important role in facilitating electron transfer from the bacteria to the electrode. This is the first study that demonstrates that Citrobacter species can transfer electrons to extracellular electron acceptors. Citrobacter strain SX-1 is capable of generating electricity from a wide range of substrates in MFCs. This finding increases the known diversity of power generating exoelectrogens and provids a new strain to explore the mechanisms of extracellular electron transfer from bacteria to electrode. The wide range of substrate utilization by SX-1 increases the application potential of MFCs in renewable energy generation and waste treatment. Anode properties are critical for the performance of microbial electrolysis cells (MECs). Inexpensive Fe nanoparticle modified graphite disks were used as anodes to preliminarily investigate the effects of nanoparticles on the performance of Shewanella oneidensis MR-1 in MECs. Results demonstrated that average current densities produced with Fe nanoparticle decorated anodes were up to 5.9-fold higher than plain graphite anodes. Whole genome microarray analysis of the gene expression showed that genes encoding biofilm formation were significantly up-regulated as a response to nanoparticle decorated anodes. Increased expression of genes related to nanowires, flavins and c-type cytochromes indicate that enhanced mechanisms of electron transfer to the anode may also have contributed to the observed increases in current density. The majority of the remaining differentially expressed genes were associated with electron transport and anaerobic metabolism demonstrating a systemic response to increased power loads. The carbon nanotube (CNT) is another form of nano materials. Carbon nanotube (CNT) modified graphite disks were used as anodes to investigate the effects of nanostructures on the performance S. oneidensis MR-1 in microbial electrolysis cells (MECs). The current densities produced with CNT decorated anodes were up to 5.6-fold higher than plain graphite anodes. Global transcriptome analysis showed that cytochrome c genes associated with extracellular electron transfer are up-expressed by CNT decorated anodes, which is the leading factor to contribute current increase in CNT decorated anode MECs. The up regulated genes encoded to flavin also contribute to current enhancement in CNT decorated anode MECs.
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Wang, Yi; Yan, Weiqiang; Fu, Shanshan; Hu, Shoukui; Wang, Yan; Xu, Jianguo; Ye, Changyun
2018-01-01
Staphylococcus aureus ( S. aureus ), including methicillin-resistant S. aureus (MRSA), is one of the most important human pathogens, which is responsible for bacteremia, soft-tissue infections, and food poisoning. Hence, multiple cross displacement amplification (MCDA) is employed to detect all S. aureus strains, and differentiates MRSA from methicillin-sensitive S. aureus . Multiplex MCDA (m-MCDA), which targets the nuc gene ( S. aureus -specific gene) and mecA gene (encoding penicillin-binding protein-2'), could detect S. aureus strains and identify MRSA within 85 min. Detection of the m-MCDA products is achieved using disposable lateral flow biosensors. A total of 58 strains, including various species of Gram-positive and Gram-negative strains, are used for evaluating and optimizing m-MCDA assays. The optimal amplification condition is found to be 63°C for 40 min, with detection limits at 100 fg DNA/reaction for nuc and mecA genes in the pure cultures, and 10 CFU/tube for nuc and mecA genes in the blood samples. The analytical specificity of m-MCDA assay is of 100%, and no cross-reactions to non- S. aureus strains are produced according to the specificity testing. Particularly, two additional components, including AUDG enzyme and dUTP, are added into the m-MCDA amplification mixtures, which are used for eliminating the unwanted results arising from carryover contamination. Thus, the m-MCDA technique appears to be a simple, rapid, sensitive, and reliable assay to detect all S. aureus strains, and identify MRSA infection for appropriate antibiotic therapy.
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Srednik, Mariela Elizabeth; Grieben, Mario Andres; Bentancor, Adriana; Gentilini, Elida Raquel
2015-09-27
Bovine mastitis is a frequent cause of economic loss in dairy herds. Coagulase-negative staphylococci (CoNS) are increasing in importance as causeof bovine intramammary infection (IMI) throughout the world in recent years. CoNShave been isolated from milk samples collected from cows with clinical andsubclinical mastitis in several countries. Identification of mastitis pathogensis important when selection appropriate antimicrobial therapy. A total of 93 strains of Staphylococcusspp isolated from bovine clinical andsubclinical mastitis in Argentina during 2010-2013 were identified by Restriction Fragment Length Polymorphism (RFLP)-PCR using the gap gene. The isolates were tested by PCR for the presence of blaZ gene and mecA gene and were tested by disk diffusion for the susceptibilityto penicillin and cefoxitin. The most common CoNS species was S.chromogenes 46.2% (43/93), followed by S. devriesei 11.8% (11/93) and S. haemolyticus 9.7% (9/93). The blaZ gene was detected in 19 (20.4%) but only 16 (17.2%) isolates were resistant to penicillin; the mecA was detected in6 (6.5%) isolates but only 4 (4.3) were resistant to cefoxitin. The 6 mecA-positive isolates showed oxaxillinMICs ≤ 0.5 μg/ml. CoNSare important minor mastitis pathogens and can be the cause of substantial economic losses. Despite the low resistance to PEN in Argentina, the presenceof MR isolates found in this study emphasize the importance of identificationof CoNS when an IMI is present because of the potentially risk of lateraltransfer of resistance genes between staphylococcal species.
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Ali Mirani, Zulfiqar; Khan, Muhammad Naseem; Siddiqui, Anila; Khan, Fouzia; Aziz, Mubashir; Naz, Shagufta; Ahmed, Ayaz; Khan, Seema Ismat
2018-02-01
Staphylococcus aureus is a Gram-positive pathogen, well known for its resistance and versatile lifestyle. Under unfavourable conditions, it adapts biofilm mode of growth. For staphylococcal biofilm formation, production of extracellular polymeric substances (EPS) is a pre-requisite, which is regulated by ica operon-encoded enzymes. This study was designed to know the impact of ascorbic acid on biofilm formation and colony spreading processes of S. aureus and MRSA. The isolates of methicillin-resistant S. aureus (MRSA) used in present study, were recovered from different food samples. Various selective and differential media were used for identification and confirmation of S. aureus . Agar dilution method was used for determination of oxacillin and ascorbic acid resistance level. MRSA isolates were re-confirmed by E-test and by amplification of mecA gene. Tube methods and Congo-Red agar were used to study biofilm formation processes. Gene expression studies were carried on real-time reverse transcriptase-polymerase chain reaction (RT-PCR). The results revealed the presence of mecA gene belonging to SCC mecA type IV along with agr type II in the isolates. In vitro studies showed the sub-inhibitory concentration of oxacillin induced biofilm production. However, addition of sub-inhibitory dose of ascorbic acid was found to inhibit EPS production, biofilm formation and augment colony spreading on soft agar plates. The inhibition of biofilm formation and augmentation of colony spreading observed with ascorbic acid alone or in combination with oxacillin. Moreover, gene expression studies showed that ascorbic acid increases agr expression and decreases icaA gene expression. The present study concluded that ascorbic acid inhibits biofilm formation, promotes colony spreading and increases agr gene expression in MRSA.
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Goh, Shan; Loeffler, Anette; Lloyd, David H; Nair, Sean P; Good, Liam
2015-11-11
Antibiotic resistance genes can be targeted by antisense agents, which can reduce their expression and thus restore cellular susceptibility to existing antibiotics. Antisense inhibitors can be gene and pathogen specific, or designed to inhibit a group of bacteria having conserved sequences within resistance genes. Here, we aimed to develop antisense peptide nucleic acids (PNAs) that could be used to effectively restore susceptibility to β-lactams in methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Staphylococcus pseudintermedius (MRSP). Antisense PNAs specific for conserved regions of the mobilisable gene mecA, and the growth essential gene, ftsZ, were designed. Clinical MRSA and MRSP strains of high oxacillin resistance were treated with PNAs and assayed for reduction in colony forming units on oxacillin plates, reduction in target gene mRNA levels, and cell size. Anti-mecA PNA at 7.5 and 2.5 μM reduced mecA mRNA in MRSA and MRSP (p < 0.05). At these PNA concentrations, 66 % of MRSA and 92 % of MRSP cells were killed by oxacillin (p < 0.01). Anti-ftsZ PNA at 7.5 and 2.5 μM reduced ftsZ mRNA in MRSA and MRSP, respectively (p ≤ 0.05). At these PNA concentrations, 86 % of MRSA cells and 95 % of MRSP cells were killed by oxacillin (p < 0.05). Anti-ftsZ PNAs resulted in swelling of bacterial cells. Scrambled PNA controls did not affect MRSA but sensitized MRSP moderately to oxacillin without affecting mRNA levels. The antisense PNAs effects observed provide in vitro proof of concept that this approach can be used to reverse β-lactam resistance in staphylococci. Further studies are warranted as clinical treatment alternatives are needed.
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Ünal, Nilgün; Askar, Şinasi; Macun, Hasan Ceyhun; Sakarya, Fatma; Altun, Belgin; Yıldırım, Murat
2012-03-01
The aims of this study were to determine the existence of pvl gene, some toxin genes, and mecA gene in Staphylococcus aureus strains isolated from sheep milk and to examine antimicrobial resistance profiles in staphylococci from sheep and goats' milk. The milk samples were collected from 13 different small ruminant farms in Kirikkale province from February to August 2009. A total of 1,604 half-udder milk samples from 857 ewes and 66 half-udder milk samples from 33 goats were collected. Staphylococcus spp. were isolated and identified from the samples. Toxin genes and mecA gene among S. aureus strains were determined by PCR. Antimicrobial susceptibility of staphylococci was examined by the disk diffusion method on Mueller-Hinton agar, and interpreted according to the Clinical Laboratory Standards Institute (CLSI) guidelines. The prevalence of subclinical intramammary infection in both ewes and goats was 5.2%. The most prevalent subclinical mastitis agents were coagulase-negative staphylococci and S. aureus with prevalences 2.8% (n:46) and 1.3% (n = 21), respectively. The prevalence of resistances in isolated Staphylococcus spp. to penicilin G, tetracycline, erythromycin, gentamicin, and enrofloxacin were found as 26.9% (18), 7.5% (5), 6.0% (4), 3.0% (2), and 1.5% (1), respectively. Only 3 of the 21 S. aureus ewe isolates (13.4%) were shown to harbor enterotoxin genes being either seh, sej or sec. However, fourteen (66.6%) of the 21 S. aureus isolates had pvl gene while none of the isolates harbored mecA gene. In conclusion, Staphylococci were shown to be the most prevalent bacteria isolated from subclinical mastitis of ewes and goats and these isolates were susceptible to most of the antibiotics. In addition, S. aureus strains isolated from ewes were harboring few staphylococcal enterotoxin genes. However, Panton-Valentine leukocidin produced by S. aureus could be an important virulence factor and contribute to subclinical mastitis pathogenicity.
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Mat Azis, Norhidayah; Pung, Hui P; Abdul Rachman, Abdul R; Amin Nordin, Syafinaz; Sarchio, Seri N E; Suhaili, Zarizal; Mohd Desa, Mohd N
The aim of the present study was to assess and compare the antimicrobial susceptibility pattern against a panel of antibiotics and molecular and methicillin resistance-associated genotypes of 120 carriage S. aureus isolates previously isolated from a student population at two isolation events within a one-month interval. The antibiotic susceptibility of isolates was determined using the Kirby-Bauer disc-diffusion method (cefoxitin by Etest). The MRSA was screened using polymerase chain reaction for the presence of the mecA gene. The mecA-positive isolates were subjected to staphylococcal cassette chromosome (SCC) mec typing, multilocus sequence typing (MLST) and eBURST analysis. All isolates were characterized for the presence of the Panton-Valentine leukocidin (PVL) gene, an enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) pattern and the spa type. For the two occasions where S. aureus was isolated, the highest frequency of resistance was observed for penicillin (70% and 65%, respectively), with a lower rate against erythromycin and tetracycline (<12%). All isolates were susceptible to ciprofloxacin and gentamycin. As for methicillin resistance, eight isolates had minimum inhibitory concentrations (MIC) of resistant categories, but 10 isolates (8.33%) were positive for the mecA gene. The mecA-positive isolates belonged to SCCmec types I (n=9) and V (n=1). MLST was resolved for only three MRSAs, ST508 (n=1), ST88 (n=1) and ST96 (n=1). The results of the eBURST analysis showed that the MRSA isolates analyzed in the present study were potentially related to MRSA identified in other countries. Approximately half of the persistent S. aureus carriers harbored S. aureus of a similar spa type in the respective individuals during both isolation events. A persistent antimicrobial pattern and limited distinct MRSAs were observed over the short study period. The latter frequently exhibited SCCmec type I, commonly associated with hospital-acquired (HA) characteristics, but further delineation is needed to justify the origins of these bacteria. Copyright © 2016 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.
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Multiplex PCR assay to identify methicillin-resistant Staphylococcus haemolyticus.
Schuenck, Ricardo P; Pereira, Eliezer M; Iorio, Natalia L P; Dos Santos, Kátia R N
2008-04-01
Staphylococcus haemolyticus is the most frequently coagulase-negative Staphylococcus species associated with antimicrobial resistance isolated from nosocomial infections. We developed an accurate and simple multiplex PCR assay to identify methicillin-resistant S. haemolyticus (MRSH) isolates. We designed species-specific primers of the mvaA gene that encodes a 3-hydroxy-3-methylglutaryl coenzyme A involved in the mevalonate pathway of the microorganism. Simultaneously, mecA gene primers of methicillin resistance were also used. The PCR assay was established using 16 strains of different reference Staphylococcus species and validated with a collection of 147 clinical staphylococcal isolates that were also phenotypically characterized. Reliable results for the detection of MRSH isolates were obtained for 100% of the strains evaluated, showing that this PCR assay can be used for the routine microbiology laboratories. This is the first report using species-specific multiplex PCR to detect a single segment of S. haemolyticus associated with a segment of mecA gene.
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Antibiotic resistance assessment in S. aureus strains isolated from raw sheep's milk cheese.
Spanu, V; Virdis, S; Scarano, C; Cossu, F; De Santis, E P L; Cosseddu, A M
2010-06-01
In vitro activities of 16 antibiotics were tested against 36 Staphylococcus aureus (SA) strains isolated from raw sheep's milk cheese from six dairies. The minimum inhibitory concentration (MIC) was determined using a broth microdilution method (CLSI). All 36 isolates were analyzed for the presence of the accessory gene regulator gene, agr (I-IV), and genes encoding resistance to methicillin (mecA), erythromycin (ermA), penicillin (blaZ), and vancomycin (vanA-B). The isolates were also analyzed for similarities in pulsed-field gel electrophoresis (PFGE) patterns. SA strains showed resistance to ampicillin (36.1%), penicillin (33.3%), tetracycline (11.1%), and cloxacillin (2.8%) but were susceptible (>or=94.4%) to 12 out of 16 tested antimicrobials. The overall susceptibility of the strains to oxacillin, vancomycin, and erythromycin was confirmed by the absence of the mecA, vanA-B, and ermA genes. The PFGE results showed that 32 strains belonged to 10 different clusters (P1-P10) while four strains were untypeable.
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Montpart, Nuria; Rago, Laura; Baeza, Juan A; Guisasola, Albert
2015-01-01
The use of synthetic wastewater containing carbon sources of different complexity (glycerol, milk and starch) was evaluated in single chamber microbial electrolysis cell (MEC) for hydrogen production. The growth of an anodic syntrophic consortium between fermentative and anode respiring bacteria was operationally enhanced and increased the opportunities of these complex substrates to be treated with this technology. During inoculation, current intensities achieved in single chamber microbial fuel cells were 50, 62.5, and 9 A m⁻³ for glycerol, milk and starch respectively. Both current intensities and coulombic efficiencies were higher than other values reported in previous works. The simultaneous degradation of the three complex substrates favored power production and COD removal. After three months in MEC operation, hydrogen production was only sustained with milk as a single substrate and with the simultaneous degradation of the three substrates. The later had the best results in terms of current intensity (150 A m⁻³), hydrogen production (0.94 m³ m⁻³ d⁻¹) and cathodic gas recovery (91%) at an applied voltage of 0.8 V. Glycerol and starch as substrates in MEC could not avoid the complete proliferation of hydrogen scavengers, even under low hydrogen retention time conditions induced by continuous nitrogen sparging.
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Shariati, Laleh; Validi, Majid; Tabatabaiefar, Mohammad Amin; Karimi, Ali; Nafisi, Mohammad Reza
2010-12-01
Methicillin-resistant Staphylococcus aureus (MRSA) is a nosocomial pathogen. Our main objective was to compare oxacillin disk test, oxacillin E-test, and oxacillin agar screen for detection of methicillin resistance in S. aureus, using real-time PCR for mecA as the "gold standard" comparison assay. 196 S. aureus isolates were identified out of 284 Staphylococcus isolates. These isolates were screened for MRSA with several methods: disk diffusion, agar screen (6.0 μg/ml), oxacillin E-test, and real-time PCR for detection of mecA gene. Of the 196 S. aureus isolates tested, 96 isolates (49%) were mecA-positive and 100 isolates (51%) mecA-negative. All methods tested had a statistically significant agreement with real-time PCR. E-test was 100% sensitive and specific for mecA presence. The sensitivity and specificity of oxacillin agar screen method were 98 and 99%, respectively and sensitivity and specificity of oxacillin disk diffusion method were 95 and 93%, respectively. In the present study, oxacillin E-test is proposed as the best phenotypic method. For economic reasons, the oxacillin agar screen method (6.0 μg/ml), which is suitable for the detection of MRSA, is recommended due to its accuracy and low cost.
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Víquez-Molina, Gerardo; Aragón-Sánchez, Javier; Pérez-Corrales, Cristian; Murillo-Vargas, Christian; López-Valverde, María Eugenia; Lipsky, Benjamin A
2018-03-01
The aim of this study is to describe the presence of genes encoding for 4 virulence factors (pvl, eta, etb, and tsst), as well as the mecA gene conferring resistance to beta-lactam antibiotics, in patients with diabetes and a staphylococcal foot infection. We have also analyzed whether isolates of Staphylococcus aureus from bone infections have a different profile for these genes compared with those from exclusively soft tissue infections. In this cross-sectional study of a prospectively recruited series of patients admitted to the Diabetic Foot Unit, San Juan de Dios Hospital, San José, Costa Rica with a moderate or severe diabetic foot infection (DFI), we collected samples from infected soft tissue and from bone during debridement. During the study period (June 1, 2014 to May 31, 2016), we treated 379 patients for a DFI. S aureus was isolated from 101 wound samples, of which 43 were polymicrobial infections; we only included the 58 infections that were monomicrobial S aureus for this study. Infections were exclusively soft tissue in 17 patients (29.3%) while 41 (70.7%) had bone involvement (osteomyelitis). The mecA gene was detected in 35 cases (60.3%), pvl gene in 4 cases (6.9%), and tsst gene in 3 (5.2%). We did not detect etA and etB in any of the cases. There were no differences in the profile of S aureus genes encoding for virulence factors (pvl, etA, etB, and tsst) recovered from DFIs between those with just soft tissue compared to those with osteomyelitis. However, we found a significantly higher prevalence of pvl+ strains of S aureus associated with soft tissue compared with bone infections. Furthermore, we observed a significantly longer time to healing among patients infected with mecA+ (methicillin-resistant) S aureus (MRSA).
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Diagnosis of bacteremia in pediatric oncologic patients by in-house real-time PCR.
Quiles, Milene Gonçalves; Menezes, Liana Carballo; Bauab, Karen de Castro; Gumpl, Elke Kreuscher; Rocchetti, Talita Trevizani; Palomo, Flavia Silva; Carlesse, Fabianne; Pignatari, Antonio Carlos Campos
2015-07-23
Infections are the major cause of morbidity and mortality in children with cancer. Gaining a favorable prognosis for these patients depends on selecting the appropriate therapy, which in turn depends on rapid and accurate microbiological diagnosis. This study employed real-time PCR (qPCR) to identify the main pathogens causing bloodstream infection (BSI) in patients treated at the Pediatric Oncology Institute IOP-GRAACC-UNIFESP-Brazil. Antimicrobial resistance genes were also investigated using this methodology. A total of 248 samples from BACTEC® blood culture bottles and 99 whole-blood samples collected in tubes containing EDTA K2 Gel were isolated from 137 patients. All samples were screened by specific Gram probes for multiplex qPCR. Seventeen sequences were evaluated using gender-specific TaqMan probes and the resistance genes bla SHV, bla TEM, bla CTX, bla KPC, bla IMP, bla SPM, bla VIM, vanA, vanB and mecA were detected using the SYBR Green method. Positive qPCR results were obtained in 112 of the blood culture bottles (112/124), and 90 % agreement was observed between phenotypic and molecular microbial detection methods. For bacterial and fungal identification, the performance test showed: sensitivity 87 %; specificity 91 %; NPV 90 %; PPV 89 % and accuracy of 89 % when compared with the phenotypic method. The mecA gene was detected in 37 samples, extended-spectrum β-lactamases were detected in six samples and metallo-β-lactamase coding genes in four samples, with 60 % concordance between the two methods. The qPCR on whole blood detected eight samples possessing the mecA gene and one sample harboring the vanB gene. The bla KPC, bla VIM, bla IMP and bla SHV genes were not detected in this study. Real-time PCR is a useful tool in the early identification of pathogens and antimicrobial resistance genes from bloodstream infections of pediatric oncologic patients.
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Yadav, Poonam; Kumar, Parveen; Mukesh, Manishi; Kataria, R S; Yadav, Anita; Mohanty, A K; Mishra, B P
2015-04-01
Expression patterns of lipogenic genes (LPL, ABCG2, ACSS2, ACACA, SCD, BDH, LIPIN1, SREBF1, PPARα and PPARγ) were studied in milk purified MEC across different stages of lactation (15, 30, 45, 60, 90, 120 and 240 days relative to parturition) in buffalo. PPARα was the most abundant gene while ABCG2 and ACSS2 had moderate level of expression; whereas expression of SREBF and PPARγ was very low. The expression patterns of some genes (BDH1, ACSS2, and LIPIN1) across lactation were positively correlated with milk yield while negatively correlated with fat yield. SCD also showed weak correlation with milk yield (p, 0.53) and fat yield (p, -0.47). On the other hand, expression pattern of ACACA was negatively correlated with milk yield (p, -0.88) and positively correlated with fat yield (p, 0.62). Strong correlation was observed between genes involved in de novo milk fat synthesis (BDH1, ACSS2, LIPIN2 and SCD) and milk yield. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Fowoyo, P T; Ogunbanwo, S T
2017-01-31
Coagulase-negative staphylococci have become increasingly recognized as the etiological agent of some infections. A significant characteristic of coagulase-negative staphylococci especially strains isolated from animals and clinical samples is their resistance to routinely used antibiotics although, resistant strains isolated from fermented foods have not been fully reported. A total of two hundred and fifty-five CoNS isolates were subjected to antimicrobial susceptibility test using the disc diffusion technique. The minimum inhibitory concentration of the isolates to the tested antibiotics was determined using the microbroth dilution method. Methicillin resistant strains were confirmed by detection of methicillin resistant genes (mecA) and also employing cefoxitin screening test. The isolates were confirmed to be methicillin resistant by the detection of mecA genes and the cefoxitin screening test. The isolates demonstrated appreciable resistance to ampicillin (86.7%), sulfomethoxazole-trimethoprim (74.9%), amoxicillin-clavulanic acid (52.5%) and oxacillin (35.7%). Methicillin resistance was exhibited by 13 out of the 255 isolates although no mecA gene was detected. It was also observed that the methicillin resistant isolates were prevalent in these traditional foods; iru, kindirmo, nono and wara. This study has ameliorated the incidence of multiple antibiotic resistant coagulase-negative staphylococci in Nigerian fermented foods and if not tackled adequately might lead to horizontal transfer of antibiotic resistance from food to man.
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Zhang, Yan-Li; Wan, Yong-Jie; Wang, Zi-Yu; Qi, Wei-Wei; Zhou, Zheng-Rong; Huang, Rong; Wang, Feng
2010-05-07
Nuclear transfer using transgenic donor cells is an efficient way of generating transgenic goats, wherein the preparation of competent transgenic donor cells is the pivotal upstream step. We have measured the efficiency of transfection with a plasmid containing hGCase (human lysosomal acid beta-glucosidase) gene into goat FFC (fetal-derived fibroblast cells), MEC (mammary epithelial cells) and AEFC (adult ear skin-derived fibroblast cells), and the characteristics of cell cycle, apoptosis and chromosome abnormalities after transfection. The expression of genes involved in imprinting [IGF2 (insulin-like growth factor 2), IGF2R (IGF2 receptor)], apoptosis (Bax), stress (heat-shock protein, Hsp70.1), cellular connections [Cx43 (connexin 43)] and DNA methylation [DNMT1 (DNA methyltransferase 1)] in transgenic fetal cells has been investigated. The hGCase transgene was successfully detected in the transfected cell lines, and chromosomal stability remained similar in FFC and transgenic FFC (70.9 compared with 66.8%), whereas a smaller percentage (P<0.05) of cells at G(0)/G(1) in the transgenic FFC, MEC and AEFC (T-FFC, T-MEC and T-AEFC), and higher percentage (P<0.05) of apoptotic cells in T-FFC than the non-transfected controls were detected by flow cytometric analysis. Among the genes tested, the relative expressions of IGF2, IGF2R and transcripts of Cx43 were significantly higher (P<0.05) in T-FFC compared with non-transfected FFC. These novel findings on gene expression in transgenic fetal cells may have certain implications in the biopharming industry and in our understanding the low efficiency of transgenic cloning.
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de Souza, C R; Aragão, F J; Moreira, E C O; Costa, C N M; Nascimento, S B; Carvalho, L J
2009-03-24
Cassava is one of the most important tropical food crops for more than 600 million people worldwide. Transgenic technologies can be useful for increasing its nutritional value and its resistance to viral diseases and insect pests. However, tissue-specific promoters that guarantee correct expression of transgenes would be necessary. We used inverse polymerase chain reaction to isolate a promoter sequence of the Mec1 gene coding for Pt2L4, a glutamic acid-rich protein differentially expressed in cassava storage roots. In silico analysis revealed putative cis-acting regulatory elements within this promoter sequence, including root-specific elements that may be required for its expression in vascular tissues. Transient expression experiments showed that the Mec1 promoter is functional, since this sequence was able to drive GUS expression in bean embryonic axes. Results from our computational analysis can serve as a guide for functional experiments to identify regions with tissue-specific Mec1 promoter activity. The DNA sequence that we identified is a new promoter that could be a candidate for genetic engineering of cassava roots.
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Rigatti, Fabiane; Tizotti, Maísa Kraulich; Hörner, Rosmari; Domingues, Vanessa Oliveira; Martini, Rosiéli; Mayer, Letícia Eichstaedt; Khun, Fábio Teixeira; de França, Chirles Araújo; da Costa, Mateus Matiuzzi
2010-01-01
This study aimed to characterize the prevalence and susceptibility profile to oxacillin-resistant Coagulase-negative Staphylococci strains isolated from blood cultures in a teaching hospital, located in Santa Maria, RS. In addition, different methodologies for phenotypic characterization of mecA-mediated oxacillin resistance were compared with genotypic reference testing. After identification (MicroScan - Siemens), the isolates were tested for antimicrobial sensitivity using disk diffusion and automation (MicroScan - Siemens). The presence of mecA gene was identified by the polymerase chain reaction molecular technique. The most common species was Staphylococcus epidermidis (n=40, 67%). The mecA gene was detected in 54 (90%) strains, while analysis of the sensitivity profiles revealed a high rate of resistance to multiple classes of antimicrobial drugs. However, all isolates were uniformly sensitive to vancomycin and tigecycline. The cefoxitin disk was the phenotypic method that best correlated with the gold standard. Analysis of the clinical significance of CoNS isolated from hemocultures and the precise detection of oxacillin resistance represent decisive factors for the correct choice of antibiotic therapy. Although vancomycin constitutes the normal treatment in most Brazilian hospitals, reduction in its use is recommended.
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Paulovich, A G; Armour, C D; Hartwell, L H
1998-01-01
In wild-type Saccharomyces cerevisiae, a checkpoint slows the rate of progression of an ongoing S phase in response to exposure to a DNA-alkylating agent. Mutations that eliminate S phase regulation also confer sensitivity to alkylating agents, leading us to suggest that, by regulating the S phase rate, cells are either better able to repair or better able to replicate damaged DNA. In this study, we determine the effects of mutations that impair S phase regulation on the ability of excision repair-defective cells to replicate irreparably UV-damaged DNA. We assay survival after UV irradiation, as well as the genetic consequences of replicating a damaged template, namely mutation and sister chromatid exchange induction. We find that RAD9, RAD17, RAD24, and MEC3 are required for UV-induced (although not spontaneous) mutagenesis, and that RAD9 and RAD17 (but not REV3, RAD24, and MEC3) are required for maximal induction of replication-dependent sister chromatid exchange. Therefore, checkpoint genes not only control cell cycle progression in response to damage, but also play a role in accommodating DNA damage during replication. PMID:9725831
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Paulovich, A G; Armour, C D; Hartwell, L H
1998-09-01
In wild-type Saccharomyces cerevisiae, a checkpoint slows the rate of progression of an ongoing S phase in response to exposure to a DNA-alkylating agent. Mutations that eliminate S phase regulation also confer sensitivity to alkylating agents, leading us to suggest that, by regulating the S phase rate, cells are either better able to repair or better able to replicate damaged DNA. In this study, we determine the effects of mutations that impair S phase regulation on the ability of excision repair-defective cells to replicate irreparably UV-damaged DNA. We assay survival after UV irradiation, as well as the genetic consequences of replicating a damaged template, namely mutation and sister chromatid exchange induction. We find that RAD9, RAD17, RAD24, and MEC3 are required for UV-induced (although not spontaneous) mutagenesis, and that RAD9 and RAD17 (but not REV3, RAD24, and MEC3) are required for maximal induction of replication-dependent sister chromatid exchange. Therefore, checkpoint genes not only control cell cycle progression in response to damage, but also play a role in accommodating DNA damage during replication.
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Netsvyetayeva, Irina; Fraczek, Mariusz; Piskorska, Katarzyna; Golas, Marlena; Sikora, Magdalena; Mlynarczyk, Andrzej; Swoboda-Kopec, Ewa; Marusza, Wojciech; Palmieri, Beniamino; Iannitti, Tommaso
2014-03-05
The number of studies regarding the incidence of multidrug resistant strains and distribution of genes encoding virulence factors, which have colonized the post-Soviet states, is considerably limited. The aim of the study was (1) to assess the Staphylococcus (S.) aureus nasal carriage rate, including Methicillin Resistant S. aureus (MRSA) strains in adult Ukrainian population, (2) to determine antibiotic resistant pattern and (3) the occurrence of Panton Valentine Leukocidine (PVL)-, Fibronectin-Binding Protein A (FnBPA)- and Exfoliative Toxin (ET)-encoding genes. Nasal samples for S. aureus culture were obtained from 245 adults. The susceptibility pattern for several classes of antibiotics was determined by disk diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The virulence factor encoding genes, mecA, lukS-lukF, eta, etb, etd, fnbA, were detected by Polymerase Chain Reaction (PCR). The S. aureus nasal carriage rate was 40%. The prevalence of nasal MRSA carriage in adults was 3.7%. LukS-lukF genes were detected in over 58% of the strains. ET-encoding genes were detected in over 39% of the strains and the most prevalent was etd. The fnbA gene was detected in over 59% of the strains. All MRSA isolates tested were positive for the mecA gene. LukS-lukF genes and the etd gene were commonly co-present in MRSA, while lukS-lukF genes and the fnbA gene were commonly co-present in Methicillin Sensitive S. aureus (MSSA) isolates. No significant difference was detected between the occurrence of lukS-lukF genes (P > 0.05) and the etd gene (P > 0.05) when comparing MRSA and MSSA. The occurrence of the fnbA gene was significantly more frequent in MSSA strains (P < 0.05). In Ukraine, S. aureus is a common cause of infection. The prevalence of S. aureus nasal carriage in our cohort of patients from Ukraine was 40.4%. We found that 9.1% of the strains were classified as MRSA and all MRSA isolates tested positive for the mecA gene. We also observed a high prevalence of PVL- and ET- encoding genes among S. aureus nasal carriage strains. A systematic surveillance system can help prevent transmission and spread of drug resistant toxin producing S. aureus strains.
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Comparison of complex effluent treatability in different bench scale microbial electrolysis cells.
Ullery, Mark L; Logan, Bruce E
2014-10-01
A range of wastewaters and substrates were examined using mini microbial electrolysis cells (mini MECs) to see if they could be used to predict the performance of larger-scale cube MECs. COD removals and coulombic efficiencies corresponded well between the two reactor designs for individual samples, with 66-92% of COD removed for all samples. Current generation was consistent between the reactor types for acetate (AC) and fermentation effluent (FE) samples, but less consistent with industrial (IW) and domestic wastewaters (DW). Hydrogen was recovered from all samples in cube MECs, but gas composition and volume varied significantly between samples. Evidence for direct conversion of substrate to methane was observed with two of the industrial wastewater samples (IW-1 and IW-3). Overall, mini MECs provided organic treatment data that corresponded well with larger scale reactor results, and therefore it was concluded that they can be a useful platform for screening wastewater sources. Copyright © 2014 Elsevier Ltd. All rights reserved.
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An in vitro model of murine middle ear epithelium.
Mulay, Apoorva; Akram, Khondoker M; Williams, Debbie; Armes, Hannah; Russell, Catherine; Hood, Derek; Armstrong, Stuart; Stewart, James P; Brown, Steve D M; Bingle, Lynne; Bingle, Colin D
2016-11-01
Otitis media (OM), or middle ear inflammation, is the most common paediatric disease and leads to significant morbidity. Although understanding of underlying disease mechanisms is hampered by complex pathophysiology it is clear that epithelial abnormalities underpin the disease. There is currently a lack of a well-characterised in vitro model of the middle ear (ME) epithelium that replicates the complex cellular composition of the middle ear. Here, we report the development of a novel in vitro model of mouse middle ear epithelial cells (mMECs) at an air-liquid interface (ALI) that recapitulates the characteristics of the native murine ME epithelium. We demonstrate that mMECs undergo differentiation into the varied cell populations seen within the native middle ear. Proteomic analysis confirmed that the cultures secrete a multitude of innate defence proteins from their apical surface. We showed that the mMECs supported the growth of the otopathogen, nontypeable Haemophilus influenzae (NTHi), suggesting that the model can be successfully utilised to study host-pathogen interactions in the middle ear. Overall, our mMEC culture system can help to better understand the cell biology of the middle ear and improve our understanding of the pathophysiology of OM. The model also has the potential to serve as a platform for validation of treatments designed to reverse aspects of epithelial remodelling that underpin OM development. © 2016. Published by The Company of Biologists Ltd.
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Drougka, Eleanna; Foka, Antigoni; Koutinas, Christos K; Jelastopulu, Eleni; Giormezis, Nikolaos; Farmaki, Ourania; Sarrou, Styliani; Anastassiou, Evangelos D; Petinaki, Efthimia; Spiliopoulou, Iris
2016-04-01
Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) prevalence among companion animals and veterinary personnel (VP) was investigated. Strains' molecular characteristics were evaluated in order to assess S. aureus transmission. Specimens (224) from colonized and infected sites of 102 animals (92 dogs, 10 cats) and 18 VP were collected during 2012 and 2013. Antibiotic susceptibility was performed by the disk diffusion method and Etest. mecA, mecC, tst (toxic shock syndrome toxin) and lukF/lukS-PV (Panton-Valentine leukocidin, PVL) genes were investigated by PCR. Genotypes were identified by Multi Locus Sequence Typing (MLST), Staphylococcal Cassette Chromosome mec (SCCmec), accessory gene regulator group (agr), spa and Pulsed Field Gel Electrophoresis (PFGE). S. aureus prevalence among pets and VP was 36.3% (37/102) and 38.9% (7/18), respectively. Younger companion animals, those living in rural areas, having a disease upon admission or Coagulase-negative staphylococci co-carriage showed significantly higher prevalence of S. aureus isolation (p<0.05). Twenty-six pets and five VP carried PVL-positive S. aureus. In total, 60 S. aureus strains were recovered (53 from pets, seven from VP) of which 16 were MRSA (26.7%), 12 mecA- and four mecC-positive. MRSA showed higher resistance rates against other antimicrobials as compared to methicillin-susceptible ones. Strains were classified by MLST in 13 STs, with the predominance of ST80 and ST15. In MRSA, SCCmec types II, IV and XI were identified. The most frequent spa types were t5559 and t7558. Fifty-six strains were classified into 15 PFGE types. Comparison of genetic markers shows that identical or very similar strains disseminate among animals and VP. Companion animals harbor PVL-positive clones constituting a possible source for transmission to humans. Copyright © 2016 Elsevier B.V. All rights reserved.
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Molecular & phenotypic characterization of Staphylococcus epidermidis in implant related infections
Prasad, Sujata; Nayak, N.; Satpathy, G.; Nag, H.L.; Venkatesh, P.; Ramakrishnan, S.; Ghose, Supriyo; Nag, T.C.
2012-01-01
Background & objectives: The discrimination between the Staphylococcus epidermidis colonizing the deep seated indwelling devices and those which are mere commensals has always been a challenge for the clinical microbiologist. This study was aimed to characterize the S. epidermidis isolates obtained from device related infection for their phenotypic and molecular markers of virulence and to see whether these markers can be used to differentiate the pathogenic S. epidermidis from the commensals. Methods: Fifty five S. epidermidis isolates from various device related infections such as endophthalmitis following intra-ocular lens (IOL) implantation, intravascular (IV) catheter related sepsis and orthopaedic implant infections, were studied for slime production, biotyping, antibiotic sensitivity; and mec A and ica positivity by the recommended procedures. Results: Twenty three (41.8%) isolates were multi-drug resistant, 26 (65.2%) were slime producers, 30 (54.5%) were adherent, 23 (41.8%) possessed the intercellular adhesin (ica) gene, and 28 (50.9%) harboured the mec A gene. Biotypes I and III were the commonest, most members of which were multi- drug resistant. Twenty two (73.3%) of the 30 adherent bacteria were slime producers as opposed to only 4 (16%) of the 25 non-adherent bacteria (P<0.001). A vast majority i.e. 21 (91.3%) of the 23 ica positive organisms were adherent to artificial surfaces in contrast to only 9 (28.1%) of the 32 non-ica positive organisms (P<0.001). Twenty (86.9%) of the 23 ica positive bacteria were slime producers, as opposed to only 6 (18.7%) of the 32 ica negative bacteria (P<0.001). Of the 23 multi-drug resistant isolates, 19 (82.6%) carried the mec A gene. Interpretation & conclusions: The present findings showed that ica AB and mec A were the two important virulence markers of S. epidermidis in implant infections and slime was responsible for the sessile mode of attachment on the devices. PMID:23041744
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USDA-ARS?s Scientific Manuscript database
Expression of genes for lipid biosynthetic enzymes during initiation of lactation in humans is unknown. Our objective was to study mRNA expression of lipid metabolic enzymes in human mammary epithelial cell (MEC) in conjunction with the measurement of milk fatty acid (FA) composition during secretor...
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Hau, Samantha J; Frana, Timothy; Sun, Jisun; Davies, Peter R; Nicholson, Tracy L
2017-08-01
Zinc resistance in livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) sequence type 398 (ST398) is primarily mediated by the czrC gene colocated with the mecA gene, encoding methicillin resistance, within the type V staphylococcal cassette chromosome mec (SCC mec ) element. Because czrC and mecA are located within the same mobile genetic element, it has been suggested that the use of zinc in feed as an antidiarrheal agent has the potential to contribute to the emergence and spread of methicillin-resistant S. aureus (MRSA) in swine, through increased selection pressure to maintain the SCC mec element in isolates obtained from pigs. In this study, we report the prevalence of the czrC gene and phenotypic zinc resistance in U.S. swine-associated LA-MRSA ST5 isolates, MRSA ST5 isolates from humans with no swine contact, and U.S. swine-associated LA-MRSA ST398 isolates. We demonstrated that the prevalence of zinc resistance in U.S. swine-associated LA-MRSA ST5 isolates was significantly lower than the prevalence of zinc resistance in MRSA ST5 isolates from humans with no swine contact and swine-associated LA-MRSA ST398 isolates, as well as prevalences from previous reports describing zinc resistance in other LA-MRSA ST398 isolates. Collectively, our data suggest that selection pressure associated with zinc supplementation in feed is unlikely to have played a significant role in the emergence of LA-MRSA ST5 in the U.S. swine population. Additionally, our data indicate that zinc resistance is associated with the multilocus sequence type lineage, suggesting a potential link between the genetic lineage and the carriage of resistance determinants. IMPORTANCE Our data suggest that coselection thought to be associated with the use of zinc in feed as an antimicrobial agent is not playing a role in the emergence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) ST5 in the U.S. swine population. Additionally, our data indicate that zinc resistance is more associated with the multilocus sequence type lineage, suggesting a potential link between the genetic lineage and the carriage of resistance markers. This information is important for public health professionals, veterinarians, producers, and consumers. Copyright © 2017 American Society for Microbiology.
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Save, Etienne; Sargolini, Francesca
2017-01-01
It is now widely accepted that the entorhinal cortex (EC) plays a pivotal role in the processing of spatial information and episodic memory. The EC is segregated into two sub-regions, the medial EC (MEC) and the lateral EC (LEC) but a comprehensive understanding of their roles across multiple behavioral contexts remains unclear. Considering that it is still useful to investigate the impact of lesions of EC on behavior, we review the contribution of lesion approach to our knowledge of EC functions. We show that the MEC and LEC play different roles in the processing of spatial and non-spatial information. The MEC is necessary to the use of distal but not proximal landmarks during navigation and is crucial for path integration, in particular integration of linear movements. Consistent with predominant hypothesis, the LEC is important for combining the spatial and non-spatial aspects of the environment. However, object exploration studies suggest that the functional segregation between the MEC and the LEC is not as clearly delineated and is dependent on environmental and behavioral factors. Manipulation of environmental complexity and therefore of cognitive demand shows that the MEC and the LEC are not strictly necessary to the processing of spatial and non-spatial information. In addition we suggest that the involvement of these sub-regions can depend on the kind of behavior, i.e., navigation or exploration, exhibited by the animals. Thus, the MEC and the LEC work in a flexible manner to integrate the “what” and “where” information in episodic memory upstream the hippocampus. PMID:29163076
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Roth, Fabian C; Beyer, Katinka M; Both, Martin; Draguhn, Andreas; Egorov, Alexei V
2016-12-01
The entorhinal cortex (EC) is a critical component of the medial temporal lobe (MTL) memory system. Local networks within the MTL express a variety of state-dependent network oscillations that are believed to organize neuronal activity during memory formation. The peculiar pattern of sharp wave-ripple complexes (SPW-R) entrains neurons by a very fast oscillation at ∼200 Hz in the hippocampal areas CA3 and CA1 and then propagates through the "output loop" into the EC. The precise mechanisms of SPW-R propagation and the resulting cellular input patterns in the mEC are, however, largely unknown. We therefore investigated the activity of layer V (LV) principal neurons of the medial EC (mEC) during SPW-R oscillations in horizontal mouse brain slices. Intracellular recordings in the mEC were combined with extracellular monitoring of propagating network activity. SPW-R in CA1 were regularly followed by negative field potential deflections in the mEC. Propagation of SPW-R activity from CA1 to the mEC was mostly monosynaptic and excitatory, such that synaptic input to mEC LV neurons directly reflected unit activity in CA1. Comparison with propagating network activity from CA3 to CA1 revealed a similar role of excitatory long-range connections for both regions. However, SPW-R-induced activity in CA1 involved strong recruitment of rhythmic synaptic inhibition and corresponding fast field oscillations, in contrast to the mEC. These differences between features of propagating SPW-R emphasize the differential processing of network activity by each local network of the hippocampal output loop. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.
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Panda, Sasmita; Kar, Sarita; Choudhury, Ranginee; Sharma, Savitri; Singh, Durg V
2014-03-01
We developed a multiplex PCR to detect the presence of methicillin- (mecA), cadmium/zinc-(czrC) and antiseptic-resistant (qacA/B) staphylococci and to identify Panton-Valentine leukocidin (PVL)-positive and -negative Staphylococcus aureus and coagulase-negative staphylococci (CoNS) from infected and healthy eyes. The assay was validated on 177 staphylococci comprising of 55 each of S. aureus and CoNS isolated from infected eyes and five S. aureus and 62 CoNS isolated from healthy eyes and nine direct ocular samples. Nine direct ocular samples for in situ testing consisted of corneal scrapings (4), conjunctiva swabs (2) and others (3). Multiplex PCR result was correlated with genotype data obtained with single PCR and dot-blot assay. The control strains that were positive in multiplex PCR for 16S rRNA, nuc, mecA, pvl, czrC and qacA/B genes were also positive in the dot-blot assay. The specificity of amplified genes obtained with reference strains was further confirmed by DNA sequencing. The single step-hexaplex PCR method can be used for rapid detection of mecA, nuc, pvl, czrC and qacA/B genes in staphylococci with simultaneous identification of PVL-positive and -negative S. aureus and CoNS from a variety of ocular samples. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.
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Ndahi, M D; Kwaga, J K P; Bello, M; Kabir, J; Umoh, V J; Yakubu, S E; Nok, A J
2014-03-01
The bacterial genera Listeria and Staphylococcus have been frequently isolated from food products and are responsible for a number of animal and human diseases. The aim of the study was to simultaneously isolate and characterize L. monocytogenes and Staphylococcus species from 300 samples of raw meat and meat products, to determine the susceptibility of the organisms to commonly used antimicrobial agents and to determine the presence of haemolysin A (hyl) virulence gene in L. monocytogenes and staphylococcal cassette chromosome mecA (SCCmec) gene in the Staph. aureus isolates using PCR. Of the 85 Listeria isolates tested, 12 L. monocytogenes were identified and tested for their sensitivity to 14 antimicrobial agents. All the 12 isolates (100%) were resistant to nine antimicrobial agents, but however sensitive to gentamicin. Only one isolate was found to harbour the hylA gene. Twenty-nine isolates were confirmed as Staph. aureus by the Microbact 12S identification system and were all presumptively identified as methicillin-resistant Staph. aureus species using oxacillin-resistant Staph. aureus basal medium (ORSAB). The 29 Staph. aureus isolates were tested for their sensitivity to 16 antimicrobial agents, and 11 were resistant to methicillin. None of the 11 Staph. aureus isolates harboured the methicillin resistance, mecA gene. Listeria monocytogenes and Staphylococcus aureus are important agents of foodborne diseases. Occurrence of these infectious agents was established in meat and meat products in Zaria, Nigeria. Majority of isolates obtained from this study, displayed multidrug resistance to commonly used antimicrobial agents, including methicillin resistance among the Staph. aureus isolates. The potential virulence of L. monocytogenes found in ready-to-eat food was documented by the carriage of hly A gene by one of the isolates. A different mechanism of methicillin resistance or different homologue of mec A gene may be circulating among Nigerian isolates. © 2013 The Society for Applied Microbiology.
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Awad, Amal; Ramadan, Hazem; Nasr, Sherif; Ateya, Ahmed; Atwa, Samar
2017-01-01
Staphylococcus aureus is commonly associated with mastitis in dairy herds with potential public health implications. This study was conducted to investigate the existence of S. aureus in mastitic milk and to determine the antimicrobial resistance profiles of the isolated strains as well as the resistance and virulence associated genes. Two hundred quarter milk samples were collected from 3 dairy farms at Dakahliya (n = 2) and Damietta (n = 1) Governorates, Egypt from September to December 2016. Conventional culturing and Polymerase Chain Reaction (PCR) assays targeting nuc (thermonuclease) and coa (coagulase) genes were performed. Isolates were tested for its susceptibility against 14 antimicrobial agents using disk diffusion method. All the isolates were screened for the presence of β-lactamases (blaZ, mecA) and virulence associated (pvl and tst) genes by PCR. The S. aureus was detected in 42% (84/200) of the total examined milk samples. Regarding the antibiogram results, S. aureus revealed a high resistance against ampicillin (95.2%) and penicillin (83.3%) and a lower resistance was observed against gentamicin (23.8%), amikacin (16.7%) and ciprofloxacin (14.3%). Multidrug resistances were detected in 83.3% of the isolated S. aureus. Of the 70 penicillin-resistant S. aureus isolates, blaZ gene was identified in 67 (95.7%) isolates. Fifty percent of S. aureus isolates harbored the specific amplicon of mecA gene. Markedly, all mecA positive strains displayed multidrug resistance and were also positive for blaZ gene. The virulence determinants pvl and tst were detected in 7.1 and 11.9% of the isolated S. aureus, respectively. Presence of multidrug resistant and toxin producing S. aureus in dairy farms pose a major risk to public health. Therefore, this study highlighted the importance of developing an efficient control program to inhibit the transmission of S. aureus, particularly multidrug resistant strains to humans.
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Multiplexed in vivo His-tagging of enzyme pathways for in vitro single-pot multienzyme catalysis.
Wang, Harris H; Huang, Po-Yi; Xu, George; Haas, Wilhelm; Marblestone, Adam; Li, Jun; Gygi, Steven P; Forster, Anthony C; Jewett, Michael C; Church, George M
2012-02-17
Protein pathways are dynamic and highly coordinated spatially and temporally, capable of performing a diverse range of complex chemistries and enzymatic reactions with precision and at high efficiency. Biotechnology aims to harvest these natural systems to construct more advanced in vitro reactions, capable of new chemistries and operating at high yield. Here, we present an efficient Multiplex Automated Genome Engineering (MAGE) strategy to simultaneously modify and co-purify large protein complexes and pathways from the model organism Escherichia coli to reconstitute functional synthetic proteomes in vitro. By application of over 110 MAGE cycles, we successfully inserted hexa-histidine sequences into 38 essential genes in vivo that encode for the entire translation machinery. Streamlined co-purification and reconstitution of the translation protein complex enabled protein synthesis in vitro. Our approach can be applied to a growing area of applications in in vitro one-pot multienzyme catalysis (MEC) to manipulate or enhance in vitro pathways such as natural product or carbohydrate biosynthesis.
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Martins Simões, P; Rasigade, J-P; Lemriss, H; Butin, M; Ginevra, C; Lemriss, S; Goering, R V; Ibrahimi, A; Picaud, J C; El Kabbaj, S; Vandenesch, F; Laurent, F
2013-12-01
Multiresistant Staphylococcus capitis pulsotype NRCS-A has been reported to be a major pathogen causing nosocomial bacteremia in preterm infants. We report that the NRCS-A strain CR01 harbors a novel 60.9-kb composite staphylococcal cassette chromosome mec (SCCmec) element, composed of an SCCmec with strong homologies to Staphylococcus aureus ST398 SCCmec and of an SCCcad/ars/cop harboring resistance genes for cadmium, arsenic, and copper. Whole-genome-based comparisons of published S. capitis strains suggest that strain CR01 acquired the two elements independently.
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Correlation of the expression of CD32 and CD180 receptors on CLL cells and MEC1 cell line.
Tsertsvadze, T; Mitskevich, N; Ghirdaladze, D; Porakishvili, N
2015-03-01
Chronic Lymphocytic Leukemia (CLL) presents with clonal expansion and accumulation of CD5+CD19+CD23+ cells in peripheral lymphoid organs and tissues and in bone marrow. CLL is supposedly driven by exogenous and/or endogenous (auto)antigen(s) and there is increasing evidence that CLL cells receive microenvironmental signals which support their growth, survival and expansion in vivo. We have previously shown that powerful signals are received by CLL cells through CD180 orphan toll-like receptor. Additional accessory signals could be generated through FcγRII (CD32), since both are expressed on CLL cells as well as on control B cells. Here we studied correlation of the expression of CD32 and CD180 on CLL cells as well as on MEC1 cell line. Peripheral blood mononuclear cells (PBMC) from CLL patients and age-matched healthy volunteers were separated, stained with appropriate antibodies to CD19, CD32 and CD180 and analysed by flow cytometry. CD32 and CD180 expression on MEC1 cells was studied at different time-points. The data was statistically analysed using the Mann-Whitney non-parametrical test. Our data indicates that expression of CD32 is significantly increased on CLL cells compared to control B cells as well as in long-term MEC1 cell culture. In contrast, CD180 expression on MEC1 cells significantly decreased throughout 0-96h of MEC1 cell culture. We have recently shown that CD180 ligation can redirect sIgM-mediated signaling from pro-survival to pro-apoptotic. This data indicates that a drop in the expression of CD180 on cycling CLL cells might lead to a weakening of this effect and enhance further survival and expansion of CLL cells in proliferative centres of lymphoid tissues. Since MEC1 cells are derived from a CLL patient with mutated IGVH genes (M-CLL) negative correlation between CD180 and CD32 expression on cycling MEC1 cells could be limited to M-CLL.
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Ho, Ya-Chi; Chang, Shan-Chwen; Lin, Su-Ru; Wang, Wei-Kung
2009-01-01
Persistent methicillin-resistant Staphylococcus aureus (MRSA) bacteremia is known to be a poor prognostic factor. While several PCR assays for the detection of MRSA in various clinical samples were recently reported, the possibility that a quantitative PCR assay could be used to quantify and monitor MRSA bacteremia has not been explored. In this study, we established a quantitative real-time PCR assay for the mecA gene using known copy numbers of a plasmid containing mecA DNA as a standard and the previously described mecA-specific primers and probe (P. Francois et al., J. Clin. Microbiol. 41:254-260, 2003). We employed this assay to examine 250 sequential whole-blood samples from 20 adult patients, including 13 survivors and 7 nonsurvivors, with culture-proven MRSA bacteremia at the intensive care units of National Taiwan University Hospital between 1 July 2006 and 31 January 2007. The levels of mecA DNA in the nonsurvivors were significantly higher than those in the survivors during the three periods of bacteremia examined (days 0 to 2, 3 to 5, and 6 to 8) (P = 0.003 by two-tailed Mann-Whitney U test). Moreover, the nonsurvivors had higher mecA DNA levels than the survivors after 3 days and 7 days of anti-MRSA therapy (medians for nonsurvivors and survivors at 3 days, 5.86 and 4.30 log copies/ml, respectively; medians for nonsurvivors and survivors at 7 days, 5.21 and 4.36 log copies/ml, respectively; P = 0.02 and P = 0.04, respectively, by two-tailed Mann-Whitney U test). Together, these findings suggest that the level of mecA DNA in blood could potentially be used to monitor MRSA bacteremia and evaluate responses to therapy. PMID:19279177
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Shariati, Laleh; Validi, Majid; Hasheminia, Ali Mohammad; Ghasemikhah, Reza; Kianpour, Fariborz; Karimi, Ali; Nafisi, Mohammad Reza; Tabatabaiefar, Mohammad Amin
2016-01-01
A diversity of virulence factors work together to create the pathogenicity of Staphylococcus aureus. These factors include cell surface components that promote adherence to surfaces as well as exoproteins such as Panton-Valentine leukocidin (PVL), encoded by the luk-PV genes, that invade or bypass the immune system and are toxic to the host, thereby enhancing the severity of infections caused by methicillin-resistant Staphylococcus aureus (MRSA). The aim of this study was to determine the frequency of PVL-positive MRSA strains by real-time PCR and their antibiotic susceptibility patterns by phenotypic test. In total, 284 Staphylococcus isolates, identified by phenotypic methods from clinical samples of Shahrekord University Hospitals, Shahrekord, Iran, were tested for nuc, mecA, and PVL genes by TaqMan real-time PCR. The antibiotic susceptibility patterns of PVL-containing MRSA strains were determined via the disk diffusion method. In total, 196 isolates (69%) were nuc positive (i.e., S. aureus); of those isolates, 96 (49%) were mecA positive (MRSA). Eighteen (18.8%) of the 96 MRSA positive and 3 (3%) of the 100 methicillin-susceptible Staphylococcus aureus (MSSA) strains were PVL positive. PVL-positive MRSA strains were mostly recovered from tracheal specimens. Eight PVL-positive MRSA strains were resistant to all the tested antibiotics except vancomycin. A significant correlation (P = 0.001) was found between the mecA positivity and the presence of luk-PV genes. Community acquired (CA)-MRSA is becoming a public health concern in many parts of the world, including Asian countries. The variable prevalence of luk-PV-positive MRSA isolates in different regions and their rather high frequency in pneumonia necessitate the application of rapid diagnostic methods such as real-time PCR to improve treatment effectiveness.
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NASA Technical Reports Server (NTRS)
Fleming, M. L.
1980-01-01
Four possible arrangements of the materials experiment carrier (MEC) and power system (PS) thermal control loops were defined which would provide one kW of heat rejection for each kW of power to the MEC payload. These arrangements were compared to the baseline reference concept which provides only 16 kW heat rejection to show the cost of obtaining symmetry in terms of dollars, weight, complexity, growth potential, ease of integration, technology and total launch weight. The results of these comparisons was that the concept which splits the PS thermal control loop into two systems, one to reject PS waste heat and one payload waste heat, appeared favorable. The fluid selection study resulted in recommendation of FC72 as the MEC heat transport fluid based on the thermal and physical characteristics. The coatings reviewed indicated anodized and alodine treated aluminum surfaces or silver teflon are the best choices for the MEC vehicle where durability is an important factor. For high temperature radiators silver teflon or zinc orthotitanate are recommended choices.
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Mutational landscape of yeast mutator strains.
Serero, Alexandre; Jubin, Claire; Loeillet, Sophie; Legoix-Né, Patricia; Nicolas, Alain G
2014-02-04
The acquisition of mutations is relevant to every aspect of genetics, including cancer and evolution of species on Darwinian selection. Genome variations arise from rare stochastic imperfections of cellular metabolism and deficiencies in maintenance genes. Here, we established the genome-wide spectrum of mutations that accumulate in a WT and in nine Saccharomyces cerevisiae mutator strains deficient for distinct genome maintenance processes: pol32Δ and rad27Δ (replication), msh2Δ (mismatch repair), tsa1Δ (oxidative stress), mre11Δ (recombination), mec1Δ tel1Δ (DNA damage/S-phase checkpoints), pif1Δ (maintenance of mitochondrial genome and telomere length), cac1Δ cac3Δ (nucleosome deposition), and clb5Δ (cell cycle progression). This study reveals the diversity, complexity, and ultimate unique nature of each mutational spectrum, composed of punctual mutations, chromosomal structural variations, and/or aneuploidies. The mutations produced in clb5Δ/CCNB1, mec1Δ/ATR, tel1Δ/ATM, and rad27Δ/FEN1 strains extensively reshape the genome, following a trajectory dependent on previous events. It comprises the transmission of unstable genomes that lead to colony mosaicisms. This comprehensive analytical approach of mutator defects provides a model to understand how genome variations might accumulate during clonal evolution of somatic cell populations, including tumor cells.
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Zhang, Hongwei; Ma, Luyao; Ma, Lina; Hua, Marti Z; Wang, Shuo; Lu, Xiaonan
2017-02-21
Methicillin-resistant Staphylococcus aureus (MRSA) is considered as one of the leading causes of food poisonings worldwide. Due to the high prevalence and extensive challenges in clinical treatment, a rapid and accurate detection method is required to differentiate MRSA from other S. aureus isolated from foods. Since the methicillin resistance of S. aureus is due to the acquisition of the mecA gene from staphylococcal chromosome cassette, the presence of the mecA gene is interpreted as a marker for the identification of MRSA. In this study, a low-cost lateral flow immunoassay (LFI) strip was used to detect the mecA amplicons subsequent to polymerase chain reaction (PCR). The specificity of this PCR-LFI assay was tested between MRSA and methicillin-susceptive S. aureus. Both the test line and control line were shown up on the LFI strip for MRSA, whereas only the control line developed for methicillin-susceptive S. aureus. The detection limit of PCR-LFI assay was 20fg for genomic DNA (100 times more sensitive than gel electrophoresis) and 2×10 0 CFU per 100g of pork products after enrichment at 37°C for 48h. The total detection time of using LFI was 3min, which was faster than the conventional electrophoresis (~45min). With the performance of PCR-LFI, 7 out of 42 S. aureus isolates were identified to be MRSA from imported pork products, which was consistent to the standardized minimum inhibitory concentration assay. This mecA-based PCR-LFI strip can be used for rapid and accurate detection of MRSA isolated from commercial pork products. Copyright © 2016. Published by Elsevier B.V.
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[Molecular study of methicillin-resistant Staphylococcus haemolyticus in a Mexican hospital].
Castro, Natividad; Loaiza-Loeza, María Salomé; Calderón-Navarro, Amparo; Sánchez, Alejandro; Silva-Sánchez, Jesús
2006-01-01
To perform the molecular characterization of methicillin-resistant Staphylococcus haemolyticus (MRSH) clinical isolates from patients in a Mexican hospital. Sixty three Staphylococcus ssp. isolates collected from September 2000 to October 2002 were analyzed. Antimicrobial susceptibility was determined by disk diffusion method and the presence of the mecA gene was detected by PCR technique. Isolates characterization was carried out by pulsed field gel electrophoresis (PFGE). The frequency of S. haemolyticus was 25.5% (18 of 63 clinical isolates), all S. haemolyticus isolates were methicillin-resistant and they were positive for the mecA gene. A major pattern (A) with 8 subtypes was identified. This clone was distributed during the 20 months period. Most of them were isolated from the surgery (55%) and pediatric services (27.5%). The methicillin-resistant S. haemolyticus permanence as pathogen in this hospital, suggest the implementation of control programs in order to decrease the prevalence of this multiresistant pathogen.
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Osman, Kamelia; Badr, Jihan; Al-Maary, Khalid S.; Moussa, Ihab M. I.; Hessain, Ashgan M.; Girah, Zeinab M. S. Amin; Abo-shama, Usama H.; Orabi, Ahmed; Saad, Aalaa
2016-01-01
The use of antibiotics in farm management (growing crops and raising animals) has become a major area of concern. Its implications is the consequent emergence of antibiotic resistant bacteria (ARB) and accordingly their access into the human food chain with passage of antibiotic resistance genes (ARG) to the normal human intestinal microbiota and hence to other pathogenic bacteria causative human disease. Therefore, we pursued in this study to unravel the frequency and the quinolone resistance determining region, mecA and cfr genes of methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), methicillin-resistant coagulase-negative staphylococci (MRCNS) and methicillin-susceptible coagulase-negative staphylococci (MSCNS) isolated from the retail trade of ready-to-eat raw chicken meat samples collected during 1 year and sold across the Great Cairo area. The 50 Staphylococcus isolated from retail raw chicken meat were analyzed for their antibiotic resistance phenotypic profile on 12 antibiotics (penicillin, oxacillin, methicillin, ampicillin-sulbactam, erythromycin, tetracycline, clindamycin, gentamicin, ciprofloxacin, chloramphenicol, sulfamethoxazole-trimethoprim, and vancomycin) and their endorsement of the quinolone resistance determining region, mecA and cfr genes. The isolation results revealed 50 isolates, CPS (14) and CNS (36), representing ten species (S. aureus, S. hyicus, S. epidermedius, S. lugdunensis, S. haemolyticus, S. hominus, S. schleiferi, S. cohnii, S. intermedius, and S. lentus). Twenty seven isolates were methicillin-resistant. Out of the characterized 50 staphylococcal isolates, three were MRSA but only 2/3 carried the mecA gene. The ARG that bestows resistance to quinolones, β-lactams, macrolides, lincosamides, and streptogramin B [MLS(B)] in MRSA and MR-CNS were perceived. According to the available literature, the present investigation was a unique endeavor into the identification of the quinolone-resistance-determining-regions, the identification of MRSA and MR-CNS from retail chicken meat in Egypt. In addition, these isolates might indicate the promulgation of methicillin, oxacillin and vancomycin resistance in the community and imply food safety hazards. PMID:27920760
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Mkannez, Ghada; Gagné-Ouellet, Valérie; Nsaibia, Mohamed Jalloul; Boulanger, Marie-Chloé; Rosa, Mickael; Argaud, Deborah; Hadji, Fayez; Gaudreault, Nathalie; Rhéaume, Gabrielle; Bouchard, Luigi; Bossé, Yohan; Mathieu, Patrick
2018-05-02
Calcific aortic valve disease (CAVD) is characterized by the osteogenic transition of valve interstitial cells (VICs). In CAVD, lysophosphatidic acid (LysoPA), a lipid mediator with potent osteogenic activity, is produced in the aortic valve (AV) and is degraded by membrane-associated phospholipid phosphatases (PLPPs). We thus hypothesized that a dysregulation of PLPPs could participate to the osteogenic reprograming of VICs during CAVD. The expression of PLPPs was examined in human control and mineralized AVs and comprehensive analyses were performed to document the gene regulation and impact of PLPPs on the osteogenic transition of VICs. We found that PLPP3 gene and enzymatic activity were downregulated in mineralized AVs. Multidimensional gene profiling in 21 human AVs showed that expression of PLPP3 was inversely correlated with the level of 5-methylcytosine (5meC) located in an intronic mammalian interspersed repeat element (MIR). Bisulfite pyrosequencing in a larger series of 67 AVs confirmed that 5meC in intron 1 was increased by 2.2-fold in CAVD compared to control AVs. In isolated cells, epigenome editing with CRISPR-Cas9 system containing a deficient Cas9 fused with DNA methyltransferase (dCas9-DNMT) was used to increase 5meC in the intronic enhancer and showed that it reduced significantly the expression of PLPP3. Knockdown experiments showed that lower expression of PLPP3 in VICs promotes an osteogenic program. DNA methylation of a MIR-based enhancer downregulates the expression of PLPP3 and promotes the mineralization of the AV.
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Antimicrobial Resistance Genes in Pigeons from Public Parks in Costa Rica.
Blanco-Peña, K; Esperón, F; Torres-Mejía, A M; de la Torre, A; de la Cruz, E; Jiménez-Soto, M
2017-11-01
Antimicrobial resistance is known to be an emerging problem, but the extent of the issue remains incomplete. The aim of this study was to determine the presence or absence of nine resistance genes (bla TEM , catI, mecA, qnrS, sulI, sulII, tet(A), tet(Q), vanA) in the faeces of 141 pigeons from four urban parks in Alajuela, Guadalupe, Tres Ríos and San José in Costa Rica. The genes were identified by real-time PCR directly from enema samples. About 30% of the samples were positive for genes catI and sulI; between 13% and 17% were positive for qnrS, sulII, tet(A) and tet(Q); and 4% were positive for bla TEM . The mecA and vanA genes were not detected. The average of antimicrobial resistance genes detected per pigeon was 2. Eight different patterns of resistance were identified, without differences in the sampling areas, being the most common pattern 2 (sulII positive samples). During rainy season, the genes more frequently found were sulI and tet(A). In conclusion, the urban inhabiting pigeons tested are currently carrying antimicrobial resistance genes, potentially acting as reservoirs of resistant bacteria and vectors to humans. To the authors' knowledge, this is the first study carried out on direct detection of resistance genes in the digestive metagenomes of pigeons. © 2017 The Authors. Zoonoses and Public Health Published by Blackwell Verlag GmbH.
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Community-associated methicillin-resistant Staphylococcus aureus causing chronic pneumonia.
Enayet, Iram; Nazeri, Ali; Johnson, Leonard B; Riederer, Kathleen; Pawlak, Joan; Saravolatz, Louis D
2006-04-01
A young woman presented with pneumonia of a 3-month duration with predominantly nodular pulmonary infiltrates. Methicillin-resistant Staphylococcus aureus was identified in multiple cultures of sputum specimens. According to findings of pulsed-field gel electrophoresis, the isolate was identical to USA 300 and carried a type IV Staphylococcus cassette chromosome mec type IV gene and the genes for Panton-Valentine leukocidin.
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Gajewska, Małgorzata; Motyl, Tomasz
2004-10-01
TGF-beta 1 is an antiproliferative and apoptogenic factor for mammary epithelial cells (MEC) acting in an auto/paracrine manner and thus considered an important local regulator of mammary tissue involution. However, the apoptogenic signaling pathway induced by this cytokine in bovine MEC remains obscure. The present study was focused on identification of molecules involved in apoptogenic signaling of transforming growth factor-beta 1 (TGF-beta 1) in the model of bovine mammary epithelial cell line (BME-UV1). Laser scanning cytometry (LSC), Western blot and electrophoretic mobility shift assay (EMSA) were used for analysis of expression and activity of TGF-beta 1-related signaling molecules. The earliest response occurring within 1-2 h after TGF-beta 1 administration was an induction and activation of R-Smads (Smad2 and Smad3) and Co-Smad (Smad4). An evident formation of Smad-DNA complexes began from 2nd hour after MEC exposure to TGF-beta 1. Similarly to Smads, proteins of AP1 complex: phosphorylated c-Jun and JunD appeared to be early reactive molecules; however, an increase in their expression was detected only in cytosolic fraction. In the next step, an increase of IGF binding protein-3 (IGFBP-3) and IGFBP-4 expression was observed from 6th hour followed by a decrease in the activity of protein kinase B (PKB/Akt), which occurred after 24 h of MEC exposure to TGF-beta 1. The decrease in PKB/Akt activity coincided in time with the decline of phosphorylated Bad expression (inactive form). Present study supported additional evidence that stimulation of insulin-like growth factor I (IGF-I) was associated with complete abrogation of TGF-beta 1-induced activation of Bad and Bax and in the consequence protection against apoptosis. In conclusion, apoptotic effect of TGF-beta 1 in bovine MEC is mediated by IGFBPs and occurs through IGF-I sequestration, resulting in inhibition of PKB/Akt-dependent survival pathway.
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Ziegel, Rebecca; Shallop, Anthony; Upadhyaya, Pramod; Jones, Roger; Tretyakova, Natalia
2004-01-20
All CG dinucleotides along exons 5-8 of the p53 tumor suppressor gene contain endogenous 5-methylcytosine (MeC). These same sites (e.g., codons 157, 158, 245, 248, and 273) are mutational hot spots in smoking-induced lung cancer. Several groups used the UvrABC endonuclease incision assay to demonstrate that methylated CG dinucleotides of the p53 gene are the preferred binding sites for the diol epoxides of bay region polycyclic aromatic hydrocarbons (PAH). In contrast, effects of endogenous cytosine methylation on the distribution of DNA lesions induced by tobacco-specific nitrosamines, e.g., 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), have not been elucidated. In the work presented here, a stable isotope labeling HPLC-ESI-MS/MS approach was employed to analyze the reactivity of the N7 and O6 positions of guanines within hemimethylated and fully methylated CG dinucleotides toward NNK-derived methylating and pyridyloxobutylating species. 15N3-labeled guanine bases were placed within synthetic DNA sequences representing endogenously methylated p53 codons 154, 157, and 248, followed by treatment with acetylated precursors to NNK diazohydroxides. HPLC-ESI-MS/MS analysis was used to determine the relative yields of N7- and O6-guanine adducts at the 15N3-labeled position. In all cases, the presence of MeC inhibited the formation of N7-methylguanine, O6-methylguanine, and O6-pyridyloxobutylguanine at a neighboring G, with the greatest decrease observed in fully methylated dinucleotides and at guanines preceded by MeC. Furthermore, the O6-Me-dG/N7-Me-G molar ratios were decreased in the presence of the 5'-neighboring MeC, suggesting that the observed decline in O6-alkylguanine adduct yields is, at least partially, a result of an altered reactivity pattern in methylated CG dinucleotides. These results indicate that, unlike N2-guanine adducts of PAH diol epoxides, NNK-induced N7- and O6-alkylguanine adducts are not preferentially formed at the endogenously methylated CG sites within the p53 tumor suppressor gene.
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Brennan, Orla M.; Deasy, Emily C.; Rossney, Angela S.; Kinnevey, Peter M.; Ehricht, Ralf; Monecke, Stefan; Coleman, David C.
2012-01-01
One hundred seventy-five isolates representative of methicillin-resistant Staphylococcus aureus (MRSA) clones that predominated in Irish hospitals between 1971 and 2004 and that previously underwent multilocus sequence typing (MLST) and staphylococcal cassette chromosome mec (SCCmec) typing were characterized by spa typing (175 isolates) and DNA microarray profiling (107 isolates). The isolates belonged to 26 sequence type (ST)-SCCmec types and subtypes and 35 spa types. The array assigned all isolates to the correct MLST clonal complex (CC), and 94% (100/107) were assigned an ST, with 98% (98/100) correlating with MLST. The array assigned all isolates to the correct SCCmec type, but subtyping of only some SCCmec elements was possible. Additional SCCmec/SCC genes or DNA sequence variation not detected by SCCmec typing was detected by array profiling, including the SCC-fusidic acid resistance determinant Q6GD50/fusC. Novel SCCmec/SCC composite islands (CIs) were detected among CC8 isolates and comprised SCCmec IIA-IIE, IVE, IVF, or IVg and a ccrAB4-SCC element with 99% DNA sequence identity to SCCM1 from ST8/t024-MRSA, SCCmec VIII, and SCC-CI in Staphylococcus epidermidis. The array showed that the majority of isolates harbored one or more superantigen (94%; 100/107) and immune evasion cluster (91%; 97/107) genes. Apart from fusidic acid and trimethoprim resistance, the correlation between isolate antimicrobial resistance phenotype and the presence of specific resistance genes was ≥97%. Array profiling allowed high-throughput, accurate assignment of MRSA to CCs/STs and SCCmec types and provided further evidence of the diversity of SCCmec/SCC. In most cases, array profiling can accurately predict the resistance phenotype of an isolate. PMID:22869569
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Alva-Murillo, Nayeli; Ochoa-Zarzosa, Alejandra; López-Meza, Joel E
2017-01-01
Bovine mammary epithelial cells (bMECs) contribute to mammary gland defense against invading pathogens, such as Staphylococcus aureus (intracellular facultative), which is recognized by TLR2. In a previous report, we showed that sodium octanoate [NaO, a medium chain fatty acid (C8)] induces (0.25 mM) or inhibits (1 mM) S. aureus internalization into bMECs and differentially regulates the innate immune response (IIR). However, the molecular mechanisms have not been described, which was the aim of this study. The results showed that α5β1 integrin membrane abundance (MA) was increased in 0.25 mM NaO-treated cells, but TLR2 or CD36 MA was not modified. When these receptors were blocked individually, 0.25 mM NaO-increased S. aureus internalization was notably reduced. Interestingly, in this condition, the IIR of the bMECs was impaired because MAPK (p38, JNK, and ERK1/2) phosphorylation and the activation of transcription factors related to these pathways were decreased. In addition, the 1 mM NaO treatment induced TLR2 MA, but neither the integrin nor CD36 MA was modified. The reduction in S. aureus internalization induced by 1 mM NaO was increased further when TLR2 was blocked. In addition, the phosphorylation levels of the MAPKs increased, and 13 transcriptional factors related to the IIR were slightly activated (CBF, CDP, c-Myb, AP-1, Ets-1/Pea-3, FAST-1, GAS/ISRE, AP-2, NFAT-1, OCT-1, RAR/DR-5, RXR/DR-1, and Stat-3). Moreover, the 1 mM NaO treatment up-regulated gene expression of IL-8 and RANTES and secretion of IL-1β. Notably, when 1 mM NaO-treated bMECs were challenged with S. aureus , the gene expression of IL-8 and IL-10 increased, while IL-1β secretion was reduced. In conclusion, our results showed that α5β1 integrin, TLR2 and CD36 are involved in 0.25 mM NaO-increased S. aureus internalization in bMECs. In addition, 1 mM NaO activates bMECs via the TLR2 signaling pathways (p38, JNK, and ERK1/2), which improves IIR before S. aureus invasion. Additionally, NaO (1 mM) might exert anti-inflammatory effects after bacterial internalization.
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Matrix mechanics controls FHL2 movement to the nucleus to activate p21 expression
Nakazawa, Naotaka; Sathe, Aneesh R.; Shivashankar, G. V.; Sheetz, Michael P.
2016-01-01
Substrate rigidity affects many physiological processes through mechanochemical signals from focal adhesion (FA) complexes that subsequently modulate gene expression. We find that shuttling of the LIM domain (domain discovered in the proteins, Lin11, Isl-1, and Mec-3) protein four-and-a-half LIM domains 2 (FHL2) between FAs and the nucleus depends on matrix mechanics. In particular, on soft surfaces or after the loss of force, FHL2 moves from FAs into the nucleus and concentrates at RNA polymerase (Pol) II sites, where it acts as a transcriptional cofactor, causing an increase in p21 gene expression that will inhibit growth on soft surfaces. At the molecular level, shuttling requires a specific tyrosine in FHL2, as well as phosphorylation by active FA kinase (FAK). Thus, we suggest that FHL2 phosphorylation by FAK is a critical, mechanically dependent step in signaling from soft matrices to the nucleus to inhibit cell proliferation by increasing p21 expression. PMID:27742790
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Soeorg, Hiie; Metsvaht, Hanna Kadri; Keränen, Evamaria Elisabet; Eelmäe, Imbi; Merila, Mirjam; Ilmoja, Mari-Liis; Metsvaht, Tuuli; Lutsar, Irja
2018-04-02
Staphylococcus haemolyticus is a common colonizer and cause of late-onset sepsis (LOS) in preterm neonates. By describing genetic relatedness, we aimed to determine whether mother's breast milk (BM) is a source of S. haemolyticus colonizing neonatal gut and skin and/or causing LOS. S. haemolyticus was isolated from stool and skin swabs of 49 BM-fed preterm neonates admitted to neonatal intensive care unit, 20 healthy BM-fed term neonates and BM of mothers once a week and typed by multilocus variable-number tandem-repeat analysis (MLVA) and multilocus sequence typing (MLST). Virulence-related genes were determined by PCR. Compared with term neonates S. haemolyticus colonized more commonly gut (35% vs 89.9%; p<0.001) and skin (50% vs 91.8%; p<0.001) of preterm neonates and mothers' BM (15% vs 38.8%). Isolates from preterm compared with term neonates and their mothers carried more commonly the mecA gene (83.5% vs 5.4%; p<0.001) and IS256 (52.4% vs 2.7%; p<0.001) and belonged to clonal complex 29 (89.1% vs 63%; p=0.014). Only 7 (14.3%) preterm and 3 (15%) term neonates were colonized in gut or on skin with MLVA-types indistinguishable from those in BM. Most frequent MLVA-types belonged to sequence type 3 or 42, comprised 71.1-78.4% of isolates from preterm neonates/mothers and caused all seven LOS episodes. LOS-causing strain colonized the gut of 4/7 and the skin of 5/7 neonates, but not BM, prior to onset of LOS. S. haemolyticus colonizing gut and skin or causing LOS in preterm neonates rarely originate from BM, but are mecA-positive strains adapted to hospital environment.
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Salimnia, Hossein; Lephart, Paul R.; Schreckenberger, Paul; DesJarlais, Sharon M.; Johnson, J. Kristie; Robinson, Gwen; Carroll, Karen C.; Greer, Amy; Morgan, Margie; Chan, Raymond; Loeffelholz, Michael; Valencia-Shelton, Frances; Jenkins, Stephen; Schuetz, Audrey N.; Daly, Judy A.; Barney, Trenda; Hemmert, Andrew; Kanack, Kristen J.
2016-01-01
Sepsis is a major cause of morbidity, mortality, and increased medical expense. Rapid diagnosis improves outcomes and reduces costs. The FilmArray blood culture identification panel (BioFire Diagnostics LLC, Salt Lake City, UT), a highly multiplexed PCR assay, can identify 24 etiologic agents of sepsis (8 Gram-positive, 11 Gram-negative, and 5 yeast species) and three antimicrobial resistance genes (mecA, vanA/B, and blaKPC) from positive blood culture bottles. It provides results in about 1 h with 2 min for assay setup. We present the results of an eight-center trial comparing the sensitivity and specificity of the panel with those of the laboratories' standard phenotypic identification techniques, as well as with molecular methods used to distinguish Acinetobacter baumannii from other members of the A. calcoaceticus-A. baumannii complex and to detect antimicrobial resistance genes. Testing included 2,207 positive aerobic blood culture samples, 1,568 clinical and 639 seeded. Samples were tested fresh or were frozen for later testing within 8 h after the bottles were flagged as positive by an automated blood culture system. At least one organism was detected by the panel in 1,382 (88.1%) of the positive clinical specimens. The others contained primarily off-panel organisms. The panel reported multiple organisms in 81 (5.86%) positive clinical specimens. The unresolved blood culture identification sensitivity for all target detections exceeded 96%, except for Klebsiella oxytoca (92.2%), which achieved 98.3% sensitivity after resolution of an unavoidable phenotypic error. The sensitivity and specificity for vanA/B and blaKPC were 100%; those for mecA were 98.4 and 98.3%, respectively. PMID:26739158
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Roza, Carolina; Puel, Jean-Luc; Kress, Michaela; Baron, Anne; Diochot, Sylvie; Lazdunski, Michel; Waldmann, Rainer
2004-01-01
Mechanosensitive cation channels are thought to be crucial for different aspects of mechanoperception, such as hearing and touch sensation. In the nematode C. elegans, the degenerins MEC-4 and MEC-10 are involved in mechanosensation and were proposed to form mechanosensitive cation channels. Mammalian degenerin homologues, the H+-gated ASIC channels, are expressed in sensory neurones and are therefore interesting candidates for mammalian mechanosensors. We investigated the effect of an ASIC2 gene knockout in mice on hearing and on cutaneous mechanosensation and visceral mechanonociception. However, our data do not support a role of ASIC2 in those facets of mechanoperception. PMID:15169849
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Hofling-Lima, Ana Luisa; Pignatari, Antonio C. C.
2014-01-01
Staphylococcus epidermidis is an abundant member of the microbiota of the human skin and wet mucosa, which is commonly associated with sight-threatening infections in eyes with predisposing factors. Ocular S. epidermidis has become notorious because of its capability to form biofilms on different ocular devices and due to the evolving rates of antimicrobial resistance. In this study, the molecular epidemiology of 30 ocular methicillin-resistant S. epidermidis (MRSE) isolates was assessed using multilocus sequence typing (MLST). Antimicrobial resistance, accessory gene-regulator and staphylococcal cassette chromosome mec (SCCmec) types, biofilm formation, and the occurrence of biofilm-associated genes were correlated with MLST clonal complexes. Sequence types (STs) frequently found in the hospital setting were rarely found in our collection. Overall, 12 different STs were detected with a predominance of ST59 (30%), ST5 and ST6 (13.3% each). Most of the isolates (93.3%) belonged to the clonal complex 2 (CC2) and grouped mainly within subcluster CC2-II (92.9%). Isolates grouped within this subcluster were frequently biofilm producers (92.3%) with a higher occurrence of the aap (84.5%) and bhp (46.1%) genes compared to icaA (19.2%). SCCmec type IV (53.8%) was predominant within CC2-II strains, while 38.4% were nontypeable. In addition, CC2-II strains were frequently multidrug resistant (80.7%) and demonstrated to be particularly resistant to ciprofloxacin (80.8%), ofloxacin (77%), azithromycin (61.5%), and gentamicin (57.7%). Our findings demonstrate the predominance of a particular MRSE cluster causing ocular infections, which was associated with high rates of antimicrobial resistance and particularly the carriage of biofilm-related genes coding for proteinaceous factors implicated in biofilm accumulation. PMID:24523473
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Shariati, Laleh; Validi, Majid; Hasheminia, Ali Mohammad; Ghasemikhah, Reza; Kianpour, Fariborz; Karimi, Ali; Nafisi, Mohammad Reza; Tabatabaiefar, Mohammad Amin
2016-01-01
Background: A diversity of virulence factors work together to create the pathogenicity of Staphylococcus aureus. These factors include cell surface components that promote adherence to surfaces as well as exoproteins such as Panton-Valentine leukocidin (PVL), encoded by the luk-PV genes, that invade or bypass the immune system and are toxic to the host, thereby enhancing the severity of infections caused by methicillin-resistant Staphylococcus aureus (MRSA). Objectives: The aim of this study was to determine the frequency of PVL-positive MRSA strains by real-time PCR and their antibiotic susceptibility patterns by phenotypic test. Materials and Methods: In total, 284 Staphylococcus isolates, identified by phenotypic methods from clinical samples of Shahrekord University Hospitals, Shahrekord, Iran, were tested for nuc, mecA, and PVL genes by TaqMan real-time PCR. The antibiotic susceptibility patterns of PVL-containing MRSA strains were determined via the disk diffusion method. Results: In total, 196 isolates (69%) were nuc positive (i.e., S. aureus); of those isolates, 96 (49%) were mecA positive (MRSA). Eighteen (18.8%) of the 96 MRSA positive and 3 (3%) of the 100 methicillin-susceptible Staphylococcus aureus (MSSA) strains were PVL positive. PVL-positive MRSA strains were mostly recovered from tracheal specimens. Eight PVL-positive MRSA strains were resistant to all the tested antibiotics except vancomycin. A significant correlation (P = 0.001) was found between the mecA positivity and the presence of luk-PV genes. Conclusions: Community acquired (CA)-MRSA is becoming a public health concern in many parts of the world, including Asian countries. The variable prevalence of luk-PV-positive MRSA isolates in different regions and their rather high frequency in pneumonia necessitate the application of rapid diagnostic methods such as real-time PCR to improve treatment effectiveness. PMID:27099685
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Mammalian Homologs of Yeast Checkpoint Genes
2002-07-01
pathway is sensitive to various forms of DNA damage Developmental Biology throughout the cell cycle . The DNA replication check- Yale University point...components would be ordered into pathways for mammalian checkpoint function, with emphasis on p53 regulation, cell cycle regulation, and complementation...structurally related to the human tumor suppressor ATM. MEC1 and RAD53, two essential genes, play a central role in DNA damage checkpoints at all cell cycle
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Kim, Su Young; Kim, Ju; Jeong, Seung-Il; Jahng, Kwang Yeop; Yu, Kang-Yeol
2015-01-01
The antimicrobial killing activity toward methicillin-resistant Staphylococcus aureus (MRSA) has been a serious emerging global issue. In a continuing search for compounds with antibacterial activity against several microorganisms including S. aureus and MRSA, an n-hexane extract of Magnolia officinalis was found to contain magnolol. This compound exhibited potent activity against S. aureus, standard methicillin-susceptible S. aureus (MSSA), and MRSA as well as clinical MRSA isolates. When combined with oxacillin, the antibacterial activities of magnolol and honokiol against the MRSA strain were increased compared to single treatment without antibiotics at 10 µg/mL and 25 µg/mL, respectively. These activities of magnolol and honokiol were dose dependent. Also, magnolol showed synergistic effects with oxacillin against 13 clinical isolates of MRSA. It was determined that magnolol and honokiol had a synergistic effect with oxacillin against MRSA strain. Furthermore, the magnolol inhibited the expression of the resistant genes, mecA, mecI, femA, and femB, in mRNA. We concluded that the antibacterial activity of magnolol against MRSA strain is more related to the mecI's pathway and components of the cell wall than mecR1. Therefore, the results obtained in this study suggest that the combination of magnolol and antibiotics could lead to the development of new combination antibiotics against MRSA infection. PMID:26357651
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Kim, Su Young; Kim, Ju; Jeong, Seung-Il; Jahng, Kwang Yeop; Yu, Kang-Yeol
2015-01-01
The antimicrobial killing activity toward methicillin-resistant Staphylococcus aureus (MRSA) has been a serious emerging global issue. In a continuing search for compounds with antibacterial activity against several microorganisms including S. aureus and MRSA, an n-hexane extract of Magnolia officinalis was found to contain magnolol. This compound exhibited potent activity against S. aureus, standard methicillin-susceptible S. aureus (MSSA), and MRSA as well as clinical MRSA isolates. When combined with oxacillin, the antibacterial activities of magnolol and honokiol against the MRSA strain were increased compared to single treatment without antibiotics at 10 µg/mL and 25 µg/mL, respectively. These activities of magnolol and honokiol were dose dependent. Also, magnolol showed synergistic effects with oxacillin against 13 clinical isolates of MRSA. It was determined that magnolol and honokiol had a synergistic effect with oxacillin against MRSA strain. Furthermore, the magnolol inhibited the expression of the resistant genes, mecA, mecI, femA, and femB, in mRNA. We concluded that the antibacterial activity of magnolol against MRSA strain is more related to the mecI's pathway and components of the cell wall than mecR1. Therefore, the results obtained in this study suggest that the combination of magnolol and antibiotics could lead to the development of new combination antibiotics against MRSA infection.
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Kool, Dorien M; Zhu, Baoli; Rijpstra, W Irene C; Jetten, Mike S M; Ettwig, Katharina F; Sinninghe Damsté, Jaap S
2012-12-01
The recently described bacterium "Candidatus Methylomirabilis oxyfera" couples the oxidation of the important greenhouse gas methane to the reduction of nitrite. The ecological significance of "Ca. Methylomirabilis oxyfera" is still underexplored, as our ability to identify the presence of this bacterium is thus far limited to DNA-based techniques. Here, we investigated the lipid composition of "Ca. Methylomirabilis oxyfera" to identify new, gene-independent biomarkers for the environmental detection of this bacterium. Multiple "Ca. Methylomirabilis oxyfera" enrichment cultures were investigated. In all cultures, the lipid profile was dominated up to 46% by the fatty acid (FA) 10-methylhexadecanoic acid (10MeC(16:0)). Furthermore, a unique FA was identified that has not been reported elsewhere: the monounsaturated 10-methylhexadecenoic acid with a double bond at the Δ7 position (10MeC(16:1Δ7)), which comprised up to 10% of the total FA profile. We propose that the typical branched fatty acids 10MeC(16:0) and 10MeC(16:1Δ7) are key and characteristic components of the lipid profile of "Ca. Methylomirabilis oxyfera." The successful detection of these fatty acids in a peatland from which one of the enrichment cultures originated supports the potential of these unique lipids as biomarkers for the process of nitrite-dependent methane oxidation in the environment.
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Clonal profile, virulence and resistance of Staphylococcus aureus isolated from sheep milk.
Martins, Katheryne Benini; Faccioli-Martins, Patricia Yoshida; Riboli, Danilo Flávio Moraes; Pereira, Valéria Cataneli; Fernandes, Simone; Oliveira, Aline A; Dantas, Ariane; Zafalon, Luiz Francisco; da Cunha, Maria de Lourdes Ribeiro de Souza
2015-06-01
The objective of this study was to characterize the clonal profile, virulence factors and antimicrobial resistance, particularly oxacillin resistance, of Staphylococcus aureus isolated from sheep milk. Milk samples were collected from all teats for the California Mastitis Test (CMT), somatic cell count, identification of S. aureus, investigation in these strains of genes encoding toxins (sea, seb, sec, sed, tst), biofilm (icaA, icaC, icaD, bap), leukocidin (luk-PV) oxacillin resistance by mecA gene detection and susceptibility testing (12 antibiotics). Messenger RNA expression was evaluated by RT-PCR in isolates carrying toxin and biofilm genes. Biofilm formation was also evaluated phenotypically by adherence to polystyrene plates. The clonal profile of S. aureus was investigated by pulsed-field gel electrophoresis. A total of 473 milk samples were collected from 242 animals on three farms and 20 S. aureus strains were isolated and none carried the mecA gene. The two sec gene-positive isolates and the isolates carrying the tst and luk-PV genes were positive by RT-PCR. Staphylococcus aureus isolated from the three flocks studied showed high susceptibility to the drugs tested and none was biofilm producer, indicating that biofilm formation was not a virulence factor causing infection by these strains. The typing of 17 S. aureus isolates revealed the presence of a common clone on the three farms studied, and the presence and expression of the sec and tst genes in one strain of this clone suggest the possible acquisition of virulence genes by this clone, a fact that is important for animal health and food hygiene.
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Presence of the optrA Gene in Methicillin-Resistant Staphylococcus sciuri of Porcine Origin.
Fan, Run; Li, Dexi; Wang, Yang; He, Tao; Feßler, Andrea T; Schwarz, Stefan; Wu, Congming
2016-12-01
A total of 57 methicillin-resistant Staphylococcus aureus (MRSA) isolates and 475 methicillin-resistant coagulase-negative staphylococci (MRCoNS) collected from pigs in the Guangdong province of China in 2014 were investigated for the presence of the novel oxazolidinone-phenicol resistance gene optrA The optrA gene was detected in 6.9% (n = 33) of the MRCoNS, all of which were Staphylococcus sciuri isolates, but in none of the MRSA isolates. Five optrA-carrying methicillin-resistant (MR) S. sciuri isolates also harbored the multiresistance gene cfr Pulsed-field gel electrophoresis (PFGE) and dru typing of the 33 optrA-carrying MR S. sciuri isolates revealed 25 patterns and 5 sequence types, respectively. S1 nuclease PFGE and Southern blotting confirmed that optrA was located in the chromosomal DNAs of 29 isolates, including 1 cfr-positive isolate. The remaining four isolates harbored a ∼35-kb pWo28-3-like plasmid on which optrA and cfr were located together with other resistance genes, as confirmed by sequence analysis. Six different types of genetic environments (types I to VI) of the chromosome-borne optrA genes were identified; these types had the optrA gene and its transcriptional regulator araC in common. Tn558 was found to be associated with araC-optrA in types II to VI. The optrA gene in types II and III was found in close proximity to the ccr gene complex of the respective staphylococcal cassette chromosome mec element (SCCmec). Since oxazolidinones are last-resort antimicrobial agents for the control of serious infections caused by methicillin-resistant staphylococci in humans, the location of the optrA gene close to the ccr complex is an alarming observation. Copyright © 2016, American Society for Microbiology. All Rights Reserved.
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Systematic identification of fragile sites via genome-wide location analysis of γ-H2AX
Szilard, Rachel K.; Jacques, Pierre-Étienne; Laramée, Louise; Cheng, Benjamin; Galicia, Sarah; Bataille, Alain R.; Yeung, ManTek; Mendez, Megan; Bergeron, Maxime; Robert, François; Durocher, Daniel
2011-01-01
Phosphorylation of histone H2AX is an early response to DNA damage in eukaryotes. In Saccharomyces cerevisiae, DNA damage or replication fork stalling results in histone H2A phosphorylation to yield γ-H2A (yeast γ-H2AX) in a Mec1 (ATR)- and Tel1 (ATM)- dependent manner. Here, we describe the genome-wide location analysis of γ-H2A as a strategy to identify loci prone to engage the Mec1 and Tel1 pathways. Remarkably, γ-H2A enrichment overlaps with loci prone to replication fork stalling and is caused by the action of Mec1 and Tel1, indicating that these loci are prone to breakage. Moreover, about half the sites enriched for γ-H2A map to repressed protein-coding genes, and histone deacetylases are necessary for formation of γ-H2A at these loci. Finally, our work indicates that high resolution mapping of γ-H2AX is a fruitful route to map fragile sites in eukaryotic genomes. PMID:20139982
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Feßler, Andrea T; Thomas, Patricia; Mühldorfer, Kristin; Grobbel, Mirjam; Brombach, Julian; Eichhorn, Inga; Monecke, Stefan; Ehricht, Ralf; Schwarz, Stefan
2018-05-01
Antimicrobial resistance of Staphylococcus aureus is a major problem in human and veterinary medicine. The aim of this study was to characterise S. aureus isolates from wild and zoo animals mainly associated with bacterial infections. In total, 23 S. aureus isolates, including nine from wild animals and 14 from zoo animals, were obtained during routine diagnostics. All isolates were subjected to multilocus sequence typing (MLST), spa typing, macrorestriction analysis with subsequent SmaI pulsed-field gelelectrophoresis (PFGE), antimicrobial susceptibility testing and S. aureus-specific DNA-microarray analysis. Resistant isolates were also tested for their respective resistance genes by PCR. Isolates from zoo animals and wildlife showed a high diversity of MLST types, spa types and PFGE patterns. Nineteen different spa types were identified, including three novel types and 16 main macrorestriction patterns. Only few isolates were resistant to members of four classes of antimicrobial agents and harboured the respective resistance genes (β-lactams [blaZ, mecA, mecC], tetracyclines [tet(K), tet(L)] and chloramphenicol [cat pC221 ]) or mutations (fluoroquinolones). The DNA microarray analysis identified one isolate from a zoo animal harbouring the toxic shock syndrome toxin gene tst1. Moreover, several enterotoxin genes were detected in five S. aureus isolates. All isolates were negative for Panton-Valentine leukocidin (PVL) genes, but the animal-associated leukocidin genes lukM/lukF-P83 were found in three isolates from two animals. Copyright © 2018 Elsevier B.V. All rights reserved.
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Ciftci, Alper; Findik, Arzu; Onuk, Ertan Emek; Savasan, Serap
2009-01-01
This study aimed to detect methicillin resistant and slime producing Staphylococcus aureus in cases of bovine mastitis. A triplex PCR was optimized targetting 16S rRNA, nuc and mecA genes for detection of Staphylococcus species, S. aureus and methicillin resistance, respectively. Furthermore, for detection of slime producing strains, a PCR assay targetting icaA and icaD genes was performed. In this study, 59 strains were detected as S. aureus by both conventional tests and PCR, and 13 of them were found to be methicillin resistant and 4 (30.7%) were positive for mecA gene. Although 22 of 59 (37.2%) S. aureus isolates were slime-producing in Congo Red Agar, in PCR analysis only 15 were positive for both icaA and icaD genes. Sixteen and 38 out of 59 strains were positive for icaA and icaD gene, respectively. Only 2 of 59 strains were positive for both methicillin resistance and slime producing, phenotypically, suggesting lack of correlation between methicillin resistance and slime production in these isolates. In conclusion, the optimized triplex PCR in this study was useful for rapid and reliable detection of methicillin resistant S. aureus. Furthermore, only PCR targetting icaA and icaD may not sufficient to detect slime production and further studies targetting other ica genes should be conducted for accurate evaluation of slime production characters of S. aureus strains. PMID:24031354
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Janmeda, Mamta; Kharadi, Vishnu; Pandya, Gaurav; Brahmkshtri, Balkrishna; Ramani, Umed; Tyagi, Kuldeep
2017-01-01
Aim: Aim of the study was to study the relative gene expression of genes associated with fatty acid synthesis at 60 days postpartum (pp) in bovine mammary epithelial cells (MECs) of Surti and Jafarabadi buffaloes. Materials and Methods: A total of 10 healthy Surti and Jafarabadi buffaloes of each breed were selected at random from Livestock Research Station, Navsari and Cattle Breeding Farm, Junagadh, Gujarat, respectively, for this study. Milk sample was collected from each selected buffalo at day 60 pp from these two breeds to study relative gene expression of major milk fat genes using non-invasive approach of obtaining primary bovine MECs (pBMEC) from milk samples. Results: In this study overall, the relative expression of the six major milk lipogenic genes butyrophilin subfamily 1 member A1 (BTN1A1), stearoyl-CoA desaturase (SCD), lipoprotein lipase (LPL), glycerol-3-phosphate acyltransferase mitochondrial (GPAM), acetyl-coenzyme A carboxylase alpha (ACACA), and lipin (LPIN) did not show changes in expression patterns at 60th day of lactation in both Surti and Jafarabadi buffaloes. Conclusion: The pBMEC can be successfully recovered from 1500 ml of milk of Surti and Jafarabadi buffaloes using antibody-mediated magnetic bead separation and can be further used for recovering RNA for down step quantification of major milk lipogenic gene expression. The relative expression of the six major milk lipogenic genes BTN1A1, SCD, LPL, GPAM, ACACA, and LPIN did not show changes in expression patterns in both Surti and Jafarabadi buffaloes, suggesting expression levels of lipogenic genes are maintained almost uniform till peak lactation without any significant difference. PMID:28620248
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Conserved binding of GCAC motifs by MEC-8, couch potato, and the RBPMS protein family
Soufari, Heddy
2017-01-01
Precise regulation of mRNA processing, translation, localization, and stability relies on specific interactions with RNA-binding proteins whose biological function and target preference are dictated by their preferred RNA motifs. The RBPMS family of RNA-binding proteins is defined by a conserved RNA recognition motif (RRM) domain found in metazoan RBPMS/Hermes and RBPMS2, Drosophila couch potato, and MEC-8 from Caenorhabditis elegans. In order to determine the parameters of RNA sequence recognition by the RBPMS family, we have first used the N-terminal domain from MEC-8 in binding assays and have demonstrated a preference for two GCAC motifs optimally separated by >6 nucleotides (nt). We have also determined the crystal structure of the dimeric N-terminal RRM domain from MEC-8 in the unbound form, and in complex with an oligonucleotide harboring two copies of the optimal GCAC motif. The atomic details reveal the molecular network that provides specificity to all four bases in the motif, including multiple hydrogen bonds to the initial guanine. Further studies with human RBPMS, as well as Drosophila couch potato, confirm a general preference for this double GCAC motif by other members of the protein family and the presence of this motif in known targets. PMID:28003515
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Knierim, James J; Neunuebel, Joshua P; Deshmukh, Sachin S
2014-02-05
The hippocampus receives its major cortical input from the medial entorhinal cortex (MEC) and the lateral entorhinal cortex (LEC). It is commonly believed that the MEC provides spatial input to the hippocampus, whereas the LEC provides non-spatial input. We review new data which suggest that this simple dichotomy between 'where' versus 'what' needs revision. We propose a refinement of this model, which is more complex than the simple spatial-non-spatial dichotomy. MEC is proposed to be involved in path integration computations based on a global frame of reference, primarily using internally generated, self-motion cues and external input about environmental boundaries and scenes; it provides the hippocampus with a coordinate system that underlies the spatial context of an experience. LEC is proposed to process information about individual items and locations based on a local frame of reference, primarily using external sensory input; it provides the hippocampus with information about the content of an experience.
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Knierim, James J.; Neunuebel, Joshua P.; Deshmukh, Sachin S.
2014-01-01
The hippocampus receives its major cortical input from the medial entorhinal cortex (MEC) and the lateral entorhinal cortex (LEC). It is commonly believed that the MEC provides spatial input to the hippocampus, whereas the LEC provides non-spatial input. We review new data which suggest that this simple dichotomy between ‘where’ versus ‘what’ needs revision. We propose a refinement of this model, which is more complex than the simple spatial–non-spatial dichotomy. MEC is proposed to be involved in path integration computations based on a global frame of reference, primarily using internally generated, self-motion cues and external input about environmental boundaries and scenes; it provides the hippocampus with a coordinate system that underlies the spatial context of an experience. LEC is proposed to process information about individual items and locations based on a local frame of reference, primarily using external sensory input; it provides the hippocampus with information about the content of an experience. PMID:24366146
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Pisanu, Salvatore; Marogna, Gavino; Cubeddu, Tiziana; Pagnozzi, Daniela; Cacciotto, Carla; Campesi, Franca; Schianchi, Giuseppe; Rocca, Stefano
2013-01-01
Investigating the innate immune response mediators released in milk has manifold implications, spanning from elucidation of the role played by mammary epithelial cells (MECs) in fighting microbial infections to the discovery of novel diagnostic markers for monitoring udder health in dairy animals. Here, we investigated the mammary gland response following a two-step experimental infection of lactating sheep with the mastitis-associated bacterium Streptococcus uberis. The establishment of infection was confirmed both clinically and by molecular methods, including PCR and fluorescent in situ hybridization of mammary tissues. Proteomic investigation of the milk fat globule (MFG), a complex vesicle released by lactating MECs, enabled detection of enrichment of several proteins involved in inflammation, chemotaxis of immune cells, and antimicrobial defense, including cathelicidins and calprotectin (S100A8/S100A9), in infected animals, suggesting the consistent involvement of MECs in the innate immune response to pathogens. The ability of MECs to produce and release antimicrobial and immune defense proteins was then demonstrated by immunohistochemistry and confocal immunomicroscopy of cathelicidin and the calprotectin subunit S100A9 on mammary tissues. The time course of their release in milk was also assessed by Western immunoblotting along the course of the experimental infection, revealing the rapid increase of these proteins in the MFG fraction in response to the presence of bacteria. Our results support an active role of MECs in the innate immune response of the mammary gland and provide new potential for the development of novel and more sensitive tools for monitoring mastitis in dairy animals. PMID:23774600
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Addis, Maria Filippa; Pisanu, Salvatore; Marogna, Gavino; Cubeddu, Tiziana; Pagnozzi, Daniela; Cacciotto, Carla; Campesi, Franca; Schianchi, Giuseppe; Rocca, Stefano; Uzzau, Sergio
2013-09-01
Investigating the innate immune response mediators released in milk has manifold implications, spanning from elucidation of the role played by mammary epithelial cells (MECs) in fighting microbial infections to the discovery of novel diagnostic markers for monitoring udder health in dairy animals. Here, we investigated the mammary gland response following a two-step experimental infection of lactating sheep with the mastitis-associated bacterium Streptococcus uberis. The establishment of infection was confirmed both clinically and by molecular methods, including PCR and fluorescent in situ hybridization of mammary tissues. Proteomic investigation of the milk fat globule (MFG), a complex vesicle released by lactating MECs, enabled detection of enrichment of several proteins involved in inflammation, chemotaxis of immune cells, and antimicrobial defense, including cathelicidins and calprotectin (S100A8/S100A9), in infected animals, suggesting the consistent involvement of MECs in the innate immune response to pathogens. The ability of MECs to produce and release antimicrobial and immune defense proteins was then demonstrated by immunohistochemistry and confocal immunomicroscopy of cathelicidin and the calprotectin subunit S100A9 on mammary tissues. The time course of their release in milk was also assessed by Western immunoblotting along the course of the experimental infection, revealing the rapid increase of these proteins in the MFG fraction in response to the presence of bacteria. Our results support an active role of MECs in the innate immune response of the mammary gland and provide new potential for the development of novel and more sensitive tools for monitoring mastitis in dairy animals.
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Bartels, Mette Damkjaer; Boye, Kit; Rohde, Susanne Mie; Larsen, Anders Rhod; Torfs, Herbert; Bouchy, Peggy; Skov, Robert; Westh, Henrik
2009-05-01
Rapid tests for detection of methicillin-resistant Staphylococcus aureus (MRSA) carriage are important to limit the transmission of MRSA in the health care setting. We evaluated the performance of the BD GeneOhm MRSA real-time PCR assay using a diverse collection of MRSA isolates, mainly from Copenhagen, Denmark, but also including international isolates, e.g., USA100-1100. Pure cultures of 349 MRSA isolates representing variants of staphylococcal cassette chromosome mec (SCCmec) types I to V and 103 different staphylococcal protein A (spa) types were tested. In addition, 53 methicillin-susceptible Staphylococcus aureus isolates were included as negative controls. Forty-four MRSA isolates were undetectable; of these, 95% harbored SCCmec type IVa, and these included the most-common clone in Copenhagen, spa t024-sequence type 8-IVa. The false-negative MRSA isolates were tested with new primers (analyte-specific reagent [ASR] BD GeneOhm MRSA assay) supplied by Becton Dickinson (BD). The ASR BD GeneOhm MRSA assay detected 42 of the 44 isolates that were false negative in the BD GeneOhm MRSA assay. Combining the BD GeneOhm MRSA assay with the ASR BD GeneOhm MRSA assay greatly improved the results, with only two MRSA isolates being false negative. The BD GeneOhm MRSA assay alone is not adequate for MRSA detection in Copenhagen, Denmark, as more than one-third of our MRSA isolates would not be detected. We recommend that the BD GeneOhm MRSA assay be evaluated against the local MRSA diversity before being established as a standard assay, and due to the constant evolution of SCCmec cassettes, a continuous global surveillance is advisable in order to update the assay as necessary.
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Shi, Huaiping; Shi, Hengbo; Luo, Jun; Wang, Wei; Haile, Abiel B; Xu, Huifen; Li, Jun
2014-07-01
Although research on dairy goat mammary gland have referred extensively to molecular mechanisms, research on lines of dairy goat mammary epithelial cells (MECs) are still rare. This paper sought to establish an immortal MEC line by stable transfection of human telomerase. MECs from a lactating (45 days post-parturition) Xinong Saanen dairy goat were cultured purely and subsequently transfected with a plasmid carrying the sequence of human telomerase. Immortalized MECs by human telomerase (hT-MECs) exhibited a typical cobblestone morphology and activity and expression levels of telomerase resembled that of MCF-7 cells. hT-MECs on passage 42 grew vigorously and 'S' sigmoid curves of growth were observed. Moreover, hT-MECs maintained a normal chromosome modal number of 2n=60, keratin 8 and epithelial membrane antigen (EMA) were evidently expressed, and beta-casein protein was synthesized and secreted. Beta-casein expression was enhanced by prolactin (P<0.05). Lipid droplets were found in hT-MECs, and messenger RNA levels of PPARG, SREBP, FASN, ACC and SCD in hT-MECs (passage 40) were similar to MECs (passage 7). In conclusion, the obtained hT-MEC line retained a normal morphology, growth characteristics, cytogenetics and secretory characteristics as primary MECs. Hence, it can be a representative model cell line, for molecular and functional analysis, of dairy goat MECs for an extended period of time. © 2014 Japanese Society of Animal Science.
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Mullen, Rachel D; Park, Soyoung; Rhodes, Simon J
2012-02-01
Lin-11, Isl-1, and Mec-3 (LIM)-homeodomain (HD)-class transcription factors are critical for many aspects of mammalian organogenesis. Of these, LHX3 is essential for pituitary gland and nervous system development. Pediatric patients with mutations in coding regions of the LHX3 gene have complex syndromes, including combined pituitary hormone deficiency and nervous system defects resulting in symptoms such as dwarfism, thyroid insufficiency, infertility, and developmental delay. The pathways underlying early pituitary development are poorly understood, and the mechanisms by which the LHX3 gene is regulated in vivo are not known. Using bioinformatic and transgenic mouse approaches, we show that multiple conserved enhancers downstream of the human LHX3 gene direct expression to the developing pituitary and spinal cord in a pattern consistent with endogenous LHX3 expression. Several transferable cis elements can individually guide nervous system expression. However, a single 180-bp minimal enhancer is sufficient to confer specific expression in the developing pituitary. Within this sequence, tandem binding sites recognized by the islet-1 (ISL1) LIM-HD protein are essential for enhancer activity in the pituitary and spine, and a pituitary homeobox 1 (PITX1) bicoid class HD element is required for spatial patterning in the developing pituitary. This study establishes ISL1 as a novel transcriptional regulator of LHX3 and describes a potential mechanism for regulation by PITX1. Moreover, these studies suggest models for analyses of the transcriptional pathways coordinating the expression of other LIM-HD genes and provide tools for the molecular analysis and genetic counseling of pediatric patients with combined pituitary hormone deficiency.
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Mullen, Rachel D.; Park, Soyoung
2012-01-01
Lin-11, Isl-1, and Mec-3 (LIM)-homeodomain (HD)-class transcription factors are critical for many aspects of mammalian organogenesis. Of these, LHX3 is essential for pituitary gland and nervous system development. Pediatric patients with mutations in coding regions of the LHX3 gene have complex syndromes, including combined pituitary hormone deficiency and nervous system defects resulting in symptoms such as dwarfism, thyroid insufficiency, infertility, and developmental delay. The pathways underlying early pituitary development are poorly understood, and the mechanisms by which the LHX3 gene is regulated in vivo are not known. Using bioinformatic and transgenic mouse approaches, we show that multiple conserved enhancers downstream of the human LHX3 gene direct expression to the developing pituitary and spinal cord in a pattern consistent with endogenous LHX3 expression. Several transferable cis elements can individually guide nervous system expression. However, a single 180-bp minimal enhancer is sufficient to confer specific expression in the developing pituitary. Within this sequence, tandem binding sites recognized by the islet-1 (ISL1) LIM-HD protein are essential for enhancer activity in the pituitary and spine, and a pituitary homeobox 1 (PITX1) bicoid class HD element is required for spatial patterning in the developing pituitary. This study establishes ISL1 as a novel transcriptional regulator of LHX3 and describes a potential mechanism for regulation by PITX1. Moreover, these studies suggest models for analyses of the transcriptional pathways coordinating the expression of other LIM-HD genes and provide tools for the molecular analysis and genetic counseling of pediatric patients with combined pituitary hormone deficiency. PMID:22194342
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Kilic, Abdullah; Basustaoglu, A Celal
2011-12-01
We developed and validated here a double triplex real-time PCR assay to simultaneously detect and identify Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus hominis, Staphylococcus haemolyticus and their methicillin resistance in a single reaction directly from Gram-positive cocci-in-clusters (GPCs)-positive blood culture bottles. From August 15, 2009 through February 15, 2010, 238 GPC-positive samples were collected and identified by conventional methods as 11 methicillin-resistant S. aureus (MRSA), 28 methicillin-susceptible S. aureus (MSSA), 176 MR coagulase-negative staphylococci (MRCoNS), 21 MSCoNS and two Enterococcus faecalis. The double triplex real-time PCR assay was targeted and detected tuf, nuc and mecA genes in the first tube and atlE, gap and mvaA genes in the second tube which could be run simultaneously. The detection limit of the assay was found at 10(3) CFU/ml for the atleE gene, 10(4) CFU/ml for the mva gene and 10(5) CFU/ml for gap, nuc, mecA and tuf genes based on seeding experiments. All Staphylococcus species except two S. epidermidis were correctly identified by the assay. The double triplex real-time PCR assay quickly and accurately detects S. aureus, S. epidermidis, S. hominis and S. haemolyticus and their methicillin resistance in a single reaction directly from positive blood culture bottles within 83 min. Copyright © 2011 Institut Pasteur. All rights reserved.
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Huang, Fengying; Meng, Qiuping; Tan, Guanghong; Huang, Yonghao; Wang, Hua; Mei, Wenli; Dai, Haofu
2011-06-01
To analysis and identify a bacterium strain isolated from laboratory breeding mouse far away from a hospital. Phenotype of the isolate was investigated by conventional microbiological methods, including Gram-staining, colony morphology, tests for haemolysis, catalase, coagulase, and antimicrobial susceptibility test. The mecA and 16S rRNA genes were amplified by the polymerase chain reaction (PCR) and sequenced. The base sequence of the PCR product was compared with known 16S rRNA gene sequences in the GenBank database by phylogenetic analysis and multiple sequence alignment. The isolate in this study was a gram positive, coagulase negative, and catalase positive coccus. The isolate was resistant to oxacillin, methicillin, penicillin, ampicillin, cefazolin, ciprofloxacin erythromycin, et al. PCR results indicated that the isolate was mecA gene positive and its 16S rRNA was 1 465 bp. Phylogenetic analysis of the resultant 16S rRNA indicated the isolate belonged to genus Saphylococcus, and multiple sequence alignment showed that the isolate was Saphylococcus haemolyticus with only one base difference from the corresponding 16S rRNA deposited in the GenBank. 16S rRNA gene sequencing is a suitable technique for non-specialist researchers. Laboratory animals are possible sources of lethal pathogens, and researchers must adapt protective measures when they manipulate animals. Copyright © 2011 Hainan Medical College. Published by Elsevier B.V. All rights reserved.
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Kim, Jae Seok; Kim, Han Sung; Song, Wonkeun; Cho, Hyoun Chan; Lee, Kyu Man; Kim, Eui Chong
2007-04-01
Many methicillin-resistant Staphylococcus aureus (MRSA) isolates in Korea possess a specific profile of staphylococcal enterotoxins in that the toxic shock syndrome toxin gene (tst) coexists with the staphylococcal enterotoxin C gene (sec). Because the analysis of staphylococcal cassette chromosome mec (SCCmec), a mobile genetic element mecA gene encoding methicillin resistance, showed that majority of these are SCCmec type II, these MRSA isolates with tst and sec may be genetically related with each other. This study was performed to investigate the genetic relatedness of tstand sec-harboring MRSA strains isolated in Korea by using pulsed-field gel electrophoresis (PFGE). A total of 59 strains of MRSA isolates of SCCmec type II possessing tst and sec were selected for PFGE and phylogenetic analyses. These isolates were collected from 13 health care facilities during nationwide surveillance of antimicrobial resistance in 2002. The 59 MRSA isolates were clustered into 11 PFGE types, including one major group of 26 strains (44.1%) isolated from 7 healthcare facilities. Seven PFGE types contained 2 or more isolates each, comprising 55 isolates in total. Most of SCCmec type II MRSA isolates containing tst and sec showed closely related PFGE patterns. Moreover, MRSA isolates collected from different healthcare facilities showed identical PFGE patterns. These findings suggested a clonal spread of MRSA strains possessing tst and sec in Korean hospitals.
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Mono- and Di-Alkylation Processes of DNA Bases by Nitrogen Mustard Mechlorethamine.
Larrañaga, Olatz; de Cózar, Abel; Cossío, Fernando P
2017-12-06
The reactivity of nitrogen mustard mechlorethamine (mec) with purine bases towards formation of mono- (G-mec and A-mec) and dialkylated (AA-mec, GG-mec and AG-mec) adducts has been studied using density functional theory (DFT). To gain a complete overview of DNA-alkylation processes, direct chloride substitution and formation through activated aziridinium species were considered as possible reaction paths for adduct formation. Our results confirm that DNA alkylation by mec occurs via aziridine intermediates instead of direct substitution. Consideration of explicit water molecules in conjunction with polarizable continuum model (PCM) was shown as an adequate computational method for a proper representation of the system. Moreover, Runge-Kutta numerical kinetic simulations including the possible bisadducts have been performed. These simulations predicted a product ratio of 83:17 of GG-mec and AG-mec diadducts, respectively. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
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Mohammed, Jibril; Ziwa, Michael Henry; Hounmanou, Yaovi Mahuton Gildas; Kisanga, Adela; Tuntufye, Huruma Nelwike
2018-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) in raw milk can be transmitted from animals to humans, and in Tanzania raw milk is sold in local markets and consumed as purchased. This study was performed to determine the molecular characteristics and antimicrobial susceptibility pattern of MRSA strains isolated from raw bovine milk sold at local markets in Tanzania. A total of 117 raw milk samples were cultured on Baird-Parker medium to isolate S. aureus and PCR was used for amplification of gltB gene for S. aureus identification and the presence of mecA gene for methicillin-resistant strains. Coagulase-negative (CN) S. aureus were reconfirmed using tube coagulase, DNase, and API Staph tests. MRSA isolates were spa typed whereas antimicrobial susceptibility testing was performed by the disc diffusion method. Forty-six coagulase positives (CP) and two CN S. aureus were identified. Most strains were resistant to penicillin (72%), and 3 isolates: 2 CN S. aureus and 1 coagulase-negative Staphylococci (CNS), were phenotypically resistant to vancomycin, oxacillin, and cefoxitin and were confirmed to carry mecA. Resistance to clindamycin, trimethoprim-sulfamethoxazole, and tetracycline was 23.9%, 30.4%, and 41.3%, respectively. Twelve isolates exhibited multidrug resistance; however, only one mecA positive strain among the three was typeable and belonged to spa type t2603. This study reports for the first time the presence of CN variant of MRSA, which was assigned the spa type t2603, and the presence of multidrug resistant S. aureus isolates from bovine milk in Morogoro, Tanzania.
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Mistry, Hiral; Sharma, Paresh; Mahato, Sudipta; Saravanan, R; Kumar, P Anand; Bhandari, Vasundhra
2016-01-01
Bovine mastitis caused by multidrug resistant Staphylococcus aureus is a huge problem reported worldwide, resulting in prolonged antibiotic treatment and death of livestock. The current study is focused on surveillance of antibiotic susceptibility along with genotypic and phenotypic characterization of the pathogenic S. aureus strains causing mastitis in India. One hundred and sixty seven milk samples were collected from mastitis-affected cows from different farms in India resulting in thirty nine isolated S. aureus strains. Antibiotic sensitivity profiling revealed the majority of the strains (n = 24) to be multidrug resistant and eleven strains showed reduced susceptibility to vancomycin (MICs = 2μg/ml). All strains were oxacillin sensitive, but 19 strains were positive for the mecA gene, which revealed the occurrence of oxacillin susceptible mecA positive strains (OS-MRSA) for the first time from India. Additionally, 32 strains were positive for the pvl gene, a virulence determinant; of these 17 were also OS-MRSA strains. Molecular characterization based on multilocus sequence typing (MLST), spa typing, agr typing and SCCmec classification revealed strains belonging to different groups. Moreover, strains showed spa types (t2526, t9602) and MLST sequence types, ST-72, ST-88 and ST-239 which have been earlier reported in human infections. The prevalence of OS-MRSA strains indicates the importance of including both the genetic and phenotypic tests in characterizing S. aureus strains. Increased genotypic variability with strain related to human infections and pvl positive isolates indicates a worrisome situation with the possibility of bilateral transfer.
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Mohammed, Jibril; Ziwa, Michael Henry; Kisanga, Adela; Tuntufye, Huruma Nelwike
2018-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) in raw milk can be transmitted from animals to humans, and in Tanzania raw milk is sold in local markets and consumed as purchased. This study was performed to determine the molecular characteristics and antimicrobial susceptibility pattern of MRSA strains isolated from raw bovine milk sold at local markets in Tanzania. A total of 117 raw milk samples were cultured on Baird-Parker medium to isolate S. aureus and PCR was used for amplification of gltB gene for S. aureus identification and the presence of mecA gene for methicillin-resistant strains. Coagulase-negative (CN) S. aureus were reconfirmed using tube coagulase, DNase, and API Staph tests. MRSA isolates were spa typed whereas antimicrobial susceptibility testing was performed by the disc diffusion method. Forty-six coagulase positives (CP) and two CN S. aureus were identified. Most strains were resistant to penicillin (72%), and 3 isolates: 2 CN S. aureus and 1 coagulase-negative Staphylococci (CNS), were phenotypically resistant to vancomycin, oxacillin, and cefoxitin and were confirmed to carry mecA. Resistance to clindamycin, trimethoprim-sulfamethoxazole, and tetracycline was 23.9%, 30.4%, and 41.3%, respectively. Twelve isolates exhibited multidrug resistance; however, only one mecA positive strain among the three was typeable and belonged to spa type t2603. This study reports for the first time the presence of CN variant of MRSA, which was assigned the spa type t2603, and the presence of multidrug resistant S. aureus isolates from bovine milk in Morogoro, Tanzania. PMID:29721021
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Ruppé, Etienne; Barbier, François; Mesli, Yasmine; Maiga, Aminata; Cojocaru, Radu; Benkhalfat, Mokhtar; Benchouk, Samia; Hassaine, Hafida; Maiga, Ibrahim; Diallo, Amadou; Koumaré, Abdel Karim; Ouattara, Kalilou; Soumaré, Sambou; Dufourcq, Jean-Baptiste; Nareth, Chhor; Sarthou, Jean-Louis; Andremont, Antoine; Ruimy, Raymond
2009-02-01
In staphylococci, methicillin (meticillin) resistance (MR) is mediated by the acquisition of the mecA gene, which is carried on the size and composition variable staphylococcal cassette chromosome mec (SCCmec). MR has been extensively studied in Staphylococcus aureus, but little is known about MR coagulase-negative staphylococci (MR-CoNS). Here, we describe the diversity of SCCmec structures in MR-CoNS from outpatients living in countries with contrasting environments: Algeria, Mali, Moldova, and Cambodia. Their MR-CoNS nasal carriage rates were 29, 17, 11, and 31%, respectively. Ninety-six MR-CoNS strains, comprising 75 (78%) Staphylococcus epidermidis strains, 19 (20%) Staphylococcus haemolyticus strains, 1 (1%) Staphylococcus hominis strain, and 1 (1%) Staphylococcus cohnii strain, were analyzed. Eighteen different SCCmec types were observed, with 28 identified as type IV (29%), 25 as type V (26%), and 1 as type III (1%). Fifteen strains (44%) were untypeable for their SCCmec. Thirty-four percent of MR-CoNS strains contained multiple ccr copies. Type IV and V SCCmec were preferentially associated with S. epidermidis and S. haemolyticus, respectively. MR-CoNS constitute a widespread and highly diversified MR reservoir in the community.
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Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) at ambient freshwater beaches
Fogarty, Lisa R.; Haack, Sheridan K.; Johnson, Heather E.; Brennan, Angela K.; Isaacs, Natasha M.; Spencer, Chelsea
2015-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) are a threat to human health worldwide, and although detected at marine beaches, they have been largely unstudied at freshwater beaches. Genes indicating S. aureus (SA; femA) and methicillin resistance (mecA) were detected at 11 and 12 of 13 US Great Lakes beaches and in 18% or 27% of 287 recreational water samples, respectively. Eight beaches had mecA + femA (potential MRSA) detections. During an intensive study, higher bather numbers, staphylococci concentrations, and femA detections were found in samples collected after noon than before noon. Local population density, beach cloud cover, and beach wave height were significantly correlated with SA or MRSA detection frequency. The Panton-Valentine leukocidin gene, associated with community-acquired MRSA, was detected in 12 out of 27 potential MRSA samples. The femA gene was detected less frequently at beaches that met US enterococci criteria or EU enterococci ‘excellent’ recreational water quality, but was not related to Escherichia coli-defined criteria. Escherichia coli is often the only indicator used to determine water quality at US beaches, given the economic and healthcare burden that can be associated with infections caused by SA and MRSA, monitoring of recreational waters for non-fecal bacteria such as staphylococci and/or SA may be warranted.
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Prevalence and Characterization of Staphylococcus aureus Strains in the Pork Chain Supply in Chile.
Velasco, Valeria; Vergara, José L; Bonilla, Ana M; Muñoz, Javier; Mallea, Alejandra; Vallejos, Diego; Quezada-Aguiluz, Mario; Campos, Jorge; Rojas-García, Pedro
2018-05-01
The detection of methicillin-resistant Staphylococcus aureus (MRSA) and other emerging strains in meat-producing animals and retail meat has increased the risk of contamination of food. The aim of this study was to determine the prevalence and characterize S. aureus strains isolated from the pork chain supply in Chile. A total of 487 samples were collected: 332 samples from pigs at farms and slaughterhouses (nasal, n = 155; skin, n = 177); 85 samples from carcasses at slaughterhouses; and 70 meat samples at supermarkets and retail stores. The isolation of S. aureus was carried out by selective enrichment and culture media. Biochemical testing (API ® Staph) and PCR (detection of the nuc and mecA genes) were used to confirm S. aureus and MRSA strains. The agglutination test was used to determine the protein PBP2'. Enterotoxins (SEA, SEB, SEC, SED) were determined by agglutination test and the se genes by PCR method. Oxacillin and cefoxitin susceptibility testing were carried out using the diffusion method. The overall prevalence of S. aureus in the pork meat supply was 33.9%. A higher prevalence was detected on carcasses (56.5%), in pigs sampled at farms (40.6%) than in pigs sampled at slaughterhouses (23.3%) and in nonpackaged retail meat (43.1%) than packaged retail meat (5.3%) (p ≤ 0.05). No significant differences (p > 0.05) were found between the prevalence in pigs (28.3%) and pork meat (32.9%) and between natural pig farming (33.3%) and conventional production (52.8%). The mecA gene and the protein PBP2' were not detected in S. aureus strains. Two S. aureus strains exhibited oxacillin and cefoxitin resistance, and one S. aureus strain was resistant to cefoxitin. One S. aureus strain isolated from a meat sample was positive for enterotoxin SEB. Although the mecA gene was not detected, oxacillin-resistant and seb-producing S. aureus strains were detected, which represent a risk in the pork chain supply.
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Khosravi, Azar Dokht; Jenabi, Atefeh; Montazeri, Effat Abbasi
2017-12-01
Today Methicillin-Resistant Staphylococcus aureus (MRSA) have acquired multiple resistance to a wide range of antibiotics including aminoglycosides. So, this study was aimed to investigate the rate of aminoglycoside resistance and the frequency of aminoglycoside resistance mediated genes of aac(Ia)-2, aph(3)-IIIa and ant(4')-Ia among MRSA strains. A total of 467 staphylococci isolates were collected from various clinical samples. S. aureus strains were identified by standard culture and identification criteria and investigating of presence of 16S rRNA and nuc genes. Cefoxitin disk diffusion, and oxacillin-salt agar screening methods were used to detect the MRSA strains with subsequent molecular identification for the presence of mecA gene. Antibiotic susceptibility of MRSA strains against aminoglycoside antibiotics was evaluated by using agar disk diffusion method. Multiplex PCR for the presence of aac(Ia)-2, aph(3)-IIIa and ant(4')-Ia encoding genes for aminoglycosides were performed for MRSA strains. From total staphylococci tested isolates, 262 (56.1%) were identified as S. aureus, of which 161 (61.45%) were detected as MRSA and all comprised mecA gene. The resistance pattern of MRSA strains to aminoglycoside antibiotics were: gentamicin 136 (84.5%); amikacin 125 (77.6%); kanamycin 139 (86.3%); tobramycin 132 (82%); and neomycin 155 (96.3%). The frequency of aac(Ia)-2, aph(3)-IIIa, and ant(4')-Ia genes among MRSA strains, were 64%, 42% and 11.8% respectively. In conclusion, as MRSA strains are of great concern in human infections, the results of present study could provide a useful resource for health sectors for choosing appropriate antibiotics for the effective treatment of infections due to MRSA strains. Copyright © 2017. Published by Elsevier Taiwan.
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Bulgari, Omar; Dong, Xianwen; Roca, Alfred L; Caroli, Anna M; Loor, Juan J
2017-01-01
Innate immune responses induced by in vitro stimulation of primary mammary epithelial cells (MEC) using Gram-negative lipopolysaccharide (LPS) and Gram-positive lipoteichoic acid (LTA) bacterial cell wall components are well- characterized in bovine species. The objective of the current study was to characterize the downstream regulation of the inflammatory response induced by Toll-like receptors in primary goat MEC (pgMEC). We performed quantitative real-time RT-PCR (qPCR) to measure mRNA levels of 9 genes involved in transcriptional regulation or antibacterial activity: Toll-like receptor 2 ( TLR2 ), Toll-like receptor 4 ( TLR4 ), prostaglandin-endoperoxide synthase 2 ( PTGS2 ), interferon induced protein with tetratricopeptide repeats 3 ( IFIT3 ), interferon regulatory factor 3 ( IRF3 ), myeloid differentiation primary response 88 ( MYD88 ), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 ( NFKB1 ), Toll interacting protein ( TOLLIP ), and lactoferrin ( LTF ). Furthermore, we analyzed 7 cytokines involved in Toll-like receptor signaling pathways: C-C motif chemokine ligand 2 ( CCL2 ), C-C motif chemokine ligand 5 ( CCL5 ), C-X-C motif chemokine ligand 6 ( CXCL6 ), interleukin 8 ( CXCL8 ), interleukin 1 beta ( IL1B ), interleukin 6 ( IL6 ), and tumor necrosis factor alpha ( TNF ). Stimulation of pgMEC with LPS for 3 h led to an increase in expression of CCL2 , CXCL6 , IL6 , CXCL8 , PTGS2 , IFIT3 , MYD88 , NFKB1 , and TLR4 ( P < 0.05). Except for IL6 , and PTGS2 , the same genes had greater expression than controls at 6 h post-LPS ( P < 0.05). Expression of CCL5 , PTGS2 , IFIT3 , NFKB1 , TLR4 , and TOLLIP was greater than controls after 3 h of incubation with LTA ( P < 0.05). Compared to controls, stimulation with LTA for 6 h led to greater expression of PTGS2 , IFIT3 , NFKB1 , and TOLLIP ( P < 0.05) whereas the expression of CXCL6 , CXCL8 , and TLR4 was lower ( P < 0.05). At 3 h incubation with both toxins compared to controls a greater expression ( P < 0.05) of CCL2 , CCL5 , CXCL6 , CXCL8 , IL6 , PTGS2 , IFIT3 , IRF3 , MYD88 , and NFKB1 was detected. After 6 h of incubation with both toxins, the expression of CCL2 , CXCL6 , IFIT3 , MYD88 , NFKB1 , and TLR4 was higher than the controls ( P < 0.05). Data indicate that in the goat MEC, LTA induces a weaker inflammatory response than LPS. This may be related to the observation that gram-positive bacteria cause chronic mastitis more often than gram-negative infections.
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Mahmoud, Mohamed; Parameswaran, Prathap; Torres, César I; Rittmann, Bruce E
2014-01-01
Pre-fermentation of poorly biodegradable landfill leachate (BOD5/COD ratio of 0.32) was evaluated for enhanced current density (j), Coulombic efficiency (CE), Coulombic recovery (CR), and removal of organics (BOD5 and COD) in a microbial electrolysis cell (MEC). During fermentation, the complex organic matter in the leachate was transformed to simple volatile fatty acids, particularly succinate and acetate in batch tests, but mostly acetate in semi-continuous fermentation. Carbohydrate had the highest degree of fermentation, followed by protein and lipids. j, CE, CR, and BOD5 removal were much greater for an MEC fed with fermented leachate (23 A/m(3) or 16 mA/m(2), 68%, 17.3%, and 83%, respectively) compared to raw leachate (2.5 A/m(3) or 1.7 mA/m(2), 56%, 2.1%, and 5.6%, respectively). All differences support the value of pre-fermentation before an MEC for stabilization of BOD5 and enhanced electron recovery as current when treating a recalcitrant wastewater like landfill leachate. Copyright © 2013 Elsevier Ltd. All rights reserved.
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Global Epigenetic Changes May Underlie Ethnic Differences and Susceptibility to Prostate Cancer
2013-09-01
malignancy and can often be found in non-cancerous tissues; in the prostate, hypermethylation of the GSTP1 CpG has been detected in PIA lesions [8]. DNA...methylcytosine (5-meC; [9, 10]). Since the recognition that the GSTP1 CpG island was frequently hypermethylated in PCa, more than 40 genes have been reported to...has also been identified for several genes. One study demonstrated that GSTP1 hypermethylation was significantly higher in PCa samples from AA men
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76 FR 7513 - Airworthiness Directives; The Boeing Company Model 747-400 and -400F Series Airplanes
Federal Register 2010, 2011, 2012, 2013, 2014
2011-02-10
.... Cracking in the MEC drip shield and exhaust plenum has been identified as part of the water leak path into the MEC. This condition, if not corrected, could result in water penetration into the MEC, which could... of cracked MEC drip shields. We are proposing this AD to prevent water penetration into the MEC...
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Ridler, Thomas; Matthews, Peter; Phillips, Keith G; Randall, Andrew D; Brown, Jonathan T
2018-06-01
The medial entorhinal cortex (mEC) has an important role in initiation and propagation of seizure activity. Several anatomical relationships exist in neurophysiological properties of mEC neurons; however, in the context of hyperexcitability, previous studies often considered it as a homogeneous structure. Using multi-site extracellular recording techniques, ictal-like activity was observed along the dorso-ventral axis of the mEC in vitro in response to various ictogenic stimuli. This originated predominantly from ventral areas, spreading to dorsal mEC with a surprisingly slow velocity. Modulation of inhibitory tone was capable of changing the slope of ictal initiation, suggesting seizure propagation behaviours are highly dependent on levels of GABAergic function in this region. A distinct disinhibition model also showed, in the absence of inhibition, a prevalence for interictal-like initiation in ventral mEC, reflecting the intrinsic differences in mEC neurons. These findings suggest the ventral mEC is more prone to hyperexcitable discharge than the dorsal mEC, which may be relevant under pathological conditions. The medial entorhinal cortex (mEC) has an important role in the generation and propagation of seizure activity. The organization of the mEC is such that a number of dorso-ventral relationships exist in neurophysiological properties of neurons. These range from intrinsic and synaptic properties to density of inhibitory connectivity. We examined the influence of these gradients on generation and propagation of epileptiform activity in the mEC. Using a 16-shank silicon probe array to record along the dorso-ventral axis of the mEC in vitro, we found 4-aminopyridine application produces ictal-like activity originating predominantly in ventral areas. This activity spreads to dorsal mEC at a surprisingly slow velocity (138 μm s -1 ), while cross-site interictal-like activity appeared relatively synchronous. We propose that ictal propagation is constrained by differential levels of GABAergic control since increasing (diazepam) or decreasing (Ro19-4603) GABA A receptor activation, respectively, reduced or increased the slope of ictal initiation. The observation that ictal activity is predominately generated in ventral mEC was replicated using a separate 0-Mg 2+ model of epileptiform activity in vitro. By using a distinct disinhibition model (co-application of kainate and picrotoxin) we show that additional physiological features (for example intrinsic properties of mEC neurons) still produce a prevalence for interictal-like initiation in ventral mEC. These findings suggest that the ventral mEC is more likely to initiate hyperexcitable discharges than the dorsal mEC, and that seizure propagation is highly dependent on levels of GABAergic expression across the mEC. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.
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Hansen, Thomas A; Bartels, Mette D; Høgh, Silje V; Dons, Lone E; Pedersen, Michael; Jensen, Thøger G; Kemp, Michael; Skov, Marianne N; Gumpert, Heidi; Worning, Peder; Westh, Henrik
2017-01-01
Staphylococcus argenteus ( S. argenteus ) is a newly identified Staphylococcus species that has been misidentified as Staphylococcus aureus ( S. aureus ) and is clinically relevant. We identified 25 S. argenteus genomes in our collection of whole genome sequenced S. aureus . These genomes were compared to publicly available genomes and a phylogeny revealed seven clusters corresponding to seven clonal complexes. The genome of S. argenteus was found to be different from the genome of S. aureus and a core genome analysis showed that ~33% of the total gene pool was shared between the two species, at 90% homology level. An assessment of mobile elements shows flow of SCC mec cassettes, plasmids, phages, and pathogenicity islands, between S. argenteus and S. aureus . This dataset emphasizes that S. argenteus and S. aureus are two separate species that share genetic material.
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Rong, Dongli; Wu, Qingping; Xu, Mingfang; Zhang, Jumei; Yu, Shubo
2017-01-01
Staphylococcus aureus is an important food-borne opportunistic pathogen that frequently causes severe blood and tissue infections or even fatal illnesses. Although S. aureus has been extensively studied in livestock and poultry foods in China, limited information has been reported in aquatic products. Accordingly, in this study, we aimed to characterize S. aureus in aquatic products purchased from retail markets in China. In total, 320 aquatic food samples were collected from 32 provincial capitals in China. The results showed that 119 samples (37.2%, 119/320) were positive for S. aureus by both qualitative and quantitative analyses. The contamination levels of 78.2% of samples ranged from 0.3 to 10 MPN/g, and six samples exceeded 110 MPN/g. A total of 119 S. aureus isolates from positive samples were selected to evaluate virulence factors, antibiotic resistance, and molecular characteristics. All S. aureus isolates were evaluated for the presence of 11 virulence genes by multiplex polymerase chain reaction, and α-hemolysin (hlα, 84.9%), fibronectin-binding protein A (fnbA, 79.0%), S. aureus enterotoxin E (see, 53.8%), and Panton-Valentine leucocidin (pvl, 50.4%) were identified as the major genes. These genes formed 56 different profiles, with the major profile identified as pvl-hlα-fnbA (28.6%). The antimicrobial susceptibility of all isolates was analyzed through the disk diffusion method, and the results showed high resistance to β-lactams, macrolides and tetracyclines, but susceptibility to linezolid and vancomycin. In addition, 26 sequence types (STs) were obtained via multilocus sequence typing, including seven novel STs, among which ST1 (20.2%), ST15 (18.5%), and ST188 (13.4%) were the most common STs. All the isolates were mecC negative, but nine isolates carrying mecA were evaluated by staphylococcal cassette chromosome mec (SCCmec) typing, all of which were SCCmecIII or SCCmecIV types. Isolates of SCCmecIII showed a high prevalence and were multidrug resistant. Our results showed that aquatic products could be a vehicle for transmission of virulence genes and multidrug-resistant S. aureus, representing a potential public health risk. The STs identified in this study indicated the genetic diversity of S. aureus, thereby providing important basic data for the dissemination of S. aureus in aquatic products. PMID:28473827
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Hari, Ananda Rao; Katuri, Krishna P; Gorron, Eduardo; Logan, Bruce E; Saikaly, Pascal E
2016-07-01
Microbial electrolysis cells (MECs) provide a viable approach for bioenergy generation from fermentable substrates such as propionate. However, the paths of electron flow during propionate oxidation in the anode of MECs are unknown. Here, the paths of electron flow involved in propionate oxidation in the anode of two-chambered MECs were examined at low (4.5 mM) and high (36 mM) propionate concentrations. Electron mass balances and microbial community analysis revealed that multiple paths of electron flow (via acetate/H2 or acetate/formate) to current could occur simultaneously during propionate oxidation regardless of the concentration tested. Current (57-96 %) was the largest electron sink and methane (0-2.3 %) production was relatively unimportant at both concentrations based on electron balances. At a low propionate concentration, reactors supplemented with 2-bromoethanesulfonate had slightly higher coulombic efficiencies than reactors lacking this methanogenesis inhibitor. However, an opposite trend was observed at high propionate concentration, where reactors supplemented with 2-bromoethanesulfonate had a lower coulombic efficiency and there was a greater percentage of electron loss (23.5 %) to undefined sinks compared to reactors without 2-bromoethanesulfonate (11.2 %). Propionate removal efficiencies were 98 % (low propionate concentration) and 78 % (high propionate concentration). Analysis of 16S rRNA gene pyrosequencing revealed the dominance of sequences most similar to Geobacter sulfurreducens PCA and G. sulfurreducens subsp. ethanolicus. Collectively, these results provide new insights on the paths of electron flow during propionate oxidation in the anode of MECs fed with low and high propionate concentrations.
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Khairalla, Ahmed S; Wasfi, Reham; Ashour, Hossam M
2017-08-07
There is limited data on methicillin-resistant Staphylococcus aureus (MRSA) carriage in dental clinics. 1300 specimens from patients, health personnel, and environmental surfaces of a dental clinic in Egypt were tested for MRSA. Antibiotic susceptibility, biofilm formation, Staphylococcal protein A (spa) typing, SCCmec typing, and PCR-based assays were used to detect mecA, mecC, vanA, Panton-Valentine Leukocidin toxin (PVL), and toxic shock syndrome toxin-1 (tst) genes. Among 34 mecA-positive MRSA isolates, five (14.7%) were PVL-positive, seventeen (50%) were tst-positive, ten (29.4%) were vanA-positive, while none harboured mecC. MRSA hand carriage rates in patients, nurses, and dentists were 9.8%, 6.6%, and 5%. The respective nasal colonization rates were 11.1%, 6.7%, and 9.7%. 1.3% of the environmental isolates were MRSA-positive. Strong and moderate biofilm-forming isolates represented 23.5% and 29.4% of MRSA isolates. 24 MRSA isolates (70.6%) were multi-resistant and 18 (52.9%) harboured SCCmec IV. Among eight spa types, t223 (26.5%), t267 (23.5%), and t14339 (23.5%) were predominant. We noted an alarming genetic relatedness between 7 (20.6%) MRSA isolates and the epidemic EMRSA-15 clone, as well as a combined occurrence of tst and PVL in 3 (8.8%) isolates. Results suggest high MRSA pathogenicity in dental wards highlighting the need for more efficient surveillance/infection control strategies.
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Zhang, Wanjiang; Hao, Zhihui; Wang, Yang; Cao, Xingyuan; Logue, Catherine M; Wang, Bing; Yang, Jing; Shen, Jianzhong; Wu, Congming
2011-11-01
The aim of this study was to determine the epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) isolates from pet animals and veterinary staff and the characteristics of these isolates. A total of 22 MRSA isolates were isolated from nasal swabs from dogs, cats and veterinary staff in six pet hospitals in six cities, and examined for antimicrobial susceptibility, the presence of resistance genes, Panton-Valentine leukocidin gene lukF-lukS, staphylococcal chromosomal cassette (SCC) mec typing, spa tying, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing. Of 22 MRSA isolates, 21 were recovered from pet animals, and one was isolated from a member of sstaff. All 22 MRSA strains were resistant to penicillin, oxacillin, azithromycin, clindamycin and ceftriaxone, and harboured mecA, ermB and linA genes. The lukF-lukS gene was not detected in any of the MRSA isolates. Eighteen MRSA strains from Qingdao belonged to ST59-MRSA-IV-spa t437. Of four MRSA isolates from Beijing, one belonged to ST398-MRSA-V-spa t034, and three belonged to ST239-MRSA-III-spa t030 profiles. Two PFGE types (A and B) were identified. Two isolates originating from dogs and one isolate originating from a staff member in Beijing shared similar PFGE patterns. Our cumulative data suggested that cross-transmission of MRSA may have occurred between pet animals and veterinary staff. Copyright © 2011 Elsevier Ltd. All rights reserved.
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Antimycobacterial potential of the juniper berry essential oil in tap water.
Peruč, Dolores; Gobin, Ivana; Abram, Maja; Broznić, Dalibor; Svalina, Tomislav; Štifter, Sanja; Staver, Mladenka Malenica; Tićac, Brigita
2018-03-01
Mycobacterium avium complex-related diseases are often associated with poorly maintained hot water systems. This calls for the development of new control strategies. The aim of this study was to investigate the activity of essential oils (EOs) from the Mediterranean plants, common juniper, immortelle, sage, lavandin, laurel, and white cedar against Mycobacterium avium ssp. avium, Mycobacterium intracellulare, and Mycobacterium gordonae in culturing broth and freshwater as their most common habitat. To do that, we developed a new method of water microdilution to determine their minimal effective concentrations (MEC). The most active EO was the one from the common juniper with the MEC of 1.6 mg mL-1. Gas chromatography / mass spectrometry the juniper EO identified monoterpenes (70.54 %) and sesquiterpenes (25.9 %) as dominant component groups. The main monoterpene hydrocarbons were α-pinene, sabinene, and β-pinene. The juniper EO significantly reduced the cell viability of M. intracellulare and M. gordonae at MEC, and of M. avium at 2xMEC. Microscopic analysis confirmed its inhibitory effect by revealing significant morphological changes in the cell membrane and cytoplasm of all three bacteria. The mode of action of the juniper EO on the cell membrane was confirmed by a marked leakage of intracellular material. Juniper EO has a great practical potential as a complementary or alternative water disinfectant in hot water systems such as baths, swimming pools, spa pools, hot tubs, or even foot baths/whirlpools.
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Predictors and Outcomes of Mealtime Emotional Climate in Families With Preschoolers.
Saltzman, Jaclyn A; Bost, Kelly K; Musaad, Salma M A; Fiese, Barbara H; Wiley, Angela R
2018-03-01
Mealtime emotional climate (MEC) is related to parent feeding and mental health, and possibly to child food consumption. However, MEC has been inconsistently assessed with a variety of coding schemes and self-report instruments, and has not been examined longitudinally. This study aims to characterize MEC systematically using an observational, count-based coding scheme; identify whether parent feeding or mental health predict MEC; and examine whether MEC predicts child food consumption and weight. A subsample of parents (n = 74) recruited from a larger study completed questionnaires when children were about 37 months, participated in a home visit to videotape a mealtime when children were about 41 months, and completed questionnaires again when children were about 51 months old. Maternal and child positive and negative emotions were coded from videotaped mealtimes. Observational data were submitted to cluster analyses, to identify dyads with similar emotion expression patterns, or MEC. Logistic regression was used to identify predictors of MEC, and Analysis of Covariance was used to examine differences between MEC groups. Dyads were characterized as either Positive Expressers (high positive, low negative emotion) or All Expressers (similar positive and negative emotion). Increased food involvement feeding practices were related to decreased likelihood of being an All Expresser. Positive Expressers reported that their children ate more healthy food, compared with All Expressers. Observed MEC is driven by maternal emotion, and may predict child food consumption. Food involvement may promote positive MEC. Improving MEC may increase child consumption of healthy foods.
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Wojewoda, Christina M.; Sercia, Linda; Navas, Maria; Tuohy, Marion; Wilson, Deborah; Hall, Geraldine S.; Procop, Gary W.
2013-01-01
Rapid identification of pathogens from blood cultures can decrease lengths of stay and improve patient outcomes. We evaluated the accuracy of the Verigene Gram-positive blood culture (BC-GP) nucleic acid test for investigational use only (Nanosphere, Inc., Northbrook, IL) for the identification of Gram-positive bacteria from blood cultures. The detection of resistance genes (mecA in Staphylococcus aureus and Staphylococcus epidermidis and vanA or vanB in Enterococcus faecium and Enterococcus faecalis) by the BC-GP assay also was assessed. A total of 186 positive blood cultures (in BacT/Alert FA bottles) with Gram-positive cocci observed with Gram staining were analyzed using the BC-GP assay. The BC-GP results were compared with the identification and susceptibility profiles obtained with routine methods in the clinical laboratory. Discordant results were arbitrated with additional biochemical, cefoxitin disk, and repeat BC-GP testing. The initial BC-GP organism identification was concordant with routine method results for 94.6% of the blood cultures. Only 40% of the Streptococcus pneumoniae identifications were correct. The detection of the mecA gene for 69 blood cultures with only S. aureus or S. epidermidis was concordant with susceptibility testing results. For 3 of 6 cultures with multiple Staphylococcus spp., mecA detection was reported but was correlated with oxacillin resistance in a species other than S. aureus or S. epidermidis. The detection of vanA agreed with susceptibility testing results for 45 of 46 cultures with E. faecalis or E. faecium. Comparison of the mean times to results for each organism group showed that BC-GP results were available 31 to 42 h earlier than phenotypic identifications and 41 to 50 h earlier than susceptibility results. PMID:23596240
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Wojewoda, Christina M; Sercia, Linda; Navas, Maria; Tuohy, Marion; Wilson, Deborah; Hall, Geraldine S; Procop, Gary W; Richter, Sandra S
2013-07-01
Rapid identification of pathogens from blood cultures can decrease lengths of stay and improve patient outcomes. We evaluated the accuracy of the Verigene Gram-positive blood culture (BC-GP) nucleic acid test for investigational use only (Nanosphere, Inc., Northbrook, IL) for the identification of Gram-positive bacteria from blood cultures. The detection of resistance genes (mecA in Staphylococcus aureus and Staphylococcus epidermidis and vanA or vanB in Enterococcus faecium and Enterococcus faecalis) by the BC-GP assay also was assessed. A total of 186 positive blood cultures (in BacT/Alert FA bottles) with Gram-positive cocci observed with Gram staining were analyzed using the BC-GP assay. The BC-GP results were compared with the identification and susceptibility profiles obtained with routine methods in the clinical laboratory. Discordant results were arbitrated with additional biochemical, cefoxitin disk, and repeat BC-GP testing. The initial BC-GP organism identification was concordant with routine method results for 94.6% of the blood cultures. Only 40% of the Streptococcus pneumoniae identifications were correct. The detection of the mecA gene for 69 blood cultures with only S. aureus or S. epidermidis was concordant with susceptibility testing results. For 3 of 6 cultures with multiple Staphylococcus spp., mecA detection was reported but was correlated with oxacillin resistance in a species other than S. aureus or S. epidermidis. The detection of vanA agreed with susceptibility testing results for 45 of 46 cultures with E. faecalis or E. faecium. Comparison of the mean times to results for each organism group showed that BC-GP results were available 31 to 42 h earlier than phenotypic identifications and 41 to 50 h earlier than susceptibility results.
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Montazeri, Effat Abbasi; Khosravi, Azar Dokht; Jolodar, Abbas; Ghaderpanah, Mozhgan; Azarpira, Samireh
2015-05-01
Methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative staphylococci (MRCoNS) as important human pathogens are causes of nosocomial infections worldwide. Burn patients are at a higher risk of local and systemic infections with these microorganisms. A screening method for MRSA by using a multiplex polymerase chain reaction (PCR) targeting the 16S ribosomal RNA (rRNA), mecA, and nuc genes was developed. The aim of the present study was to investigate the potential of this PCR assay for the detection of MRSA strains in samples from burn patients. During an 11-month period, 230 isolates (53.11%) of Staphylococcus spp. were collected from burn patients. The isolates were identified as S. aureus by using standard culture and biochemical tests. DNA was extracted from bacterial colonies and multiplex PCR was used to detect MRSA and MRCoNS strains. Of the staphylococci isolates, 149 (64.9%) were identified as S. aureus and 81 (35.21%) were described as CoNS. Among the latter, 51 (62.97%) were reported to be MRCoNS. From the total S. aureus isolates, 132 (88.6%) were detected as MRSA and 17 (11.4%) were methicillin-susceptible S. aureus (MSSA). The presence of the mecA gene in all isolates was confirmed by using multiplex PCR as a gold standard method. This study presented a high MRSA rate in the region under investigation. The 16S rRNA-mecA-nuc multiplex PCR is a good tool for the rapid characterization of MRSA strains. This paper emphasizes the need for preventive measures and choosing effective antimicrobials against MRSA and MRCoNS infections in the burn units. Copyright © 2014 Elsevier Ltd and ISBI. All rights reserved.
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Thulin, Elisabeth; Thulin, Måns; Andersson, Dan I
2017-09-01
Mecillinam (amdinocillin) is a β-lactam antibiotic used to treat uncomplicated urinary tract infections (UTIs). We have previously shown that inactivation of the Escherichia coli cysB gene is the major cause of mecillinam resistance (Mec R ) in clinical isolates. In this study, we used different E. coli strains (laboratory and clinical isolates) that were Mec R due to cysB mutations to determine how mecillinam susceptibility was affected during growth in urine compared to growth in the commonly used growth medium Mueller Hinton (MHB). We also examined mecillinam susceptibility when bacteria were grown in urine obtained from 48 different healthy volunteers. Metabolome analysis was done on the urine samples and the association between the mecillinam susceptibility patterns of the bacteria and urine metabolite levels was studied. Two major findings with clinical significance are reported. First, Mec R E. coli cysB mutant strains (both laboratory and clinical isolates) were always more susceptible to mecillinam when grown in urine as compared to laboratory medium, with many strains showing complete phenotypic susceptibility in urine. Second, the degree of reversion to susceptibility varied between urine samples obtained from different individuals. This difference was correlated with osmolality such that in urine with low osmolality the Mec R mutants were more susceptible to mecillinam than in urine with high osmolality. This is the first example describing conditional resistance where a genetically stable antibiotic resistance can be phenotypically reverted to susceptibility by metabolites present in urine. These findings have several important clinical implications regarding the use of mecillinam to treat UTIs. First, they suggest that mecillinam can be used to treat also those clinical strains that are identified as Mec R in standard laboratory tests. Second, the results suggest that testing of mecillinam susceptibility in the laboratory ought to be performed in media that mimics urine to obtain clinically relevant susceptibility testing results. Third, these findings imply that changes in patient behavior, such as increased water intake or use of diuretics to reduce urine osmolality and increased intake of cysteine, might induce antibiotic susceptibility in an infecting Mec R E. coli strain and thereby increase treatment efficiency. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.
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Liu, Wei; Zheng, Jian-Quan; Liu, Zhen-Wei; Li, Li-Jun; Wan, Qin; Liu, Chuan-Gui
2002-12-25
To compare the difference in action sites between mecamylamine (MEC) and hexamethonium (HEX) on nicotinic receptors of sympathetic neurons, we investigated the effects of MEC and HEX on the nicotine-induced currents in cultured superior cervical ganglion neurons by whole-cell patch clamp technique. The IC(50) of MEC and HEX for antagonizing the effect of 0.08 mmol/L nicotine was 0.0012 and 0.0095 mmol/L, respectively. Both MEC and HEX accelerated the desensitization of nicotinic receptors. Furthermore, by comparing their effects at holding potentials 30, 70 and 110 mV, it was indicated that their suppressing effect on the nicotine-induced currents was voltage-dependent. However, different from that of HEX, the inhibitory effect of MEC increased with administering the mixture of MEC and nicotine at intervals of 3 min, indicating a use-dependent effect of MEC. It is concluded that the action site of MEC on nicotinic receptors of sympathetic neurons is different from that of HEX.
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Yuan, Heyang; Lu, Yaobin; Abu-Reesh, Ibrahim M; He, Zhen
2015-01-01
While microbial electrolysis cells (MECs) can simultaneously produce bioelectrochemical hydrogen and treat wastewater, they consume considerable energy to overcome the unfavorable thermodynamics, which is not sustainable and economically feasible in practical applications. This study presents a proof-of-concept system in which hydrogen can be produced in an MEC powered by theoretically predicated energy from pressure-retarded osmosis (PRO). The system consists of a PRO unit that extracts high-quality water and generates electricity from water osmosis, and an MEC for organic removal and hydrogen production. The feasibility of the system was demonstrated using simulated PRO performance (in terms of energy production and effluent quality) and experimental MEC results (e.g., hydrogen production and organic removal). The PRO and MEC models were proven to be valid. The model predicted that the PRO unit could produce 485 mL of clean water and 579 J of energy with 600 mL of draw solution (0.8 M of NaCl). The amount of the predicated energy was applied to the MEC by a power supply, which drove the MEC to remove 93.7 % of the organic compounds and produce 32.8 mL of H2 experimentally. Increasing the PRO influent volume and draw concentration could produce more energy for the MEC operation, and correspondingly increase the MEC hydraulic retention time (HRT) and total hydrogen production. The models predicted that at an external voltage of 0.9 V, the MEC energy consumption reached the maximum PRO energy production. With a higher external voltage, the MEC energy consumption would exceed the PRO energy production, leading to negative effects on both organic removal and hydrogen production. The PRO-MEC system holds great promise in addressing water-energy nexus through organic removal, hydrogen production, and water recovery: (1) the PRO unit can reduce the volume of wastewater and extract clean water; (2) the PRO effluents can be further treated by the MEC; and (3) the osmotic energy harvested from the PRO unit can be applied to the MEC for sustainable bioelectrochemical hydrogen production.
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Yu, M-H; Chen, Y-G; Yu, Y-S; Chen, C-L; Li, L-J
2010-05-01
The aim of this study was to perform the molecular characterization of methicillin-resistant Staphylococcus haemolyticus (MRSH) from clinical specimens of patients in a Chinese hospital. One hundred and thirty-three strains of S. haemolyticus collected from April 2002 to April 2003 were analyzed. Antimicrobial susceptibility to 15 antimicrobial agents was determined by the broth microdilution method. The resistant rates to penicillin G and oxacillin were higher than 90%. There were no isolates resistant to linezolid or vancomycin, and only 6.0% of the strains were resistant to teicoplanin. The positivity rate for mecA genes was 90.2% by polymerase chain reaction (PCR). Ninety MRSH (isolated from inpatients and mecA-gene-positive) were genotyped by pulsed-field gel electrophoresis (PFGE) after SmaI digestion. Twenty-five different PFGE patterns (A approximately Y) were found and a major clone (type A; n = 36) with five subtypes was identified. Clone A was detected during a 1-year period. Identical PFGE types were found in different wards and patients. The results of this study suggest the clonal spread of MRSH within our hospital. This emphasizes the need for control and prevention measures.
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Ludden, C; Brennan, G; Morris, D; Austin, B; O'Connell, B; Cormican, M
2015-10-01
Methicillin-resistant Staphylococcus aureus (MRSA) is a major public health concern associated with residence in a long-term care facility (LTCF). The aim of this prospective study was to characterize MRSA isolated from residents over a 1-year period and their physical environment over a 2-year period. MRSA was recovered from 17/64 residents (R) of a LTCF and from 42 environmental (E) sites. All isolates carried the mecA gene and lacked the mecC and Panton-Valentine leukocidin (PVL) genes. Thirteen spa types were identified with t032 being the most frequent (41% of total; n = 8R, 16E), followed by t727 (22% of total; n = 13E), and t8783 (10% of total; n = 6E). Five spa types were each represented by single isolates. Thirty-nine isolates were of spa types associated with the multilocus sequence type ST22 (t032, 41%; spa-CC22, 68%) and reflect the predominance of ST22 in Irish hospitals. The uncommon spa types t727, t8783, t1372, t3130, t10038 were present in the environment but not detected in residents and are infrequently observed in Ireland.
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Tangeretin sensitizes SGS1-deficient cells by inducing DNA damage.
Chong, Shin Yen; Wu, Meng-Ying; Lo, Yi-Chen
2013-07-03
Tangeretin, a polymethoxyflavone found in citrus peel, has been shown to have antiatherogenic, anti-inflammatory, and anticarcinogenic properties. However, the underlying target pathways are not fully characterized. We investigated the tangeretin sensitivity of yeast (Saccharomyces cerevisiae) mutants for DNA damage response or repair pathways. We found that tangeretin treatment significantly reduced (p < 0.05) survival rate, induced preferential G1 phase accumulation, and elevated the DNA double-strand break (DSB) signal γH2A in DNA repair-defective sgs1Δ cells, but had no obvious effects on wild-type cells or mutants of the DNA damage checkpoint (including tel1Δ, sml1Δ mec1Δ, sml1Δ mec1Δ tel1Δ, and rad9Δ mutants). Additionally, microarray data indicated that tangeretin treatment up-regulates genes involved in nutritional processing and down-regulates genes related to RNA processing in sgs1Δ mutants. These results suggest tangeretin may sensitize SGS1-deficient cells by increasing a marker of DNA damage and by inducing G1 arrest and possibly metabolic stress. Thus, tangeretin may be suitable for chemosensitization of cancer cells lacking DSB-repair ability.
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In vivo targeting of de novo DNA methylation by histone modifications in yeast and mouse
Morselli, Marco; Pastor, William A; Montanini, Barbara; Nee, Kevin; Ferrari, Roberto; Fu, Kai; Bonora, Giancarlo; Rubbi, Liudmilla; Clark, Amander T; Ottonello, Simone; Jacobsen, Steven E; Pellegrini, Matteo
2015-01-01
Methylation of cytosines (5meC) is a widespread heritable DNA modification. During mammalian development, two global demethylation events are followed by waves of de novo DNA methylation. In vivo mechanisms of DNA methylation establishment are largely uncharacterized. Here, we use Saccharomyces cerevisiae as a system lacking DNA methylation to define the chromatin features influencing the activity of the murine DNMT3B. Our data demonstrate that DNMT3B and H3K4 methylation are mutually exclusive and that DNMT3B is co-localized with H3K36 methylated regions. In support of this observation, DNA methylation analysis in yeast strains without Set1 and Set2 shows an increase of relative 5meC levels at the transcription start site and a decrease in the gene-body, respectively. We extend our observation to the murine male germline, where H3K4me3 is strongly anti-correlated while H3K36me3 correlates with accelerated DNA methylation. These results show the importance of H3K36 methylation for gene-body DNA methylation in vivo. DOI: http://dx.doi.org/10.7554/eLife.06205.001 PMID:25848745
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Klaschik, Sven; Lehmann, Lutz E; Steinhagen, Folkert; Book, Malte; Molitor, Ernst; Hoeft, Andreas; Stueber, Frank
2015-03-01
Staphylococcus aureus has long been recognized as a major pathogen. Methicillin-resistant strains of S. aureus (MRSA) and methicillin-resistant strains of S. epidermidis (MRSE) are among the most prevalent multiresistant pathogens worldwide, frequently causing nosocomial and community-acquired infections. In the present pilot study, we tested a polymerase chain reaction (PCR) method to quickly differentiate Staphylococci and identify the mecA gene in a clinical setting. Compared to the conventional microbiology testing the real-time PCR assay had a higher detection rate for both S. aureus and coagulase-negative Staphylococci (CoNS; 55 vs. 32 for S. aureus and 63 vs. 24 for CoNS). Hands-on time preparing DNA, carrying out the PCR, and evaluating results was less than 5 h. The assay is largely automated, easy to adapt, and has been shown to be rapid and reliable. Fast detection and differentiation of S. aureus, CoNS, and the mecA gene by means of this real-time PCR protocol may help expedite therapeutic decision-making and enable earlier adequate antibiotic treatment. © 2014 Wiley Periodicals, Inc.
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Xu, Jia; Tan, Xiao; Zhang, Xinyu; Xia, Xiaoli; Sun, Huaichang
2015-11-01
Staphylococci are the leading pathogens of bovine mastitis which is difficult to control. However, the published data on the prevalence of staphylococcal species, virulence and antibiotic resistance genes in bovine mastitis from China are limited. In this study, 104 out of 209 subclinical mastitis milk samples from a single Chinese dairy herd were cultured-positive for staphylococci (49.8%), which were further identified as coagulase-positive staphylococci (CPS) or coagulase-negative staphylococci (CNS). According to the partial tuf and/or 16S rRNA gene sequence, the 28 CPS isolates were confirmed to be Staphylococcus aureus (26.9%), and 76 CNS isolates were assigned to 13 different species (73.1%) with Staphylococcus arlettae, Staphylococcus sciuri, Staphylococcus xylosus and Staphylococcus chromogenes as the dominant species. In the 28 S. aureus isolates, the most prevalent general virulence genes were coa, Ig and eno (100%), followed by hla (96.4%), hlb (92.9%), fib (92.9%), clfA (89.3%), clfB (85.7%) and nuc (85.7%). Both exotoxin and biofilm-associated genes were significantly less prevalent than the previously reported. Although 19 different virulence gene patterns were found, only one was dominant (32.1%). The prevalence of blaZ (82.1%) or mecA gene (35.7%) was much higher than the previously reported. In the 76 CNS isolates, the virulence genes were significantly less prevalent than that in the S. aureus isolates. Among the 4 main CNS species, S. chromogenes (n = 12) was the only species with high percentage (75%) of blaZ gene, while S. sciuri (n = 12) was the only species with the high percentage (66.7%) of mecA gene. The most of antibiotic resistance genes were present as multi-resistance genes, and the antibiotic resistances were attributed by different resistance genes between resistant S. aureus and CNS isolates. These data suggest that the prevalence of staphylococcal species, virulence and antibiotic resistance in the mastitis milk from the Chinese dairy herd are different from the previously reported, and that the herd- or farm-based diagnosis of staphylococcal bovine mastitis is required. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Motallebi, Mitra; Jabalameli, Fereshteh; Asadollahi, Kheirollah; Taherikalani, Morovat; Emaneini, Mohammad
2016-08-01
The emergence of antibiotic-resistant Staphylococcus aureus in particular methicillin-resistant S. aureus (MRSA) is an important concern in burn medical centers either in Iran or worldwide. A total of 128 S. aureus isolates were collected from wound infection of burn patients during June 2013 to June 2014. Multiplex-polymerase chain reaction (MPCR) assay was performed for the characterization of the staphylococcal cassette chromosome mec (SCCmec). Genes encoding virulence factors and biofilm were targeted by PCR. Of 128 S. aureus isolates, 77 (60.1%) isolates were MRSA. Fifty four (70.1%) isolates were identified as SCCmec type IIIA. The most frequently detected toxin genes among MRSA isolates with SCCmec type IIIA were sea (64.1%) and hla (51.8%). The rate of coexistence of sea with hla and sea with hla and hlb was 37% and12.9%, respectively. The sec, eta, tst, pvl, hla and hlb genes were not detected in any of the MRSA isolates. The most prevalent genes encoding biofilm was eno, found in 61.1% of isolates, followed by fib and icaA found in 48.1% and 38.8% of the isolates, respectively. The rate of coexistence of fib + eno + icaA + icaD and fib + eno was 20.3% and 9.2%, respectively. The ebps gene was not detected in any of the isolates. In conclusion, our study indicated that the sea, hla, fib and icaA were most frequent genes encoding virulence factors among MRSA with SCCmec type IIIA isolated from burn wound infection. Moreover, the results of this study shows that the rate of coexistence of genes encoding different virulence factor were high. Copyright © 2016 Elsevier Ltd. All rights reserved.
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External and middle ear sound pressure distribution and acoustic coupling to the tympanic membrane
Bergevin, Christopher; Olson, Elizabeth S.
2014-01-01
Sound energy is conveyed to the inner ear by the diaphanous, cone-shaped tympanic membrane (TM). The TM moves in a complex manner and transmits sound signals to the inner ear with high fidelity, pressure gain, and a short delay. Miniaturized sensors allowing high spatial resolution in small spaces and sensitivity to high frequencies were used to explore how pressure drives the TM. Salient findings are: (1) A substantial pressure drop exists across the TM, and varies in frequency from ∼10 to 30 dB. It thus appears reasonable to approximate the drive to the TM as being defined solely by the pressure in the ear canal (EC) close to the TM. (2) Within the middle ear cavity (MEC), spatial variations in sound pressure could vary by more than 20 dB, and the MEC pressure at certain locations/frequencies was as large as in the EC. (3) Spatial variations in pressure along the TM surface on the EC-side were typically less than 5 dB up to 50 kHz. Larger surface variations were observed on the MEC-side. PMID:24606269
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Bishop, Justin A; Yonescu, Raluca; Batista, Denise; Yemelyanova, Anna; Ha, Patrick K; Westra, William H
2014-01-01
High risk human papillomavirus (HPV) is firmly established as an important cause of oropharyngeal carcinoma. Recent studies have also implicated HPV as a cause of mucoepidermoid carcinoma (MEC)-a tumor of salivary gland origin that frequently harbors MAML2 translocations. The purpose of this study was to determine the prevalence of transcriptionally active HPV in a large group of MECs and to determine whether HPV infection and the MAML2 translocation are mutually exclusive events. Break-apart fluorescence in situ hybridization for MAML2 was performed on a tissue microarray containing 92 MECs. HPV testing was performed using RNA in situ hybridization targeting high risk HPV mRNA E6/E7 transcripts. Of the 71 MECs that could be evaluated by FISH, 57 (80 %) harbored the MAML2 rearrangement. HPV was not detected in any of the 57 MECs that contained a MAML2 rearrangement, in any of the 14 MECs that did not contain the rearrangement, or in any of the 21 MECs where MAML2 status was unknown. High risk HPV does not appear to play any significant role in the development of MEC. It neither complements nor replaces MAML2 translocation in the tumorigenesis of MEC.
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Kawabe, Yoshihiro; Mizobe, Kenich; Bando, Yasuhiko; Sakiyama, Koji; Taira, Fuyoko; Tomomura, Akito; Araki, Hisao; Amano, Osamu
2016-12-28
Myoepithelial cells (MECs) exist on the basal surface of acini in major exocrine glands, include myofilaments and various constructive proteins, and share characteristics with smooth muscle and epithelial cells. MECs project several ramified processes to invest acini, and possibly contract to compress acini to support the secretion by the glandular cells. However, the functional roles of MECs in salivary secretion are still unclear. We investigated morphological changes in immunostained MECs using the anti-α-smooth muscle actin (αSMA) antibody in operated or non-operated contralateral (NC) submandibular glands after partial or total resection. Furthermore, we investigated and discuss other salivary glands of rats. MECs in the parotid, sublingual and submandibular gland of adult rats exhibited different shapes and localizations. After surgery, in both operated and NC glands, the number of MECs and αSMA-immunopositive areas increased significantly. Three-dimensional analysis using a confocal laser-scanning microscope revealed that substantial and significant enhancement became evident in the number, length, and thickness of MEC-processes covering acini of the operated and NC submandibular glands. The preset findings indicate that MECs alter the morphology of their processes in operated and NC glands after surgery of the partial or total resection. It is suggested that MECs promote salivary secretion using elongated, thickened, and more ramified processes.
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Yorita, Kenji; Kuroda, Naoto; Naroda, Takushi; Tamura, Masato; Ohe, Chisato; Divatia, Mukul; Amin, Mahul B; Cubilla, Antonio L; Kazakov, Dimitry V; Hes, Ondrej; Michal, Michael; Michal, Michal
2018-04-01
Stratified mucin-producing intra-epithelial lesion (SMILE) and invasive stratified mucin-producing carcinoma (ISMC) are recently described cervical and penile lesions. We report an unusual case of mixed variant of penile squamous cell carcinomas with warty, usual and mucoepidermoid SMILE/ISMC features. A 62-year-old Japanese man had a glans penis lesion of one-and-a-half years' duration, suggesting malignancy. Partial penectomy and left inguinal lymphadenectomy were performed. Pathological evaluation revealed a mixed squamous cell carcinoma with warty, mucinous and usual features. The mucinous component resembled mucoepidermoid carcinoma (MEC) and SMILE/ISMC. Glandular differentiation was absent. All the diverse tumour components were negative for p16, which was confirmed by negative human papillomavirus (HPV) genotyping. The mucinous component was diffusely positive for cytokeratin 7 and largely negative for cytokeratin 5 and p63. Fluorescence in-situ hybridisation did not detect rearrangement in the MAML2 or EWSR1 genes. The tumour was pathological stage pT2, pN1 (AJCC prognostic stage group IIIA) and was disease-free 26 months after surgery. The lack of glands in the mucinous areas suggested that MEC should be separated from adenosquamous carcinoma (ASC). Penile SMILE/ISMC may occur without dependence upon HPV status. Further studies will be necessary to determine the pathogenesis and definition of penile SMILE/ISMC, the presence of true MEC arising from the glans penis and the clinicopathological differences of penile ASC, MEC and SMILE/ISMC. Herein, we refer to the SMILE-like penile lesion as 'mucinous penile intra-epithelial neoplasia'. © 2017 John Wiley & Sons Ltd.
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Forming system of strategic innovation management at high-tech engineering enterprises
NASA Astrophysics Data System (ADS)
Ergunova, O. T.; Lizunkov, V. G.; Malushko, E. Yu; Marchuk, V. I.; Ignatenko, A. Yu
2017-02-01
The article considers the processes of forming the strategic system of innovative activity management at the enterprises of a high-tech mechanical engineering complex (MEC) that are traditionally decisive in shaping the economic base of Russia. The authors proposed a method of designing a strategic system of innovative activity management at the MEC’s enterprises of the region which is based on the consideration of means and opportunities of the enterprise to implement intellectual activity. The proposed methodology and the empirical results constituted a ground for developing a system of strategic innovation management when performing the state-guaranteed order at the hi-tech MEC enterprises. Its implementation will help to reduce the level of uncertainty throughout the entire life cycle of an innovative activity product.
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Staphylococcus aureus is More Prevalent in Retail Beef Livers than in Pork and other Beef Cuts
Abdalrahman, Lubna S.; Wells, Harrington; Fakhr, Mohamed K.
2015-01-01
Staphylococcus aureus is one of the top five pathogens contributing to acquired foodborne illnesses causing an estimated quarter million cases every year in the US. The objectives of this study were to determine the prevalence of Methicillin Susceptible S. aureus (MSSA) and Methicillin Resistant S. aureus (MRSA) in retail beef livers, beef, and pork meats sold in Tulsa, Oklahoma and to characterize the recovered strains for their virulence and antimicrobial resistance. Ninety six chilled retail beef (50 beef livers and 46 beef other cuts), and 99 pork meat samples were collected. The prevalence in beef livers was 40/50 (80%) followed by other beef cuts 23/46 (50%) then pork 43/99 (43.3%). No isolates were positive for MRSA since none harbored the mecA or mecC gene. A total of 334 recovered S. aureus isolates (143 beef livers, 76 beef, and 115 pork isolates) were screened for their antimicrobial susceptibility against 16 different antimicrobials and their possession of 18 different toxin genes. Multidrug resistance was more prevalent in the pork isolates followed by beef then beef livers. The prevalence of enterotoxin genes such as seg, seh, and sei and the toxic shock syndrome gene tst was higher in the pork isolates than in the beef ones. The hemolysin genes, particularly hlb, were more prevalent in isolates from beef livers. Molecular typing of a subset of the recovered isolates showed that they are highly diverse where spa typing was more discriminatory than PFGE. The alarmingly high incidence of S. aureus in retail beef livers in this study should raise awareness about the food safety of such meat products. PMID:25927961
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Igbinosa, Etinosa O; Beshiru, Abeni; Akporehe, Lucy U; Oviasogie, Faith E; Igbinosa, Owen O
2016-09-24
The present study was designed to characterize methicillin-resistant staphylococci from raw meat. A total of 126 meat samples were obtained from open markets between February and April, 2015. Antimicrobial susceptibility testing was carried out using the disc diffusion method. Molecular profiling was conducted using 16S rRNA, mecA, nuc, and PVL gene signatures were detected by polymerase chain reaction assay. Fifty isolates of methicillin-resistant Staphylococcus spp. were detected in 26 (52%) pork, 14 (28%) beef and 10 (20%) chicken samples. The staphylococcal isolates were identified through partial 16S ribosomal ribonucleic acid (16S rRNA) nucleotide sequencing, and BLAST analysis of the gene sequence revealed 98%-100% staphylococcal similarity. All isolates from beef and chicken samples amplified the mecA gene, while 100% of the MRSA isolates amplified the PVL gene. The multidrug resistance profile (resistant to ≥1 antimicrobial agent in ≥3 classes of antimicrobial agents) of the staphylococcal isolates showed that 7 isolates were resistant to methicillin, penicillin, clindamycin, chloramphenicol, trimethoprim-sulfamethoxazole, kanamycin, amoxicillin, cloxacillin, erythromycin, vancomycin, and gentamycin. There was a significant regression effect from the multidrug-resistant profile on the number of isolates (p < 0.05) suggesting a consequence of the dissemination of resistant strains within bacterial populations. The findings of the present study indicate that raw meats in the Benin metropolis were possibly contaminated with pathogenic and multi-drug resistant staphylococci strains and therefore could constitute a risk to public health communities.
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Golitsina, Nina L; Danehy, Francis T; Fellows, Ross; Cretton-Scott, Erika; Standring, David N
2010-03-01
Nucleoside analogs are effective inhibitors of the hepatitis C virus (HCV) in the clinical setting. One such molecule, 2'-C-methyl-cytidine (2'-MeC), entered clinical development as NM283, a valine ester prodrug form of 2'-MeC possessing improved oral bioavailability. To be active against HCV, 2'-MeC must be converted to 2'-MeC triphosphate which inhibits NS5B, the HCV RNA-dependent RNA polymerase. Conversion of 2'-MeC to 2'-MeC monophosphate is the first step in 2'-MeC triphosphate production and is thought to be the rate-limiting step. Here we investigate which of three possible enzymes, deoxycytidine kinase (dCK), uridine-cytidine kinase 1 (UCK1), or uridine-cytidine kinase 2 (UCK2), mediate this first phosphorylation step. Purified recombinant enzymes UCK2 and dCK, but not UCK1, could phosphorylate 2'-MeC in vitro. However, siRNA knockdown experiments in three human cell lines (HeLa, Huh7 and HepG2) defined UCK2 and not dCK as the key kinase for the formation of 2'-MeC monophosphate in cultured human cells. These results underscore the importance of confirming enzymatic kinase data with appropriate cell-based assays. Finally, we present data suggesting that inefficient phosphorylation by UCK2 likely limits the antiviral activity of 2'-MeC against HCV. This paves the way for the use of a nucleotide prodrug approach to overcome this limitation.
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Castanheira, Mariana; Duncanson, Frederick P; Diekema, Daniel J; Guarro, Josep; Jones, Ronald N; Pfaller, Michael A
2012-01-01
Fusarium (n = 67) and Scedosporium (n = 63) clinical isolates were tested by two reference broth microdilution (BMD) methods against a novel broad-spectrum (active against both yeasts and molds) antifungal, E1210, and comparator agents. E1210 inhibits the inositol acylation step in glycophosphatidylinositol (GPI) biosynthesis, resulting in defects in fungal cell wall biosynthesis. Five species complex organisms/species of Fusarium (4 isolates unspeciated) and 28 Scedosporium apiospermum, 7 Scedosporium aurantiacum, and 28 Scedosporium prolificans species were identified by molecular techniques. Comparator antifungal agents included anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B. E1210 was highly active against all of the tested isolates, with minimum effective concentration (MEC)/MIC(90) values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B, respectively, for Fusarium of 0.12, >16, >16, >8, >8, 8, and 4 μg/ml. E1210 was very potent against the Scedosporium spp. tested. The E1210 MEC(90) was 0.12 μg/ml for S. apiospermum, but 1 to >8 μg/ml for other tested agents. Against S. aurantiacum, the MEC(50) for E1210 was 0.06 μg/ml versus 0.5 to >8 μg/ml for the comparators. Against S. prolificans, the MEC(90) for E1210 was only 0.12 μg/ml, compared to >4 μg/ml for amphotericin B and >8 μg/ml for itraconazole, posaconazole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparator agents. The essential agreement (EA; ±2 doubling dilutions) was >93% for all comparisons, with the exception of posaconazole and F. oxysporum species complex (SC) (60%), posaconazole and S. aurantiacum (85.7%), and voriconazole and S. aurantiacum (85.7%). In conclusion, E1210 exhibited very potent and broad-spectrum antifungal activity against azole- and amphotericin B-resistant strains of Fusarium spp. and Scedosporium spp. Furthermore, in vitro susceptibility testing of E1210 against isolates of Fusarium and Scedosporium may be accomplished using either of the CLSI or EUCAST BMD methods, each producing very similar results.
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Duncanson, Frederick P.; Diekema, Daniel J.; Guarro, Josep; Jones, Ronald N.; Pfaller, Michael A.
2012-01-01
Fusarium (n = 67) and Scedosporium (n = 63) clinical isolates were tested by two reference broth microdilution (BMD) methods against a novel broad-spectrum (active against both yeasts and molds) antifungal, E1210, and comparator agents. E1210 inhibits the inositol acylation step in glycophosphatidylinositol (GPI) biosynthesis, resulting in defects in fungal cell wall biosynthesis. Five species complex organisms/species of Fusarium (4 isolates unspeciated) and 28 Scedosporium apiospermum, 7 Scedosporium aurantiacum, and 28 Scedosporium prolificans species were identified by molecular techniques. Comparator antifungal agents included anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B. E1210 was highly active against all of the tested isolates, with minimum effective concentration (MEC)/MIC90 values (μg/ml) for E1210, anidulafungin, caspofungin, itraconazole, posaconazole, voriconazole, and amphotericin B, respectively, for Fusarium of 0.12, >16, >16, >8, >8, 8, and 4 μg/ml. E1210 was very potent against the Scedosporium spp. tested. The E1210 MEC90 was 0.12 μg/ml for S. apiospermum, but 1 to >8 μg/ml for other tested agents. Against S. aurantiacum, the MEC50 for E1210 was 0.06 μg/ml versus 0.5 to >8 μg/ml for the comparators. Against S. prolificans, the MEC90 for E1210 was only 0.12 μg/ml, compared to >4 μg/ml for amphotericin B and >8 μg/ml for itraconazole, posaconazole, and voriconazole. Both CLSI and EUCAST methods were highly concordant for E1210 and all comparator agents. The essential agreement (EA; ±2 doubling dilutions) was >93% for all comparisons, with the exception of posaconazole and F. oxysporum species complex (SC) (60%), posaconazole and S. aurantiacum (85.7%), and voriconazole and S. aurantiacum (85.7%). In conclusion, E1210 exhibited very potent and broad-spectrum antifungal activity against azole- and amphotericin B-resistant strains of Fusarium spp. and Scedosporium spp. Furthermore, in vitro susceptibility testing of E1210 against isolates of Fusarium and Scedosporium may be accomplished using either of the CLSI or EUCAST BMD methods, each producing very similar results. PMID:22083469
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Wang, Wei; Guo, Yunchang; Pei, Xiaoyan; Hu, Yujie; Bai, Li; Sun, Aiping; Liu, Jikai; Fu, Ping; Li, Fengqin
2013-11-01
To study the mecA gene distribution in 877 strains of Staphylococcus aureus isolated from the environment of pig farm and slaughter house, pig carcass and its iliac lymph nodes, and ready-to-eat foods in China as to screen the methicillin-resistant Staphylococcus aureus (MRSA), and to evaluate the antimicrobial susceptibility of MRSA. A total of 877 strains of S. aureus that had been phenotypically identified by Gram staining, catalase test, ability to coagulate rabbit plasma, API STAPH as well as analysis of nuc gene, encoding for a S. aureus specific thermonu-clease were screened for MRSA by characterizing the mecA gene. The antimicrobial susceptibility of MRSA was tested in accordance with the broth microdilution method recommended by the Clinical and Laboratory Standards Institute. Of 877 S. aureus strains tested, 71 (8.1%, 71/887) were mecA positive and identified as MRSA, among which, 48 isolates were pig-associated and 23 isolates were ready-to-eat food-associated. The frequency of pig-associated MRSA was significantly higher than that of food-associated one (chi2 = 53.040, P < 0.01). All MRSA were susceptible to linezolid, vancomycin, tigecycline, and nitrofurantoin but resistant to cefoxitin, oxacillin and benzylpenicillin. Meanwhile, 98.6% (70 strains), 95.8% (68 strains), 88.7% (63 strains), 80.3% (57 strains), 80.3% (57 strains) and 32.4% (23 strains) MRSA exhibited the resistance to clindamycin, erythromycin, tetracycline, ciprofloxacin, trimethoprim/sulfamethoxazole, and gentamicin, respectively. Besides, one strain was resistant to each of antibiotics including levofloxacin, moxifloxacin, rifampicin, and quinupristin/dalfopristin. It was worth noting that the frequency of resistance to ciprofloxacin, tetracycline, and trimethoprim/sulfamethoxazole of pig-associated MRSA was significantly higher than that of food-associated MRSA (CIP: chi2 = 29.110, P < 0.01, TET: chi2 = 18.816, P < 0.01, TMP/ SMZ: chi2 = 36.394, P < 0.01). It should be pointed out that 70 (98.6%) strains of MRSA were multi-drug resistant and eight spectrums of antimicrobial susceptibility were observed. The multi-drug resistant MRSA isolated from pig- and food-associated matrixin China is very serious.
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Osman, Kamelia M.; Amer, Aziza M.; Badr, Jihan M.; Helmy, Nashwa M.; Elhelw, Rehab A.; Orabi, Ahmed; Bakry, Magdy; Saad, Aalaa S. A.
2016-01-01
Methicillin-resistant Staphylococcus aureus (MRSA) have been found in various farm animal species throughout the world. Yet, methicillin-susceptible S. aureus (MSSA), methicillin-susceptible non-S. aureus (MS-NSA), and methicillin-resistant non-S. aureus (MR-NSA) were not investigated. Therefore, we persued to determine the diversity in their phenotypic virulence assay, phenotypic antimicrobial resistance profile and molecular characterization in one of the food chains in Egypt. Samples were collected during 2013 from beef meat at retail. Twenty seven isolates comprising five species (S. hyicus, S. aureus, S. schleiferi subsp. coagulans, S. intermedius, and S. lentus) were characterized for their antibiotic resistance phenotypic profile and antibiotic resistance genes (mecA, cfr, gyrA, gyrB, and grlA). Out of the 27 Staphylococcus isolates only one isolate was resistant to the 12 antibiotics representing nine classes. Raw beef meat sold across the Great Cairo zone, contains 66.7% of MRS, with highest prevalence was reported in S. aureus (66.7%), while the MRS non-S. aureus strains constituted 66.7% from which S. hyicus (60%), S. intermedius (33.3%), S. schleiferi subsp. coagulans (100%), and S. lentus (100%) were MRS. Seven S. aureus, six S. hyicus, four S. schleiferi subsp. coagulans, three S. intermedius, and one S. lentus isolates although being resistant to oxacillin yet, 11/27 (40.7%) carried the mecA gene. At the same time, the cfr gene was present in 2 of the nine S. aureus isolates, and totally undetectable in S. hyicus, S. schleiferi subsp. coagulans, S. intermedius, and S. lentus. Although, global researches largely focused into MRSA and MR-NSA in animals on pigs, the analysis of our results stipulates, that buffaloes and cattle could be MRSA dispersers and that this theme is not specific to pigs. Detection of MSSA virulence determinants is a must, as although oxacillin resistance may be absent yet, the MSSA may carry the virulence determinants which could be a source of perilous S. aureus for the human community. PMID:26973606
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A review on the applications of microbial electrolysis cells in anaerobic digestion.
Yu, Zhengsheng; Leng, Xiaoyun; Zhao, Shuai; Ji, Jing; Zhou, Tuoyu; Khan, Aman; Kakde, Apurva; Liu, Pu; Li, Xiangkai
2018-05-01
Anaerobic digestion (AD) has been widely used for biogas or biofuel generation from waste treatment. Because a low production rate and instability of AD occur frequently, various technologies have been applied to improvement of AD. Microbial electrolysis cells (MECs), an emerging technology, can convert organic matter into hydrogen, methane, and other value-added products. Recent studies showed that application of MEC to AD (MEC-AD) can accelerate degradation of a substrate (including recalcitrant compounds) and alter AD microbial community by enriching exoelectrogens and methanogens thus increasing biogas production. With stable microbial communities established, improvement of MEC-AD for methane production was achieved. MEC-AD process can be monitored in real-time by detecting electric signals, which linearly correlate with substrate concentrations. This review attempts to evaluate interactions among the decomposition of substrates, MEC-AD system, and the microbial community. This analysis should provide useful insights into the improvement of methane production and the performance of MEC-AD. Copyright © 2018 Elsevier Ltd. All rights reserved.
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Park, Jungyu; Lee, Beom; Tian, Donjie; Jun, Hangbae
2018-01-01
A microbial electrolysis cell (MEC) is a promising technology for enhancing biogas production from an anaerobic digestion (AD) reactor. In this study, the effects of the MEC on the rate of methane production from food waste were examined by comparing an AD reactor with an AD reactor combined with a MEC (AD+MEC). The use of the MEC accelerated methane production and stabilization via rapid organic oxidation and rapid methanogenesis. Over the total experimental period, the methane production rate and stabilization time of the AD+MEC reactor were approximately 1.7 and 4.0 times faster than those of the AD reactor. Interestingly however, at the final steady state, the methane yields of both the reactors were similar to the theoretical maximum methane yield. Based on these results, the MEC did not increase the methane yield over the theoretical value, but accelerated methane production and stabilization by bioelectrochemical reactions. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Yi, Cong; Tong, Jingjing; Lu, Puzhong; Wang, Yizheng; Zhang, Jinxie; Sun, Chen; Yuan, Kangning; Xue, Renyu; Zou, Bing; Li, Nianzhong; Xiao, Shuhua; Dai, Chong; Huang, Yuwei; Xu, Liling; Li, Lin; Chen, She; Miao, Di; Deng, Haiteng; Li, Hongliang; Yu, Li
2017-04-10
Autophagy is essential for maintaining glucose homeostasis, but the mechanism by which energy deprivation activates autophagy is not fully understood. We show that Mec1/ATR, a member of the DNA damage response pathway, is essential for glucose starvation-induced autophagy. Mec1, Atg13, Atg1, and the energy-sensing kinase Snf1 are recruited to mitochondria shortly after glucose starvation. Mec1 is recruited through the adaptor protein Ggc1. Snf1 phosphorylates Mec1 on the mitochondrial surface, leading to recruitment of Atg1 to mitochondria. Furthermore, the Snf1-mediated Mec1 phosphorylation and mitochondrial recruitment of Atg1 are essential for maintaining mitochondrial respiration during glucose starvation, and active mitochondrial respiration is required for energy deprivation-activated autophagy. Thus, formation of a Snf1-Mec1-Atg1 module on mitochondria governs energy deprivation-induced autophagy by regulating mitochondrial respiration. Copyright © 2017 Elsevier Inc. All rights reserved.
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Fijałkowski, Karol; Peitler, Dorota; Karakulska, Jolanta
2016-12-05
The aim of this study was to analyse the staphylococci isolated from ready-to-eat meat products, including pork ham, chicken cold cuts, pork sausage, salami and pork luncheon meat, sliced in the store to the consumer's specifications, along with species identification and determination of antibiotic resistance. Genes encoding staphylococcal enterotoxins, staphylococcal enterotoxin-like proteins, exfoliative toxins, and toxic shock syndrome toxin 1 were also investigated. From the 41 samples, 75 different staphylococcal isolates were obtained. Based on PCR-RFLP analysis of the gap gene using AluI and HpyCH4V restriction enzymes, the isolates were identified as Staphylococcus equorum (28%), S. vitulinus (16%), S. carnosus (14%), S. succinus (11%), S. xylosus (11%), S. saprophyticus (9%), S. warneri (9%), S. haemolyticus (1%) and S. pasteuri (1%). The incidence and number of resistances to antimicrobials was found to be species but not source of isolation dependent. All S. xylosus, S. saprophyticus, S. haemolyticus and S. pasteuri isolates showed antibiotic resistance. A lower percentage of resistance was recorded for S. warneri (71%) and S. vitulinus (58%), followed by S. equorum (57%), S. carnosus (50%) and S. succinus (50%). The most frequent resistance was observed to fusidic acid (43%). The mecA gene was amplified in 4% of the staphylococci. However, phenotypic resistance to methicillin was not confirmed in any of these isolates. On the other hand, the mecA gene was not detected in any of 9% of the isolates resistant to cefoxitin. It was also found that among 75 isolates, 60 (80%) harbored from 1 to 10 out of 21 analyzed superantigenic toxin genes. The most prevalent genes were: sei (36% isolates) among enterotoxins, seln (32% isolates) among enterotoxin-like proteins and eta encoding exfoliative toxin A (37% isolates). The findings of this study further extend previous observations that, when present in food, not only S. aureus but also other species of staphylococci could be of public health significance. Copyright © 2016 Elsevier B.V. All rights reserved.
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Wang, Wei; Baloch, Zulqarnain; Jiang, Tao; Zhang, Cunshan; Peng, Zixin; Li, Fengqin; Fanning, Séamus; Ma, Aiguo; Xu, Jin
2017-01-01
Staphylococcus aureus is one of the most common causes of zoonotic agent in the world, which are attributable to the contamination of food with enterotoxins. In this study, a total of 1,150 S. aureus isolates were cultured from 27,000 retail foods items from 203 cities of 24 provinces in China in 2015 and were test for antimicrobial susceptibility. Additionally, the role of the genes responsible for the staphylococcal enterotoxins (SEA to SEE), methicillin resistance (mecA) and the toxigenic capabilities were also assessed. The results showed that 4.3% retail foods were contaminated with S. aureus, and 7.9% retail foods isolates were mecA positive. Some 97.6% of S. aureus isolates were resistant to at least one antimicrobial compound, and 57.5% of these were multi drug resistant (MDR). Resistance to penicillin (83.7%, 963/1,150), was common, followed by linezolid (67.7%, 778/1,150) and erythromycin (52.1%, 599/1,150). The isolates cultured from raw meats showed high levels of resistant to tetracycline (42.8%), ciprofloxacin (17.4%), and chloramphenicol (12.0%) and expressed a MDR phenotype (62.4%). A total of 29.7% S. aureus isolates harbored the classical SEs genes (sea, seb, sec, and sed). The sea and seb genes were the most frequent SEs genes detected. Of note, 22% of the SEs genes positive S. aureus harbored two or three SEs genes, and 16 isolates were confirmed with the capacity to simultaneously produce two or three enterotoxin types. Moreover, nearly 50% of the MRSA isolates were positive for at least one SE gene in this study. Therefore, it is important to monitor the antimicrobial susceptibility and enterotoxigenicity of MDR S. aureus and MRSA in the food chain and to use these data to develop food safety measures, designed to reduce the contamination and transmission of this bacterium. PMID:29209290
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Wang, Wei; Baloch, Zulqarnain; Jiang, Tao; Zhang, Cunshan; Peng, Zixin; Li, Fengqin; Fanning, Séamus; Ma, Aiguo; Xu, Jin
2017-01-01
Staphylococcus aureus is one of the most common causes of zoonotic agent in the world, which are attributable to the contamination of food with enterotoxins. In this study, a total of 1,150 S. aureus isolates were cultured from 27,000 retail foods items from 203 cities of 24 provinces in China in 2015 and were test for antimicrobial susceptibility. Additionally, the role of the genes responsible for the staphylococcal enterotoxins (SEA to SEE), methicillin resistance ( mecA ) and the toxigenic capabilities were also assessed. The results showed that 4.3% retail foods were contaminated with S. aureus , and 7.9% retail foods isolates were mecA positive. Some 97.6% of S. aureus isolates were resistant to at least one antimicrobial compound, and 57.5% of these were multi drug resistant (MDR). Resistance to penicillin (83.7%, 963/1,150), was common, followed by linezolid (67.7%, 778/1,150) and erythromycin (52.1%, 599/1,150). The isolates cultured from raw meats showed high levels of resistant to tetracycline (42.8%), ciprofloxacin (17.4%), and chloramphenicol (12.0%) and expressed a MDR phenotype (62.4%). A total of 29.7% S. aureus isolates harbored the classical SEs genes ( sea, seb, sec , and sed ). The sea and seb genes were the most frequent SEs genes detected. Of note, 22% of the SEs genes positive S. aureus harbored two or three SEs genes, and 16 isolates were confirmed with the capacity to simultaneously produce two or three enterotoxin types. Moreover, nearly 50% of the MRSA isolates were positive for at least one SE gene in this study. Therefore, it is important to monitor the antimicrobial susceptibility and enterotoxigenicity of MDR S. aureus and MRSA in the food chain and to use these data to develop food safety measures, designed to reduce the contamination and transmission of this bacterium.
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Osaka, Shunsuke; Okuzumi, Katsuko; Koide, Shota; Tamai, Kiyoko; Sato, Tomoaki; Tanimoto, Koichi; Tomita, Haruyoshi; Suzuki, Masahiro; Nagano, Yukiko; Shibayama, Keigo; Arakawa, Yoshichika; Nagano, Noriyuki
2018-03-01
The decline in methicillin-resistant Staphylococcus aureus (MRSA) isolation rates has become a general observation worldwide, including Japan. We hypothesized that some genetic shift in MRSA might cause this phenomenon, and therefore we investigated the genetic profiles among MRSA clinical isolates obtained from three different epidemic phases in Japan. A total of 353 MRSA isolates were selected from 202 medical facilities in 1990 (pre-epidemic phase), 2004 (epidemic phase) and 2016 (post-epidemic phase). Molecular typing was performed by PCR detection of 22 genes using the polymerase chain reaction (PCR)-based ORF typing (POT) system, including an additional eight genes including small genomic islets and seven toxin genes. Isolates with a POT1 of score 93, identified as presumed clonal complex (pCC)5-staphylococcal cassette chromosome mec (SCCmec) type II including ST5-SCCmec type II New York/Japan clone, represented the major epidemic MRSA lineage in 1990 and 2004. In 2016, however, a marked decrease in isolates with a POT1 score of 93, along with changes in the epidemiology of toxin genes carried, was noted, where the carriers of tst genes including the tst-sec combination were markedly reduced, and those possessing the seb gene alone were markedly increased. Rather, isolates with a POT1 score of 106, including pCC1 or pCC8 among the isolates with SCCmec type IV, which often links to community-associated MRSA, were predominant. Interestingly, the pCC1 and pCC8 lineages were related to sea and tst-sec carriage, respectively. Over time, a transition in MRSA genetic profiles from a POT1 score of 93 in 1990 and 2004 to 106 in 2014 was found in Japan.
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mecC-Harboring Methicillin-Resistant Staphylococcus aureus: Hiding in Plain Sight.
Ford, Bradley A
2018-01-01
Previously there was scant data on the performance of laboratory testing to detect mecC -mediated beta-lactam resistance in Staphylococcus aureus Kriegeskorte and colleagues (J Clin Microbiol 56:e00826-17, 2018, https://doi.org/10.1128/JCM.00826-17) report the performance of various clinical tests for the detection of mecC -harboring methicillin-resistant S. aureus (MRSA), which failed to identify from 0 to 41% of tested mecC -harboring MRSA isolates. Changes in practice and new test development are necessary to address the challenge of mecC -harboring MRSA. Copyright © 2017 American Society for Microbiology.
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Badali, Hamid; Fakhim, Hamed; Zarei, Fereshteh; Nabili, Mojtaba; Vaezi, Afsane; Poorzad, Nafiseh; Dolatabadi, Somayeh; Mirhendi, Hossein
2016-04-01
Black aspergilli, particularly Aspergillus niger and A. tubingensis, are the most common etiological agents of otomycosis followed by onychomycosis, pulmonary aspergillosis and aspergilloma. However, so far there is no systematic study on their antifungal susceptibility profiles. A collection of 124 clinical and environmental species of black aspergilli consisted of A. niger, A. tubingensis, A. uvarum. A. acidus and A. sydowii were verified by DNA sequencing of the partial β-tubulin gene. MICs of amphotericin B, itraconazole, voriconazole, posaconazole, and MECs of caspofungin were performed based on CLSI M38-A2. Posaconazole and caspofungin had the lowest MIC range (0.016-0.125 µg/ml and 0.008-0.031 µg/ml, respectively), followed by amphotericin B (0.25-4 µg/ml), voriconazole (0.125-16 µg/ml) and itraconazole (0.25 to >16) in an increasing order. Some strains of A. niger showed high MIC value for itraconazole and voriconazole (>16 µg/ml), in contrast only environmental isolates of A. tubingensis had high itraconazole MICs (>16 µg/ml). These results confirm that posaconazole and caspofungin are potential drugs for treatment of aspergillosis due to opportunistic agents of Aspergillus Nigri complex. However, in vivo efficacy remains to be determined.
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Momoh, Asabe Halimat; Kwaga, Jacob K P; Bello, Mohammed; Sackey, Anthony K B; Larsen, Anders Rhod
2018-04-19
Staphylococcus aureus is a commensal and pathogenic bacterium with impact on public health and livestock industry. The study investigated nasal carriage, antibiotic resistance, and molecular characterization of S. aureus in pigs and pig workers. Nasal swabs from 300 backyard-raised pigs and 101 pig workers were used for the study. Resulting isolates were confirmed using MALDI-TOF MS, tested for antibiotic resistance, and three different multiplex PCRs were used to detect enterotoxin, mecA, spaA, scn, and pvl genes. spa typing was used to annotate the isolates into MLST clonal complexes (CC). Structured questionnaire was used to access possible risk factors for S. aureus carriage. The prevalence of S. aureus in pigs and pig workers were 5.3 and 12.9%, respectively. The isolates were resistant to beta-lactams (97%), tetracycline (62%), sulfonamide (52%), aminoglycoside (20.6%), fluoroquinolone (24%), and mupirocin (3.4%). Twenty seven (93%) of the isolates carried scn, 7(24%) pvl, and 12 (41%) enterotoxin genes, respectively. Questionnaire survey showed medical-related occupation of household members was associated (p < 0.5) with S. aureus carriage. This study suggests the presence of human multidrug resistant strains of S. aureus, high carriage of pvl, and enterotoxin genes, and CC5, CC15, and CC152 were the CC-groups shared among pigs and pig workers.
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Melo-Silveira, Raniere Fagundes; Fidelis, Gabriel Pereira; Viana, Rony Lucas Silva; Soeiro, Vinícius Campelo; Silva, Rodrigo Augusto da; Machado, Daisy; Costa, Leandro Silva; Ferreira, Carmen Veríssima; Oliveira Rocha, Hugo Alexandre
2014-04-24
Neglected agricultural products (NAPs) are defined as discarded material in agricultural production. Corn cobs are a major waste of agriculture maize. Here, a methanolic extract from corn cobs (MEC) was obtained. MEC contains phenolic compounds, protein, carbohydrates (1.4:0.001:0.001). We evaluated the in vitro and in vivo antioxidant potential of MEC. Furthermore, its antiproliferative property against tumor cells was assessed through MTT assays and proteins related to apoptosis in tumor cells were examined by western blot. MEC showed no hydroxyl radical scavenger capacity, but it showed antioxidant activity in Total Antioxidant Capacity and DPPH scavenger ability assays. MEC showed higher Reducing Power than ascorbic acid and exhibited high Superoxide Scavenging activity. In tumor cell culture, MEC increased catalase, metallothionein and superoxide dismutase expression in accordance with the antioxidant tests. In vivo antioxidant test, MEC restored SOD and CAT, decreased malondialdehyde activities and showed high Trolox Equivalent Antioxidant Capacity in animals treated with CCl4. Furthermore, MEC decreased HeLa cells viability by apoptosis due an increase of Bax/Bcl-2 ratio, caspase 3 active. Protein kinase C expression increased was also detected in treated tumor cells. Thus, our findings pointed out the biotechnological potential of corn cobs as a source of molecules with pharmacological activity.
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Structural Basis of Mec1-Ddc2-RPA Assembly and Activation on Single-Stranded DNA at Sites of Damage.
Deshpande, Ishan; Seeber, Andrew; Shimada, Kenji; Keusch, Jeremy J; Gut, Heinz; Gasser, Susan M
2017-10-19
Mec1-Ddc2 (ATR-ATRIP) is a key DNA-damage-sensing kinase that is recruited through the single-stranded (ss) DNA-binding replication protein A (RPA) to initiate the DNA damage checkpoint response. Activation of ATR-ATRIP in the absence of DNA damage is lethal. Therefore, it is important that damage-specific recruitment precedes kinase activation, which is achieved at least in part by Mec1-Ddc2 homodimerization. Here, we report a structural, biochemical, and functional characterization of the yeast Mec1-Ddc2-RPA assembly. High-resolution co-crystal structures of Ddc2-Rfa1 and Ddc2-Rfa1-t11 (K45E mutant) N termini and of the Ddc2 coiled-coil domain (CCD) provide insight into Mec1-Ddc2 homodimerization and damage-site targeting. Based on our structural and functional findings, we present a Mec1-Ddc2-RPA-ssDNA composite structural model. By way of validation, we show that RPA-dependent recruitment of Mec1-Ddc2 is crucial for maintaining its homodimeric state at ssDNA and that Ddc2's recruitment domain and CCD are important for Mec1-dependent survival of UV-light-induced DNA damage. Copyright © 2017 Elsevier Inc. All rights reserved.
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Staphylococcus argenteus from rabbits in Thailand.
Indrawattana, Nitaya; Pumipuntu, Natapol; Suriyakhun, Nawarat; Jangsangthong, Arunee; Kulpeanprasit, Suphang; Chantratita, Narisara; Sookrung, Nitat; Chaicumpa, Wanpen; Buranasinsup, Shutipen
2018-06-21
Staphylococcus argenteus, a novel species of the genus Staphylococcus or a member of the S. aureus complex, is closely related to S. aureus and is usually misidentified. In this study, the presence of S. argenteus in isolated S. aureus was investigated in 67 rabbits with abscess lesions during 2014-2016. Among 19 S. aureus complex isolates, three were confirmed to be S. argenteus by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, nonribosomal peptide synthetase gene amplification, and multilocus sequence type. All S. aureus complex isolates, including the S. aureus isolates, were examined for their antimicrobial resistance phenotype by disk diffusion and for their resistance genotype by PCR assays. Among the S. argenteus isolates, one was susceptible to all antimicrobial drugs and the other two were resistant to penicillin and doxycycline. In contrast, most S. aureus isolates were resistant to penicillin (37.5%), and gentamicin (12.5%). Moreover, S. aureus isolates harbored the blaZ, mecA, aacA-aphD, and mrs(A) as well as mutations of gyrA and grlA, but S. argenteus isolates carried solely the blaZ. S. argenteus isolates were investigated for enterotoxin (sea-sed) and virulence genes by PCR. One isolate carried sea, sec, and sed, whereas the other two isolates carried only sea or sed. No isolate carried seb and see. All three S. argenteus isolates carried hla, hlb, and clfA, followed by pvl, whereas coa, spa (IgG-binding region), and spa (x region) were not detected in the three isolates. This paper presents the first identification of S. argenteus from rabbits in Thailand. S. argenteus might be pathogenic because the isolates carried virulence genes. Moreover, antimicrobial resistance was observed. Investigations of this new bacterial species should be conducted in other animal species as well as in humans. © 2018 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.
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Jin, Xiaolu; Wang, Kai; Liu, Hongyun; Hu, Fuliang; Zhao, Fengqi; Liu, Jianxin
2016-01-01
The mammary epithelial cells (MECs) of high-producing dairy cows are likely to be subject to oxidative stress (OS) due to the intensive cell metabolism. The objectives of this study were to investigate the cytoprotective effects of resveratrol against hydrogen peroxide- (H2O2-) induced OS in cultured bovine MECs (MAC-T). Pretreatment of MAC-T cells with resveratrol could rescue the decrease in cell viability and resulted in lower intracellular reactive oxygen species (ROS) accumulation after H2O2 exposure. Resveratrol helped MAC-T cells to prevent H2O2-induced endoplasmic reticulum stress and mitochondria-related cell apoptosis. Moreover, resveratrol induced mRNA expression of multiple antioxidant defense genes in MAC-T cells under normal/oxidative conditions. Nuclear factor erythroid 2-related factor 2 (Nrf2) was required for the cytoprotective effects on MAC-T cells by resveratrol, as knockdown of Nrf2 significantly abolished resveratrol-induced cytoprotective effects against OS. In addition, by using selective inhibitors, we further confirmed that the induction of Nrf2 by resveratrol was mediated through the prolonged activation of PI3K/Akt and ERK/MAPK pathways but negatively regulated by p38/MAPK pathway. Overall, resveratrol has beneficial effects on bovine MECs redox balance and may be potentially used as a therapeutic medicine against oxidative insult in lactating animals.
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Feng, Wenyi; Bachant, Jeff; Collingwood, David; Raghuraman, M K; Brewer, Bonita J
2009-12-01
Yeast replication checkpoint mutants lose viability following transient exposure to hydroxyurea, a replication-impeding drug. In an effort to understand the basis for this lethality, we discovered that different events are responsible for inviability in checkpoint-deficient cells harboring mutations in the mec1 and rad53 genes. By monitoring genomewide replication dynamics of cells exposed to hydroxyurea, we show that cells with a checkpoint deficient allele of RAD53, rad53K227A, fail to duplicate centromeres. Following removal of the drug, however, rad53K227A cells recover substantial DNA replication, including replication through centromeres. Despite this recovery, the rad53K227A mutant fails to achieve biorientation of sister centromeres during recovery from hydroxyurea, leading to secondary activation of the spindle assembly checkpoint (SAC), aneuploidy, and lethal chromosome segregation errors. We demonstrate that cell lethality from this segregation defect could be partially remedied by reinforcing bipolar attachment. In contrast, cells with the mec1-1 sml1-1 mutations suffer from severely impaired replication resumption upon removal of hydroxyurea. mec1-1 sml1-1 cells can, however, duplicate at least some of their centromeres and achieve bipolar attachment, leading to abortive segregation and fragmentation of incompletely replicated chromosomes. Our results highlight the importance of replicating yeast centromeres early and reveal different mechanisms of cell death due to differences in replication fork progression.
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Zhang, Yifeng; Angelidaki, Irini
2014-06-01
Microbial electrolysis cells (MECs) are an electricity-mediated microbial bioelectrochemical technology, which is originally developed for high-efficiency biological hydrogen production from waste streams. Compared to traditional biological technologies, MECs can overcome thermodynamic limitations and achieve high-yield hydrogen production from wide range of organic matters at relatively mild conditions. This approach greatly reduces the electric energy cost for hydrogen production in contrast to direct water electrolysis. In addition to hydrogen production, MECs may also support several energetically unfavorable biological/chemical reactions. This unique advantage of MECs has led to several alternative applications such as chemicals synthesis, recalcitrant pollutants removal, resources recovery, bioelectrochemical research platform and biosensors, which have greatly broaden the application scopes of MECs. MECs are becoming a versatile platform technology and offer a new solution for emerging environmental issues related to waste streams treatment and energy and resource recovery. Different from previous reviews that mainly focus on hydrogen production, this paper provides an up-to-date review of all the new applications of MECs and their resulting performance, current challenges and prospects of future. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Spatial and temporal variation in efficiency of the Moore egg collector
Worthington, Thomas A.; Brewer, Shannon K.; Farless, Nicole
2013-01-01
The Moore egg collector (MEC) was developed for quantitative and nondestructive capture of semibuoyant fish eggs. Previous studies have indicated that capture efficiency of the MEC was low and the use of one device did not adequately represent the spatial distribution within the water column of egg surrogates (gellan beads) of pelagic broadcast-spawning cyprinids. The objective of this study was to assess whether use of multiple MECs showed differences in spatial and temporal distribution of bead catches. Capture efficiency of three MECs was tested at four 500-m sites on the South Canadian River, a Great Plains river in Oklahoma. For each trial, approximately 100,000 beads were released and mean capture efficiency was 0.47–2.16%. Kolmogorov–Smirnov tests indicated the spatial distributions of bead catches were different among multiple MECs at three of four sites. Temporal variability in timing of peak catches of gellan beads was also evident between MECs. We concluded that the use of multiple MECs is necessary to properly sample eggs of pelagic broadcast-spawning cyprinids.
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Abroo, Soleiman; Hosseini Jazani, Nima; Sharifi, Yaeghob
2017-07-01
Methicillin-resistant Staphylococcus aureus (MRSA) is a challenge for public health, and community-acquired (CA) infections seem to be increasing among people in different areas. A total of 700 healthy student volunteers residing in dormitories of universities in Urmia, Iran, were enrolled in this study. After identification of the isolates, antibiotic susceptibility, presence of mecA and pvl genes, and staphylococcal cassette chromosome mec (SCCmec) typing were evaluated. Nasal screening identified 137 (19.6%) carriers of S aureus, and 18 (13.14%) were MRSA isolates. The antimicrobial susceptibility patterns of isolates revealed high resistance to penicillin (93.4%). All isolates were sensitive to vancomycin. The SCCmec typing showed that most MRSA strains belonged to SCCmec type IV (n = 14; 77.8%). Only 1 (5.56%) MRSA isolates carried the pvl gene. Our findings revealed the relatively high frequency of S aureus nasal carriers and the advent of multidrug resistance among these isolates. Most MRSA isolates were SCCmec type IV; the transfer of such MRSA strains from carriers to other individuals in crowded living conditions such as dormitories can act as a risk factor for outbreak of CA MRSA and is a serious threat for the study groups. Copyright © 2017 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.
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Karmakar, Amit; Dua, Parimal; Ghosh, Chandradipa
2016-01-01
Staphylococcus aureus is opportunistic human as well as animal pathogen that causes a variety of diseases. A total of 100 Staphylococcus aureus isolates were obtained from clinical samples derived from hospitalized patients. The presumptive Staphylococcus aureus clinical isolates were identified phenotypically by different biochemical tests. Molecular identification was done by PCR using species specific 16S rRNA primer pairs and finally 100 isolates were found to be positive as Staphylococcus aureus. Screened isolates were further analyzed by several microbiological diagnostics tests including gelatin hydrolysis, protease, and lipase tests. It was found that 78%, 81%, and 51% isolates were positive for gelatin hydrolysis, protease, and lipase activities, respectively. Antibiogram analysis of isolated Staphylococcus aureus strains with respect to different antimicrobial agents revealed resistance pattern ranging from 57 to 96%. Our study also shows 70% strains to be MRSA, 54.3% as VRSA, and 54.3% as both MRSA and VRSA. All the identified isolates were subjected to detection of mecA, nuc, and hlb genes and 70%, 84%, and 40% were found to harbour mecA, nuc, and hlb genes, respectively. The current investigation is highly important and informative for the high level multidrug resistant Staphylococcus aureus infections inclusive also of methicillin and vancomycin.
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The Evaluation of Methicillin Resistance in Staphylococcus aboard the International Space Station
NASA Technical Reports Server (NTRS)
Ott, C. M.; Bassinger, V. J.; Fontenot, S. L.; Castro, V. A.; Pierson, D. L.
2005-01-01
The International Space Station (ISS) represents a semi-closed environment with a high level of crewmember interaction. As community-acquired methicillin-resistant Staphylococcus aureus (MRSA) has emerged as a health concern in environments with susceptible hosts in close proximity, an evaluation of isolates of clinical and environmental Staphylococcus aureus and coagulase negative Staphylococcus was performed to determine if this trend was also present in astronauts aboard ISS or the space station itself. Rep-PCR fingerprinting analysis of archived ISS isolates confirmed our earlier studies indicating a transfer of S. aureus between crewmembers. In addition, this fingerprinting also indicated a transfer between crewmembers and their environment. While a variety of S. aureus were identified from both the crewmembers and the environment, phenotypic evaluations indicated minimal methicillin resistance. However, positive results for the Penicillin Binding Protein, indicative of the presence of the mecA gene, were detected in multiple isolates of archived Staphylococcus epidermidis and Staphylococcus haemolyticus. Phenotypic analysis of these isolates confirmed their resistance to methicillin. While MRSA has not been isolated aboard ISS, the potential exists for the transfer of the gene, mecA, from coagulase negative environmental Staphylococcus to S. aureus creating MRSA strains. This study suggests the need to expand environmental monitoring aboard long duration exploration spacecraft to include antibiotic resistance profiling.
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NASA Astrophysics Data System (ADS)
Ebata, Takayuki
2017-06-01
The nonraddiative dececy route involving trans → cis photo-isomerization from the S_1 (ππ*) state has been investigated for several trans-cinnamate derivatives, which are known as sunscreen reagents. We examined two types of substitution effects. One is structural isomer such as ortho-, meta-, and para-hydroxy-methylcinnmate (o-, m-, p-HMC). The S_1 lifetime of p-HMC is less than 8 ps at zero-point level, and it undergoes rapid S_1 → ^1nπ* → T_1 decay via multiple conical intersections. Finally, the trans → cis isomerization proceeds in the T_1 state. On the other hand, both o- and m-HMC show very slow decay. Their S_1 lifetimes are in the order of 100 ps even at the excess energy of 2000-3000 \\wn. The other is the effect of the complexity of ester group in para-subsitituted species, such as para-methoxy-methyl, -ethyl and -2ethylhexyl cinnamate (p-MMC, p-MEC, p-M2EHC). p-MMC and p-MEC show sharp S_0 → S_1 (ππ*) vibronic bands, while p-M2EHC shows only broad structureless feature even under the jet-cooled condition. In addition, we found that the S_0 → ^1nπ* absorption appears at 1000 \\wn below the S_0 → S_1 (ππ*) transition in p-MEC and p-M2EHC, but not in p-MMC. Thus, the complexity of the ester group is very important for the appearance of the ^1nπ* state.
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He, GuangLin; Li, Ye; Zou, Xing; Li, Ping; Chen, PengYu; Song, Feng; Gao, Tianzhen; Liao, Miao; Yan, Jing; Wu, Jin
2017-09-01
The demographic characteristics and genetic polymorphism data of 56 Chinese nationalities or 31 administrative divisions in Chinese mainland have repeatedly been the genetic research hotspots. While most genetic studies focused on some particular Chinese populations based on autosomal or Y-chromosomal genetic markers, the forensic characteristics and phylogenetic analyses of the seventh largest Chinese population (Yi ethnicity) on the X-chromosomal genetic markers are scarce. Here, allele frequencies and forensic statistical parameters for 19 X-chromosomal short tandem repeat loci (DXS7424-DXS101, DXS6789-DXS6809, DXS7423-DXS10134, DXS10103-HPRTB-DXS10101, DXS10159-DXS10162-DXS10164, DXS10148-DXS10135-DXS8378, and DXS7132-DXS10079-DXS10074-DXS10075) of 331 Chinese Yi individuals were obtained. All 19 X-chromosomal short tandem repeat (STR) loci in females were consistent with the Hardy-Weinberg equilibrium test. A total of 214 alleles were identified with the corresponding allele frequencies spanned from 0.0019 to 0.6106. The combined PE, PDF, and PDM were 0.9999999214, 0.9999999999999999999993, and 0.9999999999998, respectively. The high combined MEC Krüger , MEC Kishida , MEC Desmarais , and MEC Desmarais Duo were achieved as 0.9999999617638, 0.9999999999971, 0.9999999999971, and 0.9999999931538, respectively. The findings suggested that the panel of 19 X-STR loci is highly polymorphic and informative in the Yi ethnic population and can be considered to be a powerful tool in forensic complex kinship identification. Population differentiation analyses among 12 populations indicated that significant differences in genetic structure were observed in between the Yi ethnicity and the Chinese Uyghur as well as Kazakh, and genetic homogeneity existed in similar ethno-origin or geographic origin populations.
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Staphylococcus aureus toxin gene hitchhikes on a transferable antibiotic resistance element.
Otto, Michael
2010-01-01
Virulence and antibiotic resistance of the dangerous human pathogen Staphylococcus aureus are to large extent determined by the acquisition of mobile genetic elements (MGEs). Up to now, these elements were known to comprise either resistance or virulence determinants, but not a mixture of the two. Queck et al. now found a cytolysin gene of the phenol-soluble modulin (PSM) family within SCCmec elements, which contain methicillin resistance genes and are largely responsible for the spread of methicillin-resistant S. aureus (MRSA). The novel gene, called psm-mec, had a significant impact on virulence in MRSA strains that do not produce high levels of genome-encoded PSMs. This first example of a combination of toxin and resistance genes on one staphylococcal MGE shows that such bundling is possible and may lead to an even faster acquisition of toxin and resistance genes by S. aureus and other staphylococcal pathogens.
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Brooks, John P; Adeli, Ardeshir; McLaughlin, Michael R
2014-06-15
The environmental influence of farm management in concentrated animal feeding operations (CAFO) can yield vast changes to the microbial biota and ecological structure of both the pig and waste manure lagoon wastewater. While some of these changes may not be negative, it is possible that CAFOs can enrich antibiotic resistant bacteria or pathogens based on farm type, thereby influencing the impact imparted by the land application of its respective wastewater. The purpose of this study was to measure the microbial constituents of swine-sow, -nursery, and -finisher farm manure lagoon wastewater and determine the changes induced by farm management. A total of 37 farms were visited in the Mid-South USA and analyzed for the genes 16S rRNA, spaQ (Salmonella spp.), Camp-16S (Campylobacter spp.), tetA, tetB, ermF, ermA, mecA, and intI using quantitative PCR. Additionally, 16S rRNA sequence libraries were created. Overall, it appeared that finisher farms were significantly different from nursery and sow farms in nearly all genes measured and in 16S rRNA clone libraries. Nearly all antibiotic resistance genes were detected in all farms. Interestingly, the mecA resistance gene (e.g. methicillin resistant Staphylococcus aureus) was below detection limits on most farms, and decreased as the pigs aged. Finisher farms generally had fewer antibiotic resistance genes, which corroborated previous phenotypic data; additionally, finisher farms produced a less diverse 16S rRNA sequence library. Comparisons of Camp-16S and spaQ GU (genomic unit) values to previous culture data demonstrated ratios from 10 to 10,000:1 depending on farm type, indicating viable but not cultivatable bacteria were dominant. The current study indicated that swine farm management schemes positively and negatively affect microbial and antibiotic resistant populations in CAFO wastewater which has future "downstream" implications from both an environmental and public health perspective. Published by Elsevier Ltd.
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Janjua, Omer Sefvan; Qureshi, Sana Mehmood; Khan, Tariq Sarfraz; Alamgir, Wajiha
2012-01-01
Mucoepidermoid carcinoma is the most common salivary gland tumor with varying behavior among different histopathological grades. The objective of this study was to determine the expression of Bcl-2 protein in mucoepidermoid carcinoma (MEC) and to correlate with histological grades. The records of 40 cases of MEC were collected from the histopathology department. Fresh slides were prepared and fresh diagnoses were made using the grading criteria for MEC. Immunohistochemical markers for Bcl-2 were applied and the results analyzed using the chi-square test. Of 40 cases, 20 were males and 20 were females. The range in age of the patients was 6 to 67 years mean (SD) was 42.6 (1.85) years. Twenty-two were low grade (55%), 11 high grade (27.5%) and 7 (17.5%) were intermediate grade MEC. Among these 40 cases, Bcl-2 expression was positive in 24 cases and negative in 16 cases. In 22 cases of low-grade MEC, 19 were positive while only 3 were negative. In high-grade tumors, all 11 cases were found to have a negative expression of Bcl-2 protein. In intermediate-grade MEC, 5 cases showed positive expression while only 2 cases showed negative expression. Bcl-2 protein expression showed positive expression in low-grade and negative expression in high-grade MEC. Intermediate grade showed more than 50% positive results for Bcl-2. Correlation between grades of MEC and expression of Bcl-2 is statistically significant and can be used for the depicting the prognosis of MEC along with other prognostic and clinico-pathological parameters.
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Lobo, A; Saz, P; Marcos, G; Día, J L; de la Cámara, C; Ventura, T; Morales Asín, F; Fernando Pascual, L; Montañés, J A; Aznar, S
1999-06-05
The revalidation of the Mini Examen Cognoscitivo (MEC), first Spanish version (1978) of the Mini-Mental Status Examination (MMSE) and documentation of "population-based norms" should clarify the potential confusion induced by later versions of MMSE. The Zaragoza Study on the prevalence of dementia and depression in a representative sample of the elderly community (N = 1,080). MEC-35 and MEC-30 points, and validated, Spanish versions of Geriatric Mental State (GMS), History and Aetiology Schedule (HAS) and Social Status Schedule (SSS). a) validation of MEC (standardized lay interviewers) against the gold standard of psychiatric diagnosis (DSM-III-R), two months later; b) "population-based norms" in the "healthy" population, and c) comparison with other MMSE versions. The instrument fulfills criteria of "feasibility", "content", "procedural" and "construct validity". Test-retest reliability: weighted kappa = 0.637. MEC-30 (cut-off point 23/24), sensitivity = 89.8%, specificity = 75.1% (80.8% with the cut-off at 22/23), and ROC curve, AUC = 0.920. The coefficients of individual items were satisfactory and the specificity increases in MEC-35 (83.9%). Other MMSE Spanish versions have not improved these coefficients. "Population-based norms" confirm the hypothesized influence of age and education level. MEC-30 is the version with most comparable results with the MMSE in USA. The validity of MEC is confirmed in the elderly population, with the same cut-off points recommended in the original standardization. MEC-30 is the best version for international comparisons.
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Kamal, Noura M; Salem, Hend M; Dahmoush, Heba M
2017-07-01
Mucoepidermoid carcinoma (MEC) is the most common malignant salivary gland tumor which displays biological, histological and clinical diversity thus representing a challenge for its diagnosis and management. Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein identified as a tumor specific antigen due to its frequent overexpression in the majority of epithelial carcinomas and its correlation with prognosis. It is considered to be a promising biomarker used as a therapeutic target already in ongoing clinical trials. The purpose of this study was to investigate the pattern, cellular characterization and level of EpCAM expression in MEC and demonstrate its correlation with histologic grading which may benefit future clinical trials using EpCAM targeted therapy. 48 specimens (12 normal salivary gland tissue and 36 MEC) were collected and EpCAM membranous expression was evaluated by immunohistochemistry. Total immunoscore (TIS) was evaluated, the term 'EpCAM overexpression' was given for tissues showing a total immunoscore >4. A highly significant difference was observed between TIS percent values in control and different grades of MEC (p<0.001). High grade MEC (HG-MEC) was the highest EpCAM expressor. In addition, EpCAM expression pattern differed among the different grades. EpCAM expression was detected in MEC, and its overexpression correlated with increasing the histological grade. The diffuse membranous expression in HG-MEC could be of diagnostic value in relation to the patchy expression observed in both low grade and intermediate grade MEC. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Materials experiment carrier concepts definition study. Volume 3: Programmatics, part 2
NASA Technical Reports Server (NTRS)
1981-01-01
Project logic, schedule and funding information was derived to enable decisions to be made regarding implementation of MEC system development. A master schedule and cost and price estimates (ROM) were developed for a project that consists of development of an all-up MEC, its integration with payloads and its flight on one 90 day mission. In Part 2 of the study a simple initial MEC was defined to accommodate three MPS baseline payloads. The design of this initial MEC is illustrated. The project logic, detailed schedules, and ROM cost estimate relate to a project in which this initial MEC is developed, integrated with payloads and flown once for 180 days.
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0ħω MEC effect on M1 properties of middle pf-shell nuclei
NASA Astrophysics Data System (ADS)
Nakada, H.; Sebe, T.
1994-09-01
M1 properties of middle pf-shell nuclei are studied within the framework of a large-scale shell-model calculation, by including two-body operators originating from the MEC effect within the 0ħω space. This MEC effect tends to enhance the M1 matrix elements slightly. However, the 0ħω MEC effect does not change the previous results so much, which have shown notable quenching in the magnetic moments of 55Co and 57Ni due to the 0ħω CP effect, while the 0ħω MEC effect should be kept track of in discussing the M1 properties with ⪅ 10% accuracy.
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Karbuz, Adem; Karahan, Zeynep Ceren; Aldemir-Kocabaş, Bilge; Tekeli, Alper; Özdemir, Halil; Güriz, Haluk; Gökdemir, Refik; İnce, Erdal; Çiftçi, Ergin
2017-01-01
Karbuz A, Karahan ZC, Aldemir-Kocabaş B, Tekeli A, Özdemir H, Güriz H, Gökdemir R, İnce E, Çiftçi E. Evaluation of antimicrobial susceptibilities and virulence factors of Staphylococcus aureus strains isolated from community-acquired and health-care associated pediatric infections. Turk J Pediatr 2017; 59: 395-403. The aim of this study was to investigate the enterotoxins and Panton-Valentine leukocidin (PVL) gene as virulence factor, identification if antimicrobial sensitivity patterns, agr (accessory gene regulator) types and sequence types and in resistant cases to obtain SCCmec (staphylococcal cassette chromosome mec) gene types which will be helpful to decide empirical therapy and future health politics for S. aureus species. Total of 150 isolates of S. aureus were isolated from the cultures of the child patients in January 2011 and December 2012. In this study, the penicillin resistance was observed as 93.8%. PVL and mecA was detected positive in 8.7% and in 6% of all S. aureus strains, respectively. Two MRSA (methicillin resistant S.aureus) strains were detected as SCCmec type III and SCCmec type V and five MRSA strains were detected as SCCmec type IV. SET-I and SET-G were the most common detected enterotoxins. In both community-associated and healthcare-associated MRSA strains, agr type 1 was detected most commonly. The most common sequence types were ST737 in 13 patients than ST22 in eight patients and ST121 in six patients. This study highlights a necessity to review the cause of small changes in the structural genes in order to determine whether it is a cause or outcome; community-acquired and healthcare associated strains overlap.
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Nobrega, Diego B.; Naushad, Sohail; Naqvi, S. Ali; Condas, Larissa A. Z.; Saini, Vineet; Kastelic, John P.; Luby, Christopher; De Buck, Jeroen; Barkema, Herman W.
2018-01-01
Emergence and spread of antimicrobial resistance is a major concern for the dairy industry worldwide. Objectives were to determine: (1) phenotypic and genotypic prevalence of drug-specific resistance for 25 species of non-aureus staphylococci, and (2) associations between presence of resistance determinants and antimicrobial resistance. Broth micro-dilution was used to determine resistance profiles for 1,702 isolates from 89 dairy herds. Additionally, 405 isolates were sequenced to screen for resistance determinants. Antimicrobial resistance was clearly species-dependent. Resistance to quinupristin/dalfopristin was common in Staphylococcus gallinarum (prevalence of 98%), whereas S. cohnii and S. arlettae were frequently resistant to erythromycin (prevalence of 63 and 100%, respectively). Prevalence of resistance was 10% against β-lactams and tetracyclines. In contrast, resistance to antimicrobials critically important for human medicine, namely vancomycin, fluoroquinolones, linezolid and daptomycin, was uncommon (< 1%). Genes encoding multidrug-resistance efflux pumps and resistance-associated residues in deducted amino acid sequences of the folP gene were the most frequent mechanisms of resistance, regardless of species. The estimated prevalence of the mecA gene was 17% for S. epidermidis. Several genes, including blaZ, mecA, fexA, erm, mphC, msrA, and tet were associated with drug-specific resistance, whereas other elements were not. There were specific residues in gyrB for all isolates of species intrinsically resistant to novobiocin. This study provided consensus protein sequences of key elements previously associated with resistance for 25 species of non-aureus staphylococci from dairy cattle. These results will be important for evaluating effects of interventions in antimicrobial use in Canadian dairy herds. PMID:29503642
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STS-99 Technicians work in Endeavour's aft compartment of the payload bay
NASA Technical Reports Server (NTRS)
2000-01-01
Technicians work in the aft compartment of Shuttle Endeavour's payload bay, where a new Enhanced Main Events Controller (E-MEC) will be installed. The original E-MEC in Endeavour became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. Both E-MECs are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. Before workers can begin E-MEC replacement efforts at the launch pad, cryogenic reactants had to be offloaded from the orbiter and Space Shuttle ordnance disconnected. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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The Caenorhabditis elegans LET-418/Mi2 plays a conserved role in lifespan regulation.
De Vaux, Véronique; Pfefferli, Catherine; Passannante, Myriam; Belhaj, Khaoula; von Essen, Alina; Sprecher, Simon G; Müller, Fritz; Wicky, Chantal
2013-12-01
The evolutionarily conserved nucleosome-remodeling protein Mi2 is involved in transcriptional repression during development in various model systems, plays a role in embryonic patterning and germ line development, and participates in DNA repair and cell cycle progression. It is the catalytic subunit of the nucleosome remodeling and histone deacetylase (NuRD) complex, a key determinant of differentiation in mammalian embryonic stem cells. In addition, the Drosophila and C. elegans Mi2 homologs participate in another complex, the MEC complex, which also plays an important developmental role in these organisms. Here we show a new and unexpected feature of the C. elegans Mi2 homolog, LET-418/Mi2. Lack of LET-418/Mi2 results in longevity and enhanced stress resistance, a feature that we found to be conserved in Drosophila and in Arabidopsis. The fact that depletion of other components of the NuRD and the MEC complexes did not result in longevity suggests that LET-418 may regulate lifespan in a different molecular context. Genetic interaction studies suggest that let-418 could act in the germ-cell-loss pathway, downstream of kri-1 and tcer-1. On the basis of our data and on previous findings showing a role for let-418 during development, we propose that LET-418/Mi2 could be part of a system that drives development and reproduction with concomitant life-reducing effects later in life. © 2013 the Anatomical Society and John Wiley & Sons Ltd.
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Methicillin-resistant Staphylococcus aureus harboring mecC in livestock in Spain.
Ariza-Miguel, Jaime; Hernández, Marta; Fernández-Natal, Isabel; Rodríguez-Lázaro, David
2014-11-01
We report for the first time mecC-positive methicillin-resistant Staphylococcus aureus (mecC-MRSA) in livestock in Spain. One isolate (sequence type 130) was found in milk samples among 601 S. aureus isolates obtained from 229 dairy sheep farms. This finding highlights the potential for zoonotic transmission of mecC-positive MRSA and the need for surveillance programs to monitor its presence and clonal evolution. Copyright © 2014, American Society for Microbiology. All Rights Reserved.
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H2O2_COD_EPA: Measurements of hydrogen peroxide and COD concentrations for water samples from the MEC reactors.MEC_acclimation: raw data for current and voltage of the anode in the MEC reactor.This dataset is associated with the following publication:Sim, J., J. An, E. Elbeshbishy, R. Hodon, and H. Lee. Characterization and optimization of cathodic conditions for H2O2 synthesis in microbial electrochemical cells. Bioresource Technology. Elsevier Online, New York, NY, USA, 195: 31-36, (2015).
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Buiza, Cristina; Navarro, Ana; Díaz-Orueta, Unai; González, Mari Feli; Alaba, Javier; Arriola, Enrique; Hernández, Carmen; Zulaica, Amaia; Yanguas, José Javier
2011-01-01
The cognitive assessment of patients with advanced dementia needs proper screening instruments that allow obtain information about the cognitive state and resources that these individuals still have. The present work conducts a Spanish validation study of the Severe Mini Mental State Examination (SMMSE). Forty-seven patients with advanced dementia (Mini-Cognitive Examination [MEC]<11) were evaluated with the Reisberg's Global Deterioration Scale, MEC, SMMSE and Severe Cognitive Impairment Profile scales. All test items were discriminative. The test showed high internal (α=0.88), test-retest (0.64 to 1.00, P<.01) and between observers reliabilities (0.69-1.00, p<0.01), both for scores total and for each item separately. Construct validity was tested through correlations between the instrument and MEC scores (r=0.59, P<0.01). Further information on the construct validity was obtained by dividing the sample into groups that scored above or below 5 points in the MEC and recalculating their correlations with SMMSE. The correlation between the scores in the SMMSE and MEC was significant in the MEC 0-5 group (r=0.55, P<.05), but not in the MEC>5 group. Additionally, differences in scores were found in the SMMSE, but not in the MEC, between the three GDS groups (5, 6 and 7) (H=11.1, P<.05). The SMMSE is an instrument for the assessment of advanced cognitive impairment which prevents the floor effect through an extension of lower measurement range relative to that of the MEC. From our results, this rapid screening tool and easy to administer, can be considered valid and reliable. Copyright © 2010 SEGG. Published by Elsevier Espana. All rights reserved.
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Kang, Yoo Kyung; Kwon, Kyu; Ryu, Jea Sung; Lee, Ha Neul; Park, Chankyu; Chung, Hyun Jung
2017-04-19
The overuse of antibiotics plays a major role in the emergence and spread of multidrug-resistant bacteria. A molecularly targeted, specific treatment method for bacterial pathogens can prevent this problem by reducing the selective pressure during microbial growth. Herein, we introduce a nonviral treatment strategy delivering genome editing material for targeting antibacterial resistance. We apply the CRISPR-Cas9 system, which has been recognized as an innovative tool for highly specific and efficient genome engineering in different organisms, as the delivery cargo. We utilize polymer-derivatized Cas9, by direct covalent modification of the protein with cationic polymer, for subsequent complexation with single-guide RNA targeting antibiotic resistance. We show that nanosized CRISPR complexes (= Cr-Nanocomplex) were successfully formed, while maintaining the functional activity of Cas9 endonuclease to induce double-strand DNA cleavage. We also demonstrate that the Cr-Nanocomplex designed to target mecA-the major gene involved in methicillin resistance-can be efficiently delivered into Methicillin-resistant Staphylococcus aureus (MRSA), and allow the editing of the bacterial genome with much higher efficiency compared to using native Cas9 complexes or conventional lipid-based formulations. The present study shows for the first time that a covalently modified CRISPR system allows nonviral, therapeutic genome editing, and can be potentially applied as a target specific antimicrobial.
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Lee, Beom; Park, Jun-Gyu; Shin, Won-Beom; Tian, Dong-Jie; Jun, Hang-Bae
2017-06-01
Microbial electrolysis cells (MECs) are being studied to improve the efficiency of anaerobic digesters and biogas production. In the present study, we investigated the effects of electrochemical reactions in AD-MEC (anaerobic digester combined with MECs) on changes in the microbial communities of bulk sludge through 454-pyrosequencing analysis, as well as the effect of these changes on anaerobic digestion. Methanobacterium beijingense and Methanobacterium petrolearium were the dominant archaeal species in AD, while Methanosarcina thermophila and Methanobacterium formicicum were dominant in AD-MEC at steady-state. There were no substantial differences in dominant bacterial species. Clostridia class was more abundant than Bacteroidia class in both reactors. Compared to AD, AD-MEC showed a 40% increase in overall bacterial population, increasing the removal of organic matters and the conversion of volatile fatty acids (VFAs). Thus, the MEC reaction more effectively converts organic matters to VFAs and activates microbial communities favorable for methane production. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Microbial electrolysis cells for waste biorefinery: A state of the art review.
Lu, Lu; Ren, Zhiyong Jason
2016-09-01
Microbial electrolysis cells (MECs) is an emerging technology for energy and resource recovery during waste treatment. MECs can theoretically convert any biodegradable waste into H2, biofuels, and other value added products, but the system efficacy can vary significantly when using different substrates or are operated in different conditions. To understand the application niches of MECs in integrative waste biorefineries, this review provides a critical analysis of MEC system performance reported to date in terms of H2 production rate, H2 yield, and energy efficiency under a variety of substrates, applied voltages and other crucial factors. It further discusses the mutual benefits between MECs and dark fermentation and argues such integration can be a viable approach for efficient H2 production from renewable biomass. Other marketable products and system integrations that can be applied to MECs are also summarized, and the challenges and prospects of the technology are highlighted. Copyright © 2016 Elsevier Ltd. All rights reserved.
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The faulty Master Events Controller is removed from STS-99 Endeavour
NASA Technical Reports Server (NTRS)
2000-01-01
Technicians remove a faulty Enhanced Main Events Controller (E- MEC) from Shuttle Endeavour at Launch Pad 39A. The E-MEC became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. Both E-MECs are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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The faulty Master Events Controller is carried away from STS-99 Endeavour
NASA Technical Reports Server (NTRS)
2000-01-01
Workers carry away the faulty Enhanced Main Events Controller (E- MEC) from Shuttle Endeavour at Launch Pad 39A. The E-MEC became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. Both E-MECs are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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Real-time monitoring of a microbial electrolysis cell using an electrical equivalent circuit model.
Hussain, S A; Perrier, M; Tartakovsky, B
2018-04-01
Efforts in developing microbial electrolysis cells (MECs) resulted in several novel approaches for wastewater treatment and bioelectrosynthesis. Practical implementation of these approaches necessitates the development of an adequate system for real-time (on-line) monitoring and diagnostics of MEC performance. This study describes a simple MEC equivalent electrical circuit (EEC) model and a parameter estimation procedure, which enable such real-time monitoring. The proposed approach involves MEC voltage and current measurements during its operation with periodic power supply connection/disconnection (on/off operation) followed by parameter estimation using either numerical or analytical solution of the model. The proposed monitoring approach is demonstrated using a membraneless MEC with flow-through porous electrodes. Laboratory tests showed that changes in the influent carbon source concentration and composition significantly affect MEC total internal resistance and capacitance estimated by the model. Fast response of these EEC model parameters to changes in operating conditions enables the development of a model-based approach for real-time monitoring and fault detection.
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Cytosine Methylation Effects on the Repair of O6-Methylguanines within CG Dinucleotides*
Guza, Rebecca; Ma, Linan; Fang, Qingming; Pegg, Anthony E.; Tretyakova, Natalia
2009-01-01
O6-Alkyldeoxyguanine adducts induced by tobacco-specific nitrosamines are repaired by O6-alkylguanine DNA alkyltransferase (AGT), which transfers the O6-alkyl group from the damaged base to a cysteine residue within the protein. In the present study, a mass spectrometry-based approach was used to analyze the effects of cytosine methylation on the kinetics of AGT repair of O6-methyldeoxyguanosine (O6-Me-dG) adducts placed within frequently mutated 5′-CG-3′ dinucleotides of the p53 tumor suppressor gene. O6-Me-dG-containing DNA duplexes were incubated with human recombinant AGT protein, followed by rapid quenching, acid hydrolysis, and isotope dilution high pressure liquid chromatography-electrospray ionization tandem mass spectrometry analysis of unrepaired O6-methylguanine. Second-order rate constants were calculated in the absence or presence of the C-5 methyl group at neighboring cytosine residues. We found that the kinetics of AGT-mediated repair of O6-Me-dG were affected by neighboring 5-methylcytosine (MeC) in a sequence-dependent manner. AGT repair of O6-Me-dG adducts placed within 5′-CG-3′ dinucleotides of p53 codons 245 and 248 was hindered when MeC was present in both DNA strands. In contrast, cytosine methylation within p53 codon 158 slightly increased the rate of O6-Me-dG repair by AGT. The effects of MeC located immediately 5′ and in the base paired position to O6-Me-dG were not additive as revealed by experiments with hypomethylated sequences. Furthermore, differences in dealkylation rates did not correlate with AGT protein affinity for cytosine-methylated and unmethylated DNA duplexes or with the rates of AGT-mediated nucleotide flipping, suggesting that MeC influences other kinetic steps involved in repair, e.g. the rate of alkyl transfer from DNA to AGT. PMID:19531487
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Cytosine methylation effects on the repair of O6-methylguanines within CG dinucleotides.
Guza, Rebecca; Ma, Linan; Fang, Qingming; Pegg, Anthony E; Tretyakova, Natalia
2009-08-21
O(6)-alkyldeoxyguanine adducts induced by tobacco-specific nitrosamines are repaired by O(6)-alkylguanine DNA alkyltransferase (AGT), which transfers the O(6)-alkyl group from the damaged base to a cysteine residue within the protein. In the present study, a mass spectrometry-based approach was used to analyze the effects of cytosine methylation on the kinetics of AGT repair of O(6)-methyldeoxyguanosine (O(6)-Me-dG) adducts placed within frequently mutated 5'-CG-3' dinucleotides of the p53 tumor suppressor gene. O(6)-Me-dG-containing DNA duplexes were incubated with human recombinant AGT protein, followed by rapid quenching, acid hydrolysis, and isotope dilution high pressure liquid chromatography-electrospray ionization tandem mass spectrometry analysis of unrepaired O(6)-methylguanine. Second-order rate constants were calculated in the absence or presence of the C-5 methyl group at neighboring cytosine residues. We found that the kinetics of AGT-mediated repair of O(6)-Me-dG were affected by neighboring 5-methylcytosine ((Me)C) in a sequence-dependent manner. AGT repair of O(6)-Me-dG adducts placed within 5'-CG-3' dinucleotides of p53 codons 245 and 248 was hindered when (Me)C was present in both DNA strands. In contrast, cytosine methylation within p53 codon 158 slightly increased the rate of O(6)-Me-dG repair by AGT. The effects of (Me)C located immediately 5' and in the base paired position to O(6)-Me-dG were not additive as revealed by experiments with hypomethylated sequences. Furthermore, differences in dealkylation rates did not correlate with AGT protein affinity for cytosine-methylated and unmethylated DNA duplexes or with the rates of AGT-mediated nucleotide flipping, suggesting that (Me)C influences other kinetic steps involved in repair, e.g. the rate of alkyl transfer from DNA to AGT.
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A new Master Events Controller is prepared for installation in STS-99 Endeavour
NASA Technical Reports Server (NTRS)
2000-01-01
Workers in a Quality trailer in the Launch Pad 39B Area unwrap a new Enhanced Main Events Controller (E-MEC) to be installed in Shuttle Endeavour. The original E-MEC in Endeavour became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. The E-MECs are located in the orbiter's aft compartment and both are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. Before workers can begin E-MEC replacement efforts at the launch pad, cryogenic reactants must be offloaded from the orbiter and Space Shuttle ordnance disconnected. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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A new Master Events Controller is prepared for installation in STS-99 Endeavour
NASA Technical Reports Server (NTRS)
2000-01-01
A new Enhanced Main Events Controller (E-MEC) for Shuttle Endeavour sits on a table in a Quality trailer in the Launch Pad 39B area. The original E-MEC in Endeavour became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. The E-MECs are located in the orbiter's aft compartment and both are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. Before workers can begin E-MEC replacement efforts at the launch pad, cryogenic reactants must be offloaded from the orbiter and Space Shuttle ordnance disconnected. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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NASA Astrophysics Data System (ADS)
DuVal, C.; Carton, G.; Trembanis, A. C.; Edwards, M.; Miller, J. K.
2017-12-01
Munitions and explosives of concern (MEC) are present in U.S. waters as a result of past and ongoing live-fire testing and training, combat operations, and sea disposal. To identify MEC that may pose a risk to human safety during development of offshore wind facilities on the Atlantic Outer Continental Shelf (OCS), the Bureau of Ocean Energy Management (BOEM) is preparing to develop guidance on risk analysis and selection processes for methods and technologies to identify MEC in Wind Energy Areas (WEA). This study developed a process for selecting appropriate technologies and methodologies for MEC detection using a synthesis of historical research, physical site characterization, remote sensing technology review, and in-field trials. Personnel were tasked with seeding a portion of the Delaware WEA with munitions surrogates, while a second group of researchers not privy to the surrogate locations tested and optimized the selected methodology to find and identify the placed targets. This in-field trial, conducted in July 2016, emphasized the use of multiple sensors for MEC detection, and led to further guidance for future MEC detection efforts on the Atlantic OCS. An April 2017 follow on study determined the fate of the munitions surrogates after the Atlantic storm season had passed. Using regional hydrodynamic models and incorporating the recommendations from the 2016 field trial, the follow on study examined the fate of the MEC and compared the findings to existing research on munitions mobility, as well as models developed as part of the Office of Naval Research Mine-Burial Program. Focus was given to characterizing the influence of sediment type on surrogate munitions behavior and the influence of mophodynamics and object burial on MEC detection. Supporting Mine-Burial models, ripple bedforms were observed to impede surrogate scour and burial in coarse sediments, while surrogate burial was both predicted and observed in finer sediments. Further, incorporation of recommendations from the previous trial in the 2017 study led to fourfold improvement of MEC detection rates over the 2016 approach. The use of modeling to characterize local morphodynamics, MEC burial or mobility, and the impact of seasonal or episodic storm events are discussed in light of technology selection and timing for future MEC detection surveys.
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Soeorg, Hiie; Metsvaht, Tuuli; Eelmäe, Imbi; Merila, Mirjam; Treumuth, Sirli; Huik, Kristi; Jürna-Ellam, Marika; Ilmoja, Mari-Liis; Lutsar, Irja
2017-11-01
BackgroundWe aimed to determine the genetic relatedness between Staphylococcus epidermidis colonizing breast milk (BM) and BM-fed neonates during the first month of life.MethodsS. epidermidis was isolated from the stool and skin swabs of 20 healthy term and 49 preterm neonates hospitalized in the neonatal intensive care unit and from the BM of mothers once a week and typed by multilocus variable-number tandem-repeat analysis. Virulence-related genes were determined by PCR.ResultsThe gut (95%) and skin (100%) of term neonates were colonized with strains genetically similar to those in BM and carrying mecA and IS256 at low rate (both <6.7%). In preterm neonates, colonization with strains genetically similar to those in BM was low on the skin (34.7%) and in the gut in the first week of life (14.3%), but the prevalence of mecA (>90.6%) and IS256 (>61.7%) was high. By the fourth week, in the gut of preterm neonates the prevalence of mecA (73.8%) and IS256 (18.4%) decreased, but colonization with strains genetically similar to those in BM increased (83.7%).ConclusionDuring early life, the skin and gut of preterm neonates is colonized with S. epidermidis that is distinct from strains found in BM, but gradually the gut is enriched with strains genetically similar to those in BM, as in term neonates.
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Methicillin-resistant staphylococcus aureus isolates in a hospital of shanghai.
Wang, Xiaoguang; Ouyang, Lin; Luo, Lingfei; Liu, Jiqian; Song, Chiping; Li, Cuizhen; Yan, Hongjing; Wang, Ping
2017-01-24
Methicillin-resistant Staphylococcus aureus (MRSA) strains are now common both in the health care setting and in the community. Active surveillance is critical for MRSA control and prevention. Specimens of patients (200 patients with 1119 specimens) as well as medical staff and hospital setting (1000 specimens) were randomly sampled in a level 2 hospital in Shanghai from September 2011 to August 2012. Isolation, cultivation and identification of S. aureus were performed. Totally, 67 S. aureus strains were isolated. 32 S. aureus strains were isolated from patient samples; 13 (13/32, 40.6%) of the 32 S. aureus isolates were MRSA; sputum sample and patients in the department of general internal medicine were the most frequent specimen and patient group for S. aureus strains isolation. Remaining 35 S. aureus strains were isolated from the medical staff and hospital setting; 20 (20/35, 57.1%) of the 35 S. aureus isolates were MRSA; specimens sampled from doctors and nurses' hands and nose and hospital facilities were the most frequent samples to isolate S. aureus. Resistant and virulent genes detection showed that, all 33 MRSA strains were mecA positive which accounts for 49.3% of the 67 S. aureus strains; 38 isolates were Panton-Valentine leukocidin (PVL) gene positive which accounts for 56.7% of the 67 S. aureus strains; and 17 (17/67, 25.4%) isolates are mecA and PVL genes dual positive. Multidrug-resistant strains of MRSA and PVL positive S. aureus are common in patients, medical staff and hospital setting, the potential health threat is worthy of our attention.
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Analysis of the ERK1,2 transcriptome in mammary epithelial cells
Grill, Constance; Gheyas, Ferdous; Dayananth, Priya; Jin, Weihong; Ding, Wei; Qiu, Ping; Wang, Luquan; Doll, Ronald J.; English, Jessie M.
2004-01-01
MAPK (mitogen-activated protein kinase) pathways constitute major regulators of cellular transcriptional programmes. We analysed the ERK1,2 (extracellular-signal-regulated kinase 1,2) transcriptome in a non-transformed MEC (mammary epithelial cell) line, MCF-12A, utilizing rAd MEK1EE, a recombinant adenovirus encoding constitutively active MEK1 (MAPK/ERK kinase 1). rAd MEK1EE infection induced morphological changes and DNA synthesis which were inhibited by the MEK1,2 inhibitor PD184352. Hierarchical clustering of data derived from seven time points over 24 h identified 430 and 305 co-ordinately up-regulated and down-regulated genes respectively. c-Myc binding sites were identified in the promoters of most of these up-regulated genes. A total of 46 candidate effectors of the Raf/MEK/ERK1,2 pathway in MECs were identified by comparing our dataset with previously reported Raf-1-regulated genes. These analyses led to the identification of a suite of growth factors co-ordinately induced by MEK1EE, including multiple ErbB ligands, vascular endothelial growth factor and PHRP (parathyroid hormone-related protein). PHRP is the primary mediator of humoral hypercalcaemia of malignancy, and has been implicated in metastasis to bone. We demonstrate that PHRP is secreted by MEK1EE-expressing cells. This secretion is inhibited by PD184352, but not by ErbB inhibitors. Our results suggest that, in addition to anti-proliferative properties, MEK1,2 inhibitors may be anti-angiogenic and possess therapeutic utility in the treatment of PHRP-positive tumours. PMID:15109307
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Chajęcka-Wierzchowska, Wioleta; Zadernowska, Anna; Nalepa, Beata; Sierpińska, Magda; Łaniewska-Trokenheim, Łucja
2015-04-01
The aim of this work was to study the pheno- and genotypical antimicrobial resistance profile of coagulase negative staphylococci (CoNS) isolated from 146 ready-to-eat food of animal origin (cheeses, cured meats, sausages, smoked fishes). 58 strains were isolated, they were classified as Staphylococcus xylosus (n = 29), Staphylococcus epidermidis (n = 16); Staphylococcus lentus (n = 7); Staphylococcus saprophyticus (n = 4); Staphylococcus hyicus (n = 1) and Staphylococcus simulans (n = 1) by phenotypic and genotypic methods. Isolates were tested for resistance to erythromycin, clindamycin, gentamicin, cefoxitin, norfloxacin, ciprofloxacin, tetracycline, tigecycline, rifampicin, nitrofurantoin, linezolid, trimetoprim, sulphamethoxazole/trimethoprim, chloramphenicol, quinupristin/dalfopristin by the disk diffusion method. PCR was used for the detection of antibiotic resistance genes encoding: methicillin resistance--mecA; macrolide resistance--erm(A), erm(B), erm(C), mrs(A/B); efflux proteins tet(K) and tet(L) and ribosomal protection proteins tet(M). For all the tet(M)-positive isolates the presence of conjugative transposons of the Tn916-Tn1545 family was determined. Most of the isolates were resistant to cefoxitin (41.3%) followed by clindamycin (36.2%), tigecycline (24.1%), rifampicin (17.2%) and erythromycin (13.8%). 32.2% staphylococcal isolates were multidrug resistant (MDR). All methicillin resistant staphylococci harboured mecA gene. Isolates, phenotypic resistant to tetracycline, harboured at least one tetracycline resistance determinant on which tet(M) was most frequent. All of the isolates positive for tet(M) genes were positive for the Tn916-Tn1545 -like integrase family gene. In the erythromycin-resistant isolates, the macrolide resistance genes erm(C) or msr(A/B) were present. Although coagulase-negative staphylococci are not classical food poisoning bacteria, its presence in food could be of public health significance due to the possible spread of antibiotic resistance. Copyright © 2014 Elsevier Ltd. All rights reserved.
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López, Yolanda; Samudio, Margarita; Fariña, Norma; Castillo, Verónica; Abente, Sonia; Nentwich, Martin M; González-Britez, Nilsa; Laspina, Florentina; Carron, Agustín; Cibils, Diógenes; de Kaspar, Herminia Miño
2017-08-01
In this prospective study, multiplex polymerase chain reaction (PCR) was used to identify genes encoding virulence factors (ica, atlE and mecA) in Coagulase-negative Staphylococcus (CNS) isolates from the ocular microbiota of patients undergoing cataract surgery and to investigate possible changes in the CNS profile due to antibiotic prophylaxis. Between 09/2011 and 08/2013, patients undergoing cataract surgery were recruited at the Department of Ophthalmology, National University of Asuncion, Paraguay. In the eye to be operated on, patients received moxifloxacin 0.5 % eye drops four times at the day before surgery and a last drop 1 hour before surgery (T1). The other eye remained as control (T0). Conjunctival swabs were taken from both eyes 1 hour after the last drop. The presence of genes encoding biofilm formation (ica and atlE) and methicillin resistance (mecA) was detected by a multiplex PCR. Of the 162 patients (162 study eyes, 162 fellow eye as control group), 87 (53.7 %) eyes were positive for CNS at T0 yielding 96 CNS isolates; 70 eyes (43.2 %) were positive at T1 yielding 77 CNS isolates. For this study, 43 CNS isolates (44.8 %) from T0 and 45 (64.3 %) from T1 were used. Of the total isolates, 81.8 % (72/88) had at least one virulence factor gene (37/43 from T0 and 35/45 from T1) (p = 0.314). Simultaneous detection of ica and atlE genes was higher in T0 (58.0 %) than T1 (46.7 %), but the difference was not significant (p = 0.28). A high frequency of genes encoding virulence factors was observed in the coagulase-negative Staphylococcus isolates. The use of moxifloxacin did not significantly modify the CNS virulence factor profiles.
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Carlesse, Fabianne; Cappellano, Paola; Quiles, Milene Gonçalves; Menezes, Liana Carballo; Petrilli, Antonio Sérgio; Pignatari, Antonio Carlos
2016-09-01
Bloodstream infections (BSIs) are the major cause of mortality in cancer patients. Molecular techniques are used for rapid diagnosis of BSI, allowing early therapy and improving survival. We aimed to establish whether real-time quantitative polymerase chain reaction (qPCR) could improve early diagnosis and therapy in paediatric cancer patients, and describe the predominant pathogens of BSI and their antimicrobial susceptibility. Blood samples were processed by the BACTEC system and microbial identification and susceptibility tests were performed by the Phoenix system. All samples were screened by multiplex 16 s rDNA qPCR. Seventeen species were evaluated using sex-specific TaqMan probes and resistance genes blaSHV, blaTEM, blaCTX, blaKPC, blaIMP, blaSPM, blaVIM, vanA, vanB and mecA were screened by SYBR Green reactions. Therapeutic efficacy was evaluated at the time of positive blood culture and at final phenotypic identification and antimicrobial susceptibility results. We analyzed 69 episodes of BSI from 64 patients. Gram-positive bacteria were identified in 61 % of the samples, Gram-negative bacteria in 32 % and fungi in 7 %. There was 78.2 % of agreement between the phenotypic and molecular methods in final species identification. The mecA gene was detected in 81.4 % of Staphylococcus spp., and 91.6 % were concordant with the phenotypic method. Detection of vanA gene was 100 % concordant. The concordance for Gram-negative susceptibilities was 71.4 % for Enterobacteriaceae and 50 % for Pseudomonas aeruginosa. Therapy was more frequently inadequate in patients who died, and the molecular test was concordant with the phenotypic susceptibility test in 50 %. qPCR has potential indication for early identification of pathogens and antimicrobial resistance genes from BSI in paediatric cancer patients and may improve antimicrobial therapy.
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Hau, Samantha J.; Frana, Timothy; Sun, Jisun; Davies, Peter R.
2017-01-01
ABSTRACT Zinc resistance in livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) sequence type 398 (ST398) is primarily mediated by the czrC gene colocated with the mecA gene, encoding methicillin resistance, within the type V staphylococcal cassette chromosome mec (SCCmec) element. Because czrC and mecA are located within the same mobile genetic element, it has been suggested that the use of zinc in feed as an antidiarrheal agent has the potential to contribute to the emergence and spread of methicillin-resistant S. aureus (MRSA) in swine, through increased selection pressure to maintain the SCCmec element in isolates obtained from pigs. In this study, we report the prevalence of the czrC gene and phenotypic zinc resistance in U.S. swine-associated LA-MRSA ST5 isolates, MRSA ST5 isolates from humans with no swine contact, and U.S. swine-associated LA-MRSA ST398 isolates. We demonstrated that the prevalence of zinc resistance in U.S. swine-associated LA-MRSA ST5 isolates was significantly lower than the prevalence of zinc resistance in MRSA ST5 isolates from humans with no swine contact and swine-associated LA-MRSA ST398 isolates, as well as prevalences from previous reports describing zinc resistance in other LA-MRSA ST398 isolates. Collectively, our data suggest that selection pressure associated with zinc supplementation in feed is unlikely to have played a significant role in the emergence of LA-MRSA ST5 in the U.S. swine population. Additionally, our data indicate that zinc resistance is associated with the multilocus sequence type lineage, suggesting a potential link between the genetic lineage and the carriage of resistance determinants. IMPORTANCE Our data suggest that coselection thought to be associated with the use of zinc in feed as an antimicrobial agent is not playing a role in the emergence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) ST5 in the U.S. swine population. Additionally, our data indicate that zinc resistance is more associated with the multilocus sequence type lineage, suggesting a potential link between the genetic lineage and the carriage of resistance markers. This information is important for public health professionals, veterinarians, producers, and consumers. PMID:28526788
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Gajewska, Malgorzata; McNally, Sara
2017-01-01
Three-dimensional (3D) cell cultures on reconstituted basement membrane (rBM) enable the study of complex interactions between extracellular matrix (ECM) components and epithelial cells, which are crucial for the establishment of cell polarity and functional development of epithelia. 3D cultures of mammary epithelial cells (MECs) on Matrigel (a laminin-rich ECM derived from the Engelbreth-Holm-Swarm (EHS) murine tumor) promote interactions of MECs with the matrix via integrins, leading to formation of spherical monolayers of polarized cells surrounding a hollow lumen (acini). Acini closely resemble mammary alveoli found in the mammary gland. Thus, it is possible to study ECM-cell interactions and signalling pathways that regulate formation and maintenance of tissue-specific shape and functional differentiation of MECs in 3D under in vitro conditions. Here we present experimental protocols used to investigate the role of mitogen-activated protein kinase phosphatases (MKPs) during development of the alveoli-like structures by primary mouse mammary epithelial cells (PMMEC) cultured on Matrigel. We present detailed protocols for PMMEC isolation, and establishment of 3D cultures using an "on top" method, use of specific kinase and phosphatases inhibitors (PD98059 and pervanadate, respectively) administered at different stages of acinus development, and give examples of analyses carried out post-culture (Western blot, immunofluorescence staining, and confocal imaging).
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Rossi, Esther Diana; Bizzarro, Tommaso; Monterossi, Giorgia; Inzani, Frediano; Fanfani, Francesco; Scambia, Giovanni; Zannoni, Gian Franco
2017-04-01
Mixed endometrial carcinomas (MECs) refer to tumors characterized by 2 or more distinct histotypes mostly that comprised endometrioid (EC) and serous/clear cell carcinomas (SC/CC). The specific quantification of these distinct components represents a challenging and critical point for both prognosis and management. Herein, we analyze a large series of MEC and compare them with EC and SC/CC. We evaluated a series of 69 MECs between January 2002 and December 2015. We compared the MEC series with 186 ECs (including 117 endometrioid G3), 31 SCs, and 38 CCs. The prognostic implication of the percentage of each component was analyzed. Among the 69 MECs, those patients older than 45 years represent the significant population, with 52.2% of them with stage III-IV disease. A similar result was found among pure SC. The comparative analysis of some prognostic parameters (multifocality, vascular invasion, and lymph node metastasis) underlined that MECs with a type II component larger than 5% represent a more aggressive entity. However, relapse, disease-free survival, mortality, and overall survival are statistically significant (P<.05) in EC-SC (SC<5%or >5%) and in EC-CC (CC<5%or >5%), whereas they are not significant (P>.05) in SC-CC (SC/CC<%or >5%). MECs, including also cases with less than 5% of SC/CC, show features as aggressive as those of pure SC/CC. In this perspective, MEC should be followed by personalized and tailored managements. The presence of different components suggests different pathogenic and metastatic processes when compared with pure carcinomas. Copyright © 2016. Published by Elsevier Inc.
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Wang, Aijie; Sun, Dan; Cao, Guangli; Wang, Haoyu; Ren, Nanqi; Wu, Wei-Min; Logan, Bruce E
2011-03-01
Hydrogen gas production from cellulose was investigated using an integrated hydrogen production process consisting of a dark fermentation reactor and microbial fuel cells (MFCs) as power sources for a microbial electrolysis cell (MEC). Two MFCs (each 25 mL) connected in series to an MEC (72 mL) produced a maximum of 0.43 V using fermentation effluent as a feed, achieving a hydrogen production rate from the MEC of 0.48 m(3) H(2)/m(3)/d (based on the MEC volume), and a yield of 33.2 mmol H(2)/g COD removed in the MEC. The overall hydrogen production for the integrated system (fermentation, MFC and MEC) was increased by 41% compared with fermentation alone to 14.3 mmol H(2)/g cellulose, with a total hydrogen production rate of 0.24 m(3) H(2)/m(3)/d and an overall energy recovery efficiency of 23% (based on cellulose removed) without the need for any external electrical energy input. Copyright © 2010 Elsevier Ltd. All rights reserved.
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STS-99 workers carry new Master Events Controller to Endeavour
NASA Technical Reports Server (NTRS)
2000-01-01
Workers carry the replacement Enhanced Main Events Controller (E- MEC) to Shuttle Endeavour at Launch Pad 39A for installation in the aft compartment of the payload bay. The original E-MEC became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. Both E-MECs are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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STS-99 workers move new Master Events Controller into aft compartment
NASA Technical Reports Server (NTRS)
2000-01-01
At Launch Pad 39A, workers move the replacement Enhanced Main Events Controller (E-MEC) into Shuttle Endeavour's aft compartment in the payload bay. The original E-MEC became suspect during the Jan. 31 launch countdown and mission STS-99 was delayed when NASA managers decided to replace it. Each Shuttle carries two enhanced master events controllers (E-MECs), which provide relays for onboard flight computers to send signals to arm and fire pyrotechnics that separate the solid rockets and external tank during assent. Both E-MECs are needed for the Shuttle to be cleared for flight. Currently Endeavour and Columbia are the only two orbiters with the E-MECs. Built by Rockwell's Satellite Space Electronics Division, Anaheim, Calif., each unit weighs 65 pounds and is approximately 20 inches long, 13 inches wide and 8 inches tall. Previously, three Shuttle flights have been scrubbed or delayed due to faulty MECs: STS-73, STS-49 and STS-41-D. The next scheduled date for launch of STS-99 is Feb. 11 at 12:30 p.m. EST.
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Ajayi, Folusho Francis; Kim, Kyoung-Yeol; Chae, Kyu-Jung; Choi, Mi-Jin; Chang, In Seop; Kim, In S
2010-03-01
Bio-hydrogen production in light-assisted microbial electrolysis cell (MEC) with a dye sensitized solar cell (DSSC) was optimized by connecting multiple MECs to a single dye (N719) sensitized solar cell (V(OC) approx. 0.7 V). Hydrogen production occurred simultaneously in all the connected MECs when the solar cell was irradiated with light. The amount of hydrogen produced in each MEC depends on the activity of the microbial catalyst on their anode. Substrate (acetate) to hydrogen conversion efficiencies ranging from 42% to 65% were obtained from the reactors during the experiment. A moderate light intensity of 430 W m(-2) was sufficient for hydrogen production in the coupled MEC-DSSC. A higher light intensity of 915 W m(-2), as well as an increase in substrate concentration, did not show any improvement in the current density due to limitation caused by the rate of microbial oxidation on the anode. A significant reduction in the surface area of the connected DSSC only showed a slight effect on current density in the coupled MEC-DSSC system when irradiated with light.
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Software Supports Distributed Operations via the Internet
NASA Technical Reports Server (NTRS)
Norris, Jeffrey; Backers, Paul; Steinke, Robert
2003-01-01
Multi-mission Encrypted Communication System (MECS) is a computer program that enables authorized, geographically dispersed users to gain secure access to a common set of data files via the Internet. MECS is compatible with legacy application programs and a variety of operating systems. The MECS architecture is centered around maintaining consistent replicas of data files cached on remote computers. MECS monitors these files and, whenever one is changed, the changed file is committed to a master database as soon as network connectivity makes it possible to do so. MECS provides subscriptions for remote users to automatically receive new data as they are generated. Remote users can be producers as well as consumers of data. Whereas a prior program that provides some of the same services treats disconnection of a user from the network of users as an error from which recovery must be effected, MECS treats disconnection as a nominal state of the network: This leads to a different design that is more efficient for serving many users, each of whom typically connects and disconnects frequently and wants only a small fraction of the data at any given time.
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Matias, C A R; Pereira, I A; Rodrigues, D P; Siciliano, S
2018-06-20
This work aimed to investigate the prevalence of Staphylococcus in wild birds seized in illegal trade and their antimicrobial resistance patterns. Cloacal samples were obtained from 109 wild birds apprehended in the street markets in Rio de Janeiro, Brazil. Staphylococcus spp. were pheno and genotypically identified and resistance profile were evaluated by CLSI guidelines and by Polymerase Chain Reaction of mecA and blaZ genes. Staphylococcus was detected in 45,9% (50/109) of the cloacal swab samples and thirty-nine (78,0%) isolates were resistant to one or more of the nine antimicrobials tested, and were also positive to mecA (12/39) or blaZ genes (14/39). High percentage of resistance was detected to ampicillin, oxacillin, cefoxitin, clindamycin and tetracycline, with absence of resistance to vancomycin. Wild birds captured and submitted to captive stress conditions of illegal trade market of Brazil may have an important role as reservoirs of Staphylococcus spp. and its antimicrobial resistance mechanisms. The significance of the present study is revealed by the zoonotic and pathogenic potential of staphylococci and that impact to public health and requires monitoring polices of wild birds health in tropical areas. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
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Kang, Sun-Young; Kim, Kang-Ju
2015-01-01
In this study, we used ethanol extract of A. princeps and investigated its antibacterial effects against MRSA. Ethanol extract of A. princeps significantly inhibited MRSA growth and organic acid production during glucose metabolism at concentrations greater than 1 mg/mL (P < 0.05). MRSA biofilm formation was observed using scanning electron microscopy (SEM) and safranin staining. A. princeps extract was found to inhibit MRSA biofilm formation at concentrations higher than 2 mg/mL significantly (P < 0.05). Bactericidal effects of the A. princeps were observed using confocal laser microscopy, which showed that A. princeps was bactericidal in a dose-dependent manner. Using real-time PCR, expression of mecA, an antibiotic-resistance gene of MRSA, was observed, along with that of sea, agrA, and sarA. A. princeps significantly inhibited mecA, sea, agrA, and sarA, mRNA expression at the concentrations greater than 1 mg/mL (P < 0.05). The phytochemical analysis of A. princeps showed a relatively high content of organic acids and glycosides. The results of this study suggest that the ethanol extract of A. princeps may inhibit proliferation, acid production, biofilm formation, and virulence gene expressions of MRSA, which may be related to organic acids and glycosides, the major components in the extract. PMID:26247012
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Choi, Na-Young; Kang, Sun-Young; Kim, Kang-Ju
2015-01-01
In this study, we used ethanol extract of A. princeps and investigated its antibacterial effects against MRSA. Ethanol extract of A. princeps significantly inhibited MRSA growth and organic acid production during glucose metabolism at concentrations greater than 1 mg/mL (P < 0.05). MRSA biofilm formation was observed using scanning electron microscopy (SEM) and safranin staining. A. princeps extract was found to inhibit MRSA biofilm formation at concentrations higher than 2 mg/mL significantly (P < 0.05). Bactericidal effects of the A. princeps were observed using confocal laser microscopy, which showed that A. princeps was bactericidal in a dose-dependent manner. Using real-time PCR, expression of mecA, an antibiotic-resistance gene of MRSA, was observed, along with that of sea, agrA, and sarA. A. princeps significantly inhibited mecA, sea, agrA, and sarA, mRNA expression at the concentrations greater than 1 mg/mL (P < 0.05). The phytochemical analysis of A. princeps showed a relatively high content of organic acids and glycosides. The results of this study suggest that the ethanol extract of A. princeps may inhibit proliferation, acid production, biofilm formation, and virulence gene expressions of MRSA, which may be related to organic acids and glycosides, the major components in the extract.
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Rapid amplification/detection of nucleic acid targets utilizing a HDA/thin film biosensor.
Jenison, Robert; Jaeckel, Heidi; Klonoski, Joshua; Latorra, David; Wiens, Jacinta
2014-08-07
Thin film biosensors exploit a flat, optically coated silicon-based surface whereupon formation of nucleic acid hybrids are enzymatically transduced in a molecular thin film that can be detected by the unaided human eye under white light. While the limit of sensitivity for detection of nucleic acid targets is at sub-attomole levels (60 000 copies) many clinical specimens containing bacterial pathogens have much lower levels of analyte present. Herein, we describe a platform, termed HDA/thin film biosensor, which performs helicase-dependant nucleic acid amplification on a thin film biosensor surface to improve the limit of sensitivity to 10 copies of the mecA gene present in methicillin-resistant strains of Staphylococcus. As double-stranded DNA is unwound by helicase it was either bound by solution-phase DNA primers to be copied by DNA polymerase or hybridized to surface immobilized probe on the thin film biosensor surface to be detected. Herein, we show that amplification reactions on the thin film biosensor are equivalent to in standard thin wall tubes, with detection at the limit of sensitivity of the assay occurring after 30 minutes of incubation time. Further we validate the approach by detecting the presence of the mecA gene in methicillin-resistant Staphylococcus aureus (MRSA) from positive blood culture aliquots with high specificity (signal/noise ratio of 105).
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Wagner, Vivian P.; Martins, Marco A.T.; Martins, Manoela D.; Warner, Kristy A.; Webber, Liana P.; Squarize, Cristiane H.; Nör, Jacques E.; Castilho, Rogerio M.
2016-01-01
Patients with mucoepidermoid carcinoma (MEC) experience low survival rates and high morbidity following treatment, yet the intrinsic resistance of MEC cells to ionizing radiation (IR) and the mechanisms underlying acquired resistance remain unexplored. Herein, we demonstrated that low doses of IR intrinsically activated NFκB in resistant MEC cell lines. Moreover, resistance was significantly enhanced in IR-sensitive cell lines when NFκB pathway was stimulated. Pharmacological inhibition of the IKK-β/IκBα/NFκB axis, using a single dose of FDA-approved Emetine, led to a striking sensitization of MEC cells to IR and a reduction in cancer stem cells. We achieved a major step towards better understanding the basic mechanisms involved in IR-adaptive resistance in MEC cell lines and how to efficiently overcome this critical problem. PMID:27682876
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Multi-stakeholder perspectives of locally commissioned enhanced optometric services
Baker, H; Harper, R A; Edgar, D F; Lawrenson, J G
2016-01-01
Objectives To explore views of all stakeholders (patients, optometrists, general practitioners (GPs), commissioners and ophthalmologists) regarding the operation of community-based enhanced optometric services. Design Qualitative study using mixed methods (patient satisfaction surveys, semi-structured telephone interviews and optometrist focus groups). Setting A minor eye conditions scheme (MECS) and glaucoma referral refinement scheme (GRRS) provided by accredited community optometrists. Participants 189 patients, 25 community optometrists, 4 glaucoma specialist hospital optometrists (GRRS), 5 ophthalmologists, 6 GPs (MECS), 4 commissioners. Results Overall, 99% (GRRS) and 100% (MECS) patients were satisfied with their optometrists’ examination. The vast majority rated the following as ‘very good’; examination duration, optometrists’ listening skills, explanations of tests and management, patient involvement in decision-making, treating the patient with care and concern. 99% of MECS patients would recommend the service. Manchester optometrists were enthusiastic about GRRS, feeling fortunate to practise in a ‘pro-optometry’ area. No major negatives were reported, although both schemes were limited to patients resident within certain postcode areas, and some inappropriate GP referrals occurred (MECS). Communication with hospitals was praised in GRRS but was variable, depending on hospital (MECS). Training for both schemes was valuable and appropriate but should be ongoing. MECS GPs were very supportive, reporting the scheme would reduce secondary care referral numbers, although some MECS patients were referred back to GPs for medication. Ophthalmologists (MECS and GRRS) expressed very positive views and widely acknowledged that these new care pathways would reduce unnecessary referrals and shorten patient waiting times. Commissioners felt both schemes met or exceeded expectations in terms of quality of care, allowing patients to be seen quicker and more efficiently. Conclusions Locally commissioned schemes can be a positive experience for all involved. With appropriate training, clear referral pathways and good communication, community optometrists can offer high-quality services that are highly acceptable to patients, health professionals and commissioners. PMID:27798000
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Kadier, Abudukeremu; Kalil, Mohd Sahaid; Chandrasekhar, Kuppam; Mohanakrishna, Gunda; Saratale, Ganesh Dattatraya; Saratale, Rijuta Ganesh; Kumar, Gopalakrishnan; Pugazhendhi, Arivalagan; Sivagurunathan, Periyasamy
2018-02-01
Microbial electrolysis cells (MECs) are perceived as a potential and promising innovative biotechnological tool that can convert carbon-rich waste biomass or wastewater into hydrogen (H 2 ) or other value-added chemicals. Undesired methane (CH 4 ) producing H 2 sinks, including methanogens, is a serious challenge faced by MECs to achieve high-rate H 2 production. Methanogens can consume H 2 to produce CH 4 in MECs, which has led to a drop of H 2 production efficiency, H 2 production rate (HPR) and also a low percentage of H 2 in the produced biogas. Organized inference related to the interactions of microbes and potential processes has assisted in understanding approaches and concepts for inhibiting the growth of methanogens and profitable scale up design. Thus, here in we review the current developments and also the improvements constituted for the reduction of microbial H 2 losses to methanogens. Firstly, the greatest challenge in achieving practical applications of MECs; undesirable microorganisms (methanogens) growth and various studied techniques for eliminating and reducing methanogens activities in MECs were discussed. Additionally, this extensive review also considers prospects for stimulating future research that could help to achieve more information and would provide the focus and path towards MECs as well as their possibilities for simultaneously generating H 2 and waste remediation. Copyright © 2017 Elsevier B.V. All rights reserved.
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Kim, Nahui; Dempsey, Catherine M.; Kuan, Chih-Jen; Zoval, Jim V.; O'Rourke, Eyleen; Ruvkun, Gary; Madou, Marc J.; Sze, Ji Y.
2007-01-01
The gravity response is an array of behavioral and physiological plasticity elicited by changes in ambient mechanical force and is an evolutionarily ancient adaptive mechanism. We show in Caenorhabditis elegans that the force of hypergravity is translated into biological signaling via a genetic pathway involving three factors: the degenerin/epithelial Na+ channel (DEG/ENaC) class of mechanosensory channels of touch receptor neurons, the neurotransmitter serotonin, and the FoxO transcription factor DAF-16 known to regulate development, energy metabolism, stress responses, and aging. After worms were exposed to hypergravity for 3 hr, their muscular and neuronal functions were preserved, but they exhibited DAF-16∷GFP nuclear accumulation in cells throughout the body and accumulated excess fat. Mutations in MEC-4/MEC-10 DEG/ENaC or its partners MEC-6, MEC-7, and MEC-9 blocked DAF-16∷GFP nuclear accumulation induced by hypergravity but did not affect DAF-16 response to other stresses. We show that exogenous serotonin and the antidepressant fluoxetine can attenuate DAF-16∷GFP nuclear accumulation in WT animals exposed to hypergravity. These results reveal a novel physiological role of the mechanosensory channel, showing that the perception of mechanical stress controls FoxO signaling pathways and that inactivation of DEG/ENaC may decouple mechanical loading and physiological responses. PMID:17720915
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NASA Astrophysics Data System (ADS)
Huang, Liping; Yao, Binglin; Wu, Dan; Quan, Xie
2014-08-01
Complete cobalt recovery from lithium cobalt oxide requires to firstly leach cobalt from particles LiCoO2 and then recover cobalt from aqueous Co(II). A self-driven microbial fuel cell (MFC)-microbial electrolysis cell (MEC) system can completely carry out these two processes, in which Co(II) is firstly released from particles LiCoO2 on the cathodes of MFCs and then reduced on the cathodes of MECs which are powered by the cobalt leaching MFCs. A cobalt leaching rate of 46 ± 2 mg L-1 h-1 with yield of 1.5 ± 0.1 g Co g-1 COD (MFCs) and a Co(II) reduction rate of 7 ± 0 mg L-1 h-1 with yield of 0.8 ± 0.0 g Co g-1 COD (MECs), as well as a overall system cobalt yield of 0.15 ± 0.01 g Co g-1 Co can be achieved in this self-driven MFC-MEC system. Coulombic efficiencies reach 41 ± 1% (anodic MFCs), 75 ± 0% (anodic MECs), 100 ± 2% (cathodic MFCs), and 29 ± 1% (cathodic MECs) whereas overall system efficiency averages 34 ± 1%. These results provide a new process of linking MFCs to MECs for complete recovery of cobalt and recycle of spent lithium ion batteries with no external energy consumption.
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Wei, Zhengkai; Zhou, Ershun; Guo, Changming; Fu, Yunhe; Yu, Yuqiang; Li, Yimeng; Yao, Minjun; Zhang, Naisheng; Yang, Zhengtao
2014-01-01
Bovine mastitis is one of the most costly and prevalent diseases in the dairy industry and is characterised by inflammatory and infectious processes. Staphylococcus aureus (S. aureus), a Gram-positive organism, is a frequent cause of subclinical, chronic mastitis. Thymol, a monocyclic monoterpene compound isolated from Thymus vulgaris, has been reported to have antibacterial properties. However, the effect of thymol on S. aureus internalization into bovine mammary epithelial cells (bMEC) has not been investigated. In this study, we evaluated the effect of thymol on S. aureus internalization into bMEC, the expression of tracheal antimicrobial peptide (TAP) and β-defensin (BNBD5), and the inhibition of NF-κB activation in bMEC infected with S. aureus. Our results showed that thymol (16-64 μg/ml) could reduce the internalization of S. aureus into bMEC and down-regulate the mRNA expression of TAP and BNBD5 in bMEC infected with S. aureus. In addition, thymol was found to inhibit S. aureus-induced nitric oxide (NO) production in bMEC and suppress S. aureus-induced NF-κB activation in a dose-dependent manner. In conclusion, these results indicated that thymol inhibits S. aureus internalization into bMEC by inhibiting NF-κB activation. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Gonzalez-Sulser, Alfredo; Parthier, Daniel; Candela, Antonio; McClure, Christina; Pastoll, Hugh; Garden, Derek; Sürmeli, Gülşen
2014-01-01
The medial septum (MS) is required for theta rhythmic oscillations and grid cell firing in the medial entorhinal cortex (MEC). While GABAergic, glutamatergic, and cholinergic neurons project from the MS to the MEC, their synaptic targets are unknown. To investigate whether MS neurons innervate specific layers and cell types in the MEC, we expressed channelrhodopsin-2 in mouse MS neurons and used patch-clamp recording in brain slices to determine the response to light activation of identified cells in the MEC. Following activation of MS axons, we observed fast monosynaptic GABAergic IPSPs in the majority (>60%) of fast-spiking (FS) and low-threshold-spiking (LTS) interneurons in all layers of the MEC, but in only 1.5% of nonstellate principal cells (NSPCs) and in no stellate cells. We also observed fast glutamatergic responses to MS activation in a minority (<5%) of NSPCs, FS, and LTS interneurons. During stimulation of MS inputs at theta frequency (10 Hz), the amplitude of GABAergic IPSPs was maintained, and spike output from LTS and FS interneurons was entrained at low (25–60 Hz) and high (60–180 Hz) gamma frequencies, respectively. By demonstrating cell type-specific targeting of the GABAergic projection from the MS to the MEC, our results support the idea that the MS controls theta frequency activity in the MEC through coordination of inhibitory circuits. PMID:25505326
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77 FR 13043 - Airworthiness Directives; The Boeing Company Airplanes
Federal Register 2010, 2011, 2012, 2013, 2014
2012-03-05
... to the top surface of the MEC drip shield. That NPRM was prompted by a report of a multi-power system loss in flight of 1, 2, and 3 alternating current electrical power systems located in the MEC. This... current electrical power systems located in the main equipment center (MEC). We are issuing this AD to...
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Booth, Clair A.; Ridler, Thomas; Murray, Tracey K.; Ward, Mark A.; de Groot, Emily; Goodfellow, Marc; Phillips, Keith G.; Randall, Andrew D.
2016-01-01
The entorhinal cortex (EC) is one of the first areas to be disrupted in neurodegenerative diseases such as Alzheimer's disease and frontotemporal dementia. The responsiveness of individual neurons to electrical and environmental stimuli varies along the dorsal–ventral axis of the medial EC (mEC) in a manner that suggests this topographical organization plays a key role in neural encoding of geometric space. We examined the cellular properties of layer II mEC stellate neurons (mEC-SCs) in rTg4510 mice, a rodent model of neurodegeneration. Dorsoventral gradients in certain intrinsic membrane properties, such as membrane capacitance and afterhyperpolarizations, were flattened in rTg4510 mEC-SCs, while other cellular gradients [e.g., input resistance (Ri), action potential properties] remained intact. Specifically, the intrinsic properties of rTg4510 mEC-SCs in dorsal aspects of the mEC were preferentially affected, such that action potential firing patterns in dorsal mEC-SCs were altered, while those in ventral mEC-SCs were unaffected. We also found that neuronal oscillations in the gamma frequency band (30–80 Hz) were preferentially disrupted in the dorsal mEC of rTg4510 slices, while those in ventral regions were comparatively preserved. These alterations corresponded to a flattened dorsoventral gradient in theta-gamma cross-frequency coupling of local field potentials recorded from the mEC of freely moving rTg4510 mice. These differences were not paralleled by changes to the dorsoventral gradient in parvalbumin staining or neurodegeneration. We propose that the selective disruption to dorsal mECs, and the resultant flattening of certain dorsoventral gradients, may contribute to disturbances in spatial information processing observed in this model of dementia. SIGNIFICANCE STATEMENT The medial entorhinal cortex (mEC) plays a key role in spatial memory and is one of the first areas to express the pathological features of dementia. Neurons of the mEC are anatomically arranged to express functional dorsoventral gradients in a variety of neuronal properties, including grid cell firing field spacing, which is thought to encode geometric scale. We have investigated the effects of tau pathology on functional dorsoventral gradients in the mEC. Using electrophysiological approaches, we have shown that, in a transgenic mouse model of dementia, the functional properties of the dorsal mEC are preferentially disrupted, resulting in a flattening of some dorsoventral gradients. Our data suggest that neural signals arising in the mEC will have a reduced spatial content in dementia. PMID:26758825
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Alva-Murillo, Nayeli; Medina-Estrada, Ivan; Báez-Magaña, Marisol; Ochoa-Zarzosa, Alejandra; López-Meza, Joel E
2015-12-01
Staphylococcus aureus is an etiological agent of human and animal diseases, and it is able to internalize into non-professional phagocytic cells (i.e. bovine mammary epithelial cells, bMECs), which is an event that is related to chronic and recurrent infections. bMECs contribute to host innate immune responses (IIR) through TLR pathogen recognition, whereby TLR2 is the most relevant for S. aureus. In a previous report, we showed that sodium butyrate (NaB, 0.5mM), which is a short chain fatty acid (SCFA), reduced S. aureus internalization into bMECs by modulating their IIR. However, the molecular mechanism of this process has not been described, which was the aim of this study. The results showed that the TLR2 membrane abundance (MA) and mRNA expression were induced by 0.5mM NaB ∼1.6-fold and ∼1.7-fold, respectively. Additionally, 0.5mM NaB induced p38 phosphorylation, but not JNK1/2 or ERK1/2 phosphorylation in bMECs, which reached the baseline when the bMECs were S. aureus-challenged. Additionally, bMECs that were treated with 0.5mM NaB (24h) showed activation of 8 transcriptional factors (AP-1, E2F-1, FAST-1, MEF-1, EGR, PPAR, ER and CBF), which were partially reverted when the bMECs were S. aureus-challenged. Additionally, 0.5mM NaB (24h) up-regulated mRNA expression of the antimicrobial peptides, TAP (∼4.8-fold), BNBD5 (∼3.2-fold) and BNBD10 (∼2.6-fold). Notably, NaB-treated and S. aureus-challenged bMECs increased the mRNA expression of all of the antimicrobial peptides that were evaluated, and this was evident for LAP and BNBD5. In the NaB-treated bMECs, we did not detect significant expression changes for IL-1β and IL-6 and only TNF-α, IL-10 and IL-8 were induced. Interestingly, the NaB-treated and S. aureus-challenged bMECs maintained the anti-inflammatory response that was induced by this SCFA. In conclusion, our results suggest that 0.5mM NaB activates bMECs via TLR2/p38, which leads to improved antimicrobial defense before/after pathogen invasion, and NaB may exert anti-inflammatory effects during infection. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Invasion in breast lesions: the role of the epithelial-stroma barrier.
Rakha, Emad A; Miligy, Islam M; Gorringe, Kylie L; Toss, Michael S; Green, Andrew R; Fox, Stephen B; Schmitt, Fernando C; Tan, Puay-Hoon; Tse, Gary M; Badve, Sunil; Decker, Thomas; Vincent-Salomon, Anne; Dabbs, David J; Foschini, Maria P; Moreno, Filipa; Wentao, Yang; Geyer, Felipe C; Reis-Filho, Jorge S; Pinder, Sarah E; Lakhani, Sunil R; Ellis, Ian O
2018-06-01
Despite the significant biological, behavioural and management differences between ductal carcinoma in situ (DCIS) and invasive carcinoma of the breast, they share many morphological and molecular similarities. Differentiation of these two different lesions in breast pathological diagnosis is based typically on the presence of an intact barrier between the malignant epithelial cells and stroma; namely, the myoepithelial cell (MEC) layer and surrounding basement membrane (BM). Despite being robust diagnostic criteria, the identification of MECs and BM to differentiate in-situ from invasive carcinoma is not always straightforward. The MEC layer around DCIS may be interrupted and/or show an altered immunoprofile. MECs may be absent in some benign locally infiltrative lesions such as microglandular adenosis and infiltrating epitheliosis, and occasionally in non-infiltrative conditions such as apocrine lesions, and in these contexts this does not denote malignancy or invasive disease with metastatic potential. MECs may also be absent around some malignant lesions such as some forms of papillary carcinoma, yet these behave in an indolent fashion akin to some DCIS. In Paget's disease, malignant mammary epithelial cells extend anteriorly from the ducts to infiltrate the epidermis of the nipple but do not typically infiltrate through the BM into the dermis. Conversely, BM-like material can be seen around invasive carcinoma cells and around metastatic tumour cell deposits. Here, we review the role of MECs and BM in breast pathology and highlight potential clinical implications. We advise caution in interpretation of MEC features in breast pathology and mindfulness of the substantive evidence base in the literature associated with behaviour and clinical outcome of lesions classified as benign on conventional morphological examination before changing classification to an invasive lesion on the sole basis of MEC characteristics. © 2017 John Wiley & Sons Ltd.
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Honjo, Kie; Hiraki, Tsubasa; Higashi, Michiyo; Noguchi, Hirotsugu; Nomoto, Mitsuharu; Yoshimura, Takuya; Batra, Surinder K; Yonezawa, Suguru; Semba, Ichiro; Nakamura, Norifumi; Tanimoto, Akihide; Yamada, Sohsuke
2018-02-01
In mucoepidermoid carcinoma (MEC), the most common salivary gland carcinoma, there is a lack of novel prognostic markers, but post-operative early recurrence strongly affects the clinical course and a poor outcome. It is critical to predict which MEC patients are prone to develop recurrence/metastases. Mucins play pivotal roles in influencing cancer biology, thus affecting cell differentiation, adhesion, carcinoma invasion, aggressiveness and/or metastatic potential. Our aim is to elucidate the significance of expression profiles for mucins, particularly MUC4 and MUC6, and their correlations with various clinicopathological features and recurrence in salivary gland MECs. We performed immunohistochemical analyses on patients with surgically resected primary MEC using antibodies against mucin core proteins MUC4/8G7 and MUC6/CLH5 in 73 paraffin-embedded samples. Recurrence was noted in 15 of 73 (20.5%) patients. MUC4 or MUC6 expression was considered to be negative when <30% or 0% of the MEC cells showed positive staining, respectively. MUC4- and/or MUC6-negative expression respectively and variably showed a significant relationship to pathological tumor high-grade, the presence of lymphovascular invasion, lymph node metastasis and/or tumor-related death. In addition, MUC4 showed significantly negative co-expression with MUC6. Kaplan-Meier analyses revealed that not only single MUC4/6-negative expression but also the combination of both predicted significantly shorter disease-free and disease-specific survivals in MECs, especially within the first two years postoperatively. Therefore, each mucin plays a pivotal role in the pathogenesis of MEC progression. The detection of MUC4 and/or MUC6 might be a powerful parameter in the clinical management of MECs in the early postsurgical phase.
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Nolte, Kurt B; Hanzlick, Randy L; Payne, Daniel C; Kroger, Andrew T; Oliver, William R; Baker, Andrew M; McGowan, Dennis E; DeJong, Joyce L; Bell, Micahel R; Guarner, Jeannette; Shieh, Wun-Ju; Zaki, Sherif R
2004-06-11
Medical examiners and coroners (ME/Cs) are essential public health partners for terrorism preparedness and response. These medicolegal investigators support both public health and public safety functions and investigate deaths that are sudden, suspicious, violent, unattended, and unexplained. Medicolegal autopsies are essential for making organism-specific diagnoses in deaths caused by biologic terrorism. This report has been created to 1) help public health officials understand the role of ME/Cs in biologic terrorism surveillance and response efforts and 2) provide ME/Cs with the detailed information required to build capacity for biologic terrorism preparedness in a public health context. This report provides background information regarding biologic terrorism, possible biologic agents, and the consequent clinicopathologic diseases, autopsy procedures, and diagnostic tests as well as a description of biosafety risks and standards for autopsy precautions. ME/Cs' vital role in terrorism surveillance requires consistent standards for collecting, analyzing, and disseminating data. Familiarity with the operational, jurisdictional, and evidentiary concerns involving biologic terrorism-related death investigation is critical to both ME/Cs and public health authorities. Managing terrorism-associated fatalities can be expensive and can overwhelm the existing capacity of ME/Cs. This report describes federal resources for funding and reimbursement for ME/C preparedness and response activities and the limited support capacity of the federal Disaster Mortuary Operational Response Team. Standards for communication are critical in responding to any emergency situation. This report, which is a joint collaboration between CDC and the National Association of Medical Examiners (NAME), describes the relationship between ME/Cs and public health departments, emergency management agencies, emergency operations centers, and the Incident Command System.
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Representation of Non-Spatial and Spatial Information in the Lateral Entorhinal Cortex
Deshmukh, Sachin S.; Knierim, James J.
2011-01-01
Some theories of memory propose that the hippocampus integrates the individual items and events of experience within a contextual or spatial framework. The hippocampus receives cortical input from two major pathways: the medial entorhinal cortex (MEC) and the lateral entorhinal cortex (LEC). During exploration in an open field, the firing fields of MEC grid cells form a periodically repeating, triangular array. In contrast, LEC neurons show little spatial selectivity, and it has been proposed that the LEC may provide non-spatial input to the hippocampus. Here, we recorded MEC and LEC neurons while rats explored an open field that contained discrete objects. LEC cells fired selectively at locations relative to the objects, whereas MEC cells were weakly influenced by the objects. These results provide the first direct demonstration of a double dissociation between LEC and MEC inputs to the hippocampus under conditions of exploration typically used to study hippocampal place cells. PMID:22065409
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P63 Positive Mucoepidermoid Tumor of the Lacrimal Sac with Associated Papilloma.
Iordanous, Yiannis; Belrose, Jillian C; Cadieux, Dani C; Chakrabarti, Subrata; Farmer, James P; Allen, Larry H
2015-01-01
We report a case of a 44-year-old man who presented with a left medial canthal mass and epiphora. Imaging was suggestive of a mass continuous with the nasolacrimal sac. Subsequent surgical exploration revealed a mass adherent to bone with invasion of the lacrimal system. Histological examination revealed a squamous/transitional cell papilloma overlying a low-grade mucoepidermoid carcinoma (MEC). Complete surgical resection was completed and pathology confirmed the diagnosis. This is the first case in which a MEC has been reported concurrently with an overlying papilloma, providing support for the hypothesis that MECs arise from papillomas in the lacrimal sac. Additionally, the tissue stained positive for p63, which is congruent with MEC immunoreactivity in the salivary gland. The description of these unique histopathological findings may assist in definitive diagnosis and improve our understanding of the pathophysiology underlying lacrimal sac MEC tumors.
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Reduced Spiking in Entorhinal Cortex during the Delay Period of a Cued Spatial Response Task
ERIC Educational Resources Information Center
Gupta, Kishan; Keller, Lauren A.; Hasselmo, Michael E.
2012-01-01
Intrinsic persistent spiking mechanisms in medial entorhinal cortex (mEC) neurons may play a role in active maintenance of working memory. However, electrophysiological studies of rat mEC units have primarily focused on spatial modulation. We sought evidence of differential spike rates in the mEC in rats trained on a T-maze, cued spatial delayed…
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Materials experiment carrier concepts definition study. Volume 1: Executive summary, part 2
NASA Technical Reports Server (NTRS)
1981-01-01
The materials experiment carrier (MEC) is an optimized carrier for near term and advanced materials processing in space (MPS) research and commercial payloads. When coupled with the space platform (SP), the MEC can provide the extended duration, high power and low acceleration environment the MPS payload typically requires. The lowest cost, technically reasonable first step MEC that meets the MPS program missions objectives with minimum programmatic risks is defined. The effectiveness of the initial MEC/space platform idea for accommodating high priority, multidiscipline, R&D and commercial MPS payloads, and conducting MPS payload oprations at affordable funding and acceptable productivity levels is demonstrated.
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Analysis about modeling MEC7000 excitation system of nuclear power unit
NASA Astrophysics Data System (ADS)
Liu, Guangshi; Sun, Zhiyuan; Dou, Qian; Liu, Mosi; Zhang, Yihui; Wang, Xiaoming
2018-02-01
Aiming at the importance of accurate modeling excitation system in stability calculation of nuclear power plant inland and lack of research in modeling MEC7000 excitation system,this paper summarize a general method to modeling and simulate MEC7000 excitation system. Among this method also solve the key issues of computing method of IO interface parameter and the conversion process of excitation system measured model to BPA simulation model. At last complete the simulation modeling of MEC7000 excitation system first time in domestic. By used No-load small disturbance check, demonstrates that the proposed model and algorithm is corrective and efficient.
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Schauer, B; Krametter-Frötscher, R; Knauer, F; Ehricht, R; Monecke, S; Feßler, A T; Schwarz, S; Grunert, T; Spergser, J; Loncaric, I
2018-02-01
The aim of this study was to determine the prevalence, the antimicrobial resistance patterns and the genetic diversity of methicillin-resistant Staphylococcus aureus (MRSA) from Austrian ruminants and New World camelids that were treated at the University of Veterinary Medicine, Vienna. Between April 2014 and January 2017, 723 nasal swabs originating from ruminants and New World camelids were examined. MRSA isolates were characterized by mecA/mecA1/mecC PCRs and by DNA microarray analysis. They were genotyped by spa typing, dru typing, MLST and MLVA. Glycopolymer fingerprinting by FTIR spectroscopy was also performed. Antimicrobial susceptibility testing was conducted by agar disk diffusion. Twelve MRSA isolates were mecA-positive, whereas three were mecC-positive. The MRSA isolates carried five different SCCmec elements, and belonged to three sequence types (ST45, ST130, ST398). The MRSA isolates displayed seven different resistance phenotypes. The present study describes for the first time mecC-carrying MRSA isolates originating from domesticated animals in Austria. More systematic studies are needed to unravel the role of ruminants and New World camelids as reservoirs for MRSA as a potential risk for zooanthropogenic transmission. Copyright © 2018 Elsevier B.V. All rights reserved.
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Jurisdiction of the Medical Ethics Committees.
Voljč, Božidar
2017-10-01
Ethical principles of assessing medical research are to the greatest extent defined by the Nuremberg Code, the Declarations of Geneva and Helsinki, and the Oviedo Convention. Pursuant to their directives various national Medical Ethics Committees (MECs) were established which assess the ethics of research according to the risk and benefit ratio of the persons involved. Following the example of other countries, medical ethics committees eventually appeared also in hospitals and some medical and educational institutions around Slovenia. Due to an increased number of ethical challenges, it is of great importance to define the jurisdiction of the Slovenian MECs in order to ensure their coordinated operation. Exclusive jurisdiction of the national MEC includes multicentre and multi-national research, drug research (phases 1-3), high-risk research and research related to doctoral theses. The jurisdiction of the sectoral MECs includes testing the conditions for research, monitoring the execution and overviewing the final reports. A more significant jurisdiction of the sectoral MEC is preserving an ethical environment in their institutions. A network of Slovenian MECs is to be organised in the form of a jurisdiction pyramid where each member has its own obligations and responsibilities and plays an important role in relation to the entire structure.
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Presence of Methicillin Resistant Staphylococcus aureus (MRSA) in sewage treatment plant.
Boopathy, Raj
2017-09-01
The presence of antibiotic resistant bacteria and antibiotic resistance genes in rural sewage treatment plants are not well reported in the literature. The aim of the present study was to study the frequency occurrence of Methicillin Resistant Staphylococcus aureus (MRSA) in a rural sewage treatment plant. This study was conducted using raw sewage as well as treated sewage from a small town sewage treatment plant in rural southeast Louisiana of USA. Results showed the presence of MRSA consistently in both raw and treated sewage. The presence of mecA gene responsible for methicillin resistance was confirmed in the raw and treated sewage water samples. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Mechanisms of antibiotic resistance in Staphylococcus aureus.
Pantosti, Annalisa; Sanchini, Andrea; Monaco, Monica
2007-06-01
Staphylococcus aureus can exemplify better than any other human pathogen the adaptive evolution of bacteria in the antibiotic era, as it has demonstrated a unique ability to quickly respond to each new antibiotic with the development of a resistance mechanism, starting with penicillin and methicillin, until the most recent, linezolid and daptomycin. Resistance mechanisms include enzymatic inactivation of the antibiotic (penicillinase and aminoglycoside-modification enzymes), alteration of the target with decreased affinity for the antibiotic (notable examples being penicillin-binding protein 2a of methicillin-resistant S. aureus and D-Ala-D-Lac of peptidoglycan precursors of vancomycin-resistant strains), trapping of the antibiotic (for vancomycin and possibly daptomycin) and efflux pumps (fluoroquinolones and tetracycline). Complex genetic arrays (staphylococcal chromosomal cassette mec elements or the vanA operon) have been acquired by S. aureus through horizontal gene transfer, while resistance to other antibiotics, including some of the most recent ones (e.g., fluoroquinolones, linezolid and daptomycin) have developed through spontaneous mutations and positive selection. Detection of the resistance mechanisms and their genetic basis is an important support to antibiotic susceptibility surveillance in S. aureus.
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Can 5-methylcytosine analogues with extended alkyl side chains guide DNA methylation?
Kotandeniya, D; Seiler, C L; Fernandez, J; Pujari, S S; Curwick, L; Murphy, K; Wickramaratne, S; Yan, S; Murphy, D; Sham, Yuk Y; Tretyakova, N Y
2018-01-25
5-Methylcytosine ( Me C) is an endogenous modification of DNA that plays a crucial role in DNA-protein interactions, chromatin structure, epigenetic regulation, and DNA repair. Me C is produced via enzymatic methylation of the C-5 position of cytosine by DNA-methyltransferases (DNMT) which use S-adenosylmethionine (SAM) as a cofactor. Hemimethylated CG dinucleotides generated as a result of DNA replication are specifically recognized and methylated by maintenance DNA methyltransferase 1 (DNMT1). The accuracy of DNMT1-mediated methylation is essential for preserving tissue-specific DNA methylation and thus gene expression patterns. In the present study, we synthesized DNA duplexes containing MeC analogues with modified C-5 side chains and examined their ability to guide cytosine methylation by the human DNMT1 protein. We found that the ability of 5-alkylcytosines to direct cytosine methylation decreased with increased alkyl chain length and rigidity (methyl > ethyl > propyl ∼ vinyl). Molecular modeling studies indicated that this loss of activity may be caused by the distorted geometry of the DNA-protein complex in the presence of unnatural alkylcytosines.
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Mammary analogue secretory carcinoma (MASC) of salivary gland in four Mexican patients.
Serrano-Arévalo, Mónica L; Mosqueda-Taylor, Adalberto; Domínguez-Malagón, Hugo; Michal, Michal
2015-01-01
The Clinco-pathological, immunohistochemical and molecular findings of four cases of Mammary Analogue Secretory Carcinoma (MASC) of salivary glands found in Mexico are described. The cases were extracted from 253 salivary gland tumors from a single institution in Mexico City. The 85 Candidates for initial selection were: low grade mucoepidermoid carcinoma (MEC) (N=70 ), Acinic cell cancinoma (AciCC) (N=14), papillary cystadenocarcinoma (N=1), and adenocarcinoma NOS (N=0). Tumors with some histological features consistent with MASC (N= 17, 6.7%) were studied by immunohistochemistry for mammaglobin, STAT5, and S-100 protein and four cases were positive (1.5%), thus the diagnosis of MASC was established, and these were submitted for molecular studies for ETV6-NTRK3. Fusion gene was demonstrated in three cases, two had been erroneously diagnosed as poorly granulated AciCC, and one as low grade MEC with microcystic pattern. Female gender predominated (3:1); one occurred in the parotid, two in minor salivary glands and one in the submaxillary gland; infiltrating borders, atypical mitosis and lymph node metastases were seen in the parotideal tumor. Two patients with major salivary gland tumors are alive and well at 10 and 20 months respectively, the two patients with minor salivary gland tumors are lost. It can be concluded that is important to think in MASC in poorly granulated AciCC and low grade MEC with microcystic pattern. Immunohistochemisty studies confirm the diagnosis, preferentially supported by molecular studies. MASC may follow aggressive behavior or transform into a high grade neoplasm.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Hasan, IIftekhar; Husain, Tausif; Uddin, Md Wasi
2015-08-24
This paper presents a nonlinear analytical model of a novel double-sided flux concentrating Transverse Flux Machine (TFM) based on the Magnetic Equivalent Circuit (MEC) model. The analytical model uses a series-parallel combination of flux tubes to predict the flux paths through different parts of the machine including air gaps, permanent magnets, stator, and rotor. The two-dimensional MEC model approximates the complex three-dimensional flux paths of the TFM and includes the effects of magnetic saturation. The model is capable of adapting to any geometry that makes it a good alternative for evaluating prospective designs of TFM compared to finite element solversmore » that are numerically intensive and require more computation time. A single-phase, 1-kW, 400-rpm machine is analytically modeled, and its resulting flux distribution, no-load EMF, and torque are verified with finite element analysis. The results are found to be in agreement, with less than 5% error, while reducing the computation time by 25 times.« less
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Analytical Modeling of a Novel Transverse Flux Machine for Direct Drive Wind Turbine Applications
DOE Office of Scientific and Technical Information (OSTI.GOV)
Hasan, IIftekhar; Husain, Tausif; Uddin, Md Wasi
2015-09-02
This paper presents a nonlinear analytical model of a novel double sided flux concentrating Transverse Flux Machine (TFM) based on the Magnetic Equivalent Circuit (MEC) model. The analytical model uses a series-parallel combination of flux tubes to predict the flux paths through different parts of the machine including air gaps, permanent magnets (PM), stator, and rotor. The two-dimensional MEC model approximates the complex three-dimensional flux paths of the TFM and includes the effects of magnetic saturation. The model is capable of adapting to any geometry which makes it a good alternative for evaluating prospective designs of TFM as compared tomore » finite element solvers which are numerically intensive and require more computation time. A single phase, 1 kW, 400 rpm machine is analytically modeled and its resulting flux distribution, no-load EMF and torque, verified with Finite Element Analysis (FEA). The results are found to be in agreement with less than 5% error, while reducing the computation time by 25 times.« less
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Specific roles for DEG/ENaC and TRP channels in touch and thermosensation in C. elegans nociceptors
Chatzigeorgiou, Marios; Yoo, Sungjae; Watson, Joseph D.; Lee, Wei-Hsiang; Spencer, W. Clay; Kindt, Katie S.; Hwang, Sun Wook; Miller, David M.; Treinin, Millet; Driscoll, Monica; Schafer, William R.
2010-01-01
Summary Polymodal nociceptors detect noxious stimuli including harsh touch, toxic chemicals, and extremes of heat and cold. The molecular mechanisms by which nociceptors are able to sense multiple qualitatively distinct stimuli are not well-understood. We show here that the C. elegans PVD neurons are mulitidendritic nociceptors that respond to harsh touch as well as cold temperatures. The harsh touch modality specifically requires the DEG/ENaC proteins MEC-10 and DEGT-1, which represent putative components of a harsh touch mechanotransduction complex. By contrast, responses to cold require the TRPA-1 channel and are MEC-10- and DEGT-1-independent. Heterologous expression of C. elegans TRPA-1 can confer cold responsiveness to other C. elegans neurons or to mammalian cells, indicating that TRPA-1 is itself a cold sensor. These results show that C. elegans nociceptors respond to thermal and mechanical stimuli using distinct sets of molecules, and identify DEG/ENaC channels as potential receptors for mechanical pain. PMID:20512132
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Santosaningsih, Dewi; Santoso, Sanarto; Setijowati, Nanik; Rasyid, Harun A; Budayanti, Nyoman S; Suata, Ketut; Widhyatmoko, Dicky B; Purwono, Priyo B; Kuntaman, Kuntaman; Damayanti, Damayanti; Prakoeswa, Cita R S; Laurens, Mitchell; van Nierop, Josephine W I; Nanninga, Geraldine L; Oudenes, Neline; de Regt, Michelle; Snijders, Susan V; Verbrugh, Henri A; Severin, Juliëtte A
2018-01-01
To define the role of Staphylococcus aureus in community settings among patients with skin and soft tissue infections (SSTI) in Indonesia. Staphylococcus aureus were cultured from anterior nares, throat and wounds of 567 ambulatory patients presenting with SSTI. The mecA gene and genes encoding Panton-Valentine leukocidin (PVL; lukF-PV and lukS-PV) and exfoliative toxin (ET; eta and etb) were determined by PCR. Clonal relatedness among methicillin-resistant S. aureus (MRSA) and PVL-positive S. aureus was analysed using multilocus variable-number tandem-repeat analysis (MLVA) typing, and multilocus sequence typing (MLST) for a subset of isolates. Staphylococcal cassette chromosome mec (SCCmec) was determined for all MRSA isolates. Moreover, determinants for S. aureus SSTI, and PVL/ET-positive vs PVL/ET-negative S. aureus were assessed. Staphylococcus aureus were isolated from SSTI wounds of 257 (45.3%) patients, eight (3.1%) of these were MRSA. Genes encoding PVL and ETs were detected in 21.8% and 17.5% of methicillin-susceptible S. aureus (MSSA), respectively. PVL-positive MRSA was not detected. Nasopharyngeal S. aureus carriage was an independent determinant for S. aureus SSTI (odds ratio [OR] 1.8). Primary skin infection (OR 5.4) and previous antibiotic therapy (OR 3.5) were associated with PVL-positive MSSA. Primary skin infection (OR 2.2) was the only factor associated with ET-positive MSSA. MLVA typing revealed two more prevalent MSSA clusters. One ST1-MRSA-SCCmec type IV isolate and a cluster of ST239-MRSA-SCCmec type III were found. Community-acquired SSTI in Indonesia was frequently caused by PVL-positive MSSA, and the hospital-associated ST239-MRSA may have spread from the hospital into the community. © 2017 John Wiley & Sons Ltd.
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Dekker, Denise; Wolters, Manuel; Mertens, Eva; Boahen, Kennedy Gyau; Krumkamp, Ralf; Eibach, Daniel; Schwarz, Norbert G; Adu-Sarkodie, Yaw; Rohde, Holger; Christner, Martin; Marks, Florian; Sarpong, Nimako; May, Jürgen
2016-11-29
Staphylococcus aureus is among the most common pathogens isolated from blood cultures in Ghana; yet the epidemiology of blood infections in rural settings is poorly described. This study aims to investigate antimicrobial susceptibility and clonal diversity of S. aureus causing bloodstream infections in two hospitals in the Ashanti Region, Ghana. Blood cultures were performed for all febrile patients (≥37.5 °C) on hospital admission. Antibiotic susceptibility testing for S. aureus isolates was carried out by the VITEK 2 system. Multiplex polymerase chain reaction (PCR) was used to detect S. aureus-specific nuc gene, Panton-Valentine leukocidin (PVL), and methicillin-resistant S. aureus (MRSA)-specific mecA and mecC genes. The population structure of S. aureus was assessed by spa typing. In total, 9,834 blood samples were cultured, out of which 0.6% (n = 56) were positive for S. aureus. Multidrug resistance (MDR) was detected in 35.7% (n = 20) of the S. aureus strains, of which one was a MRSA. The highest rate of antibiotic resistance was seen for commonly available antibiotics, including penicillin (n = 55; 98.2%), tetracycline (n = 32; 57.1%) and trimethoprim/sulfamethoxazole (n = 26; 46.4%). Of all S. aureus strains, 75.0% (n = 42) carried the PVL-encoding genes. We found 25 different spa types with t355 (n = 11; 19.6%), t314 (n = 8; 14.3%), t084 (n = 8; 14.3%) and t311 (n = 5; 8.9%) being predominant. The study exhibited an alarmingly large level of antibiotic resistance to locally available antibiotics. The frequency of genetically diverse and PVL-positive methicillin-sensitive S. aureus (MSSA) was high and could represent a reservoir for the emergence of virulent PVL-positive MRSA clones.
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Shahid, Muhammad; Wang, Jianfang; Gu, Xiaolong; Chen, Wei; Ali, Tariq; Gao, Jian; Han, Dandan; Yang, Rui; Fanning, Séamus; Han, Bo
2017-01-01
Prototheca zopfii infections are becoming global concerns in humans and animals. Bovine protothecal mastitis is characterized by deteriorating milk quality and quantity, thus imparting huge economic losses to dairy industry. Previous published studies mostly focused on the prevalence and characterization of P. zopfii from mastitis. However, the ultrastructural pathomorphological changes associated with apoptosis in bovine mammary epithelial cells (bMECs) are not studied yet. Therefore, in this study we aimed to evaluate the in vitro comparative apoptotic potential of P. zopfii genotype-I and -II on bMECs using flow cytometry, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The results showed fast growth rate and higher adhesion capability of genotype-II in bMECs as compared with genotype-I. The viability of bMECs infected with P. zopfii genotype-II was significantly decreased after 12 h (p < 0.05) and 24 h (p < 0.01) in comparison with control cells. Contrary, genotype-I couldn't show any significant effects on cell viability. Moreover, after infection of bMECs with genotype-II, the apoptosis increased significantly at 12 h (p < 0.05) and 24 h (p < 0.01) as compared with control group. Genotype-I couldn't display any significant effects on cell apoptosis. The host specificity of P. zopfii was also tested in mouse osteoblast cells, and the results suggest that genotype-I and -II could not cause any significant apoptosis in these cell lines. SEM interpreted the pathomorphological alterations in bMECs after infection. Adhesion of P. zopfii with cells and further disruption of cytomembrane validated the apoptosis caused by genotype-II under SEM. While genotype-1 couldn't cause any significant apoptosis in bMECs. Furthermore, genotype-II induced apoptotic manifested specific ultrastructure features, like cytoplasmic cavitation, swollen mitochondria, pyknosis, cytomembrane disruption, and appearance of apoptotic bodies under TEM. The findings of the current study revealed that genotype-II has the capability to invade and survive within the bMECs, thus imparting significant damages to the mammary cells which result in apoptosis. This study represents the first insights into the pathomorphological and ultrastructure features of apoptosis in bMECs induced by P. zopfii genotype-II. PMID:28752077
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Rajesh, Mathur; Wang, Gang; Jones, Roger; Tretyakova, Natalia
2005-02-15
The p53 tumor suppressor gene is a primary target in smoking-induced lung cancer. Interestingly, p53 mutations observed in lung tumors of smokers are concentrated at guanine bases within endogenously methylated (Me)CG dinucleotides, e.g., codons 157, 158, 245, 248, and 273 ((Me)C = 5-methylcytosine). One possible mechanism for the increased mutagenesis at these sites involves targeted binding of metabolically activated tobacco carcinogens to (Me)CG sequences. In the present work, a stable isotope labeling HPLC-ESI(+)-MS/MS approach was employed to analyze the formation of guanine lesions induced by the tobacco-specific lung carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) within DNA duplexes representing p53 mutational "hot spots" and surrounding sequences. Synthetic DNA duplexes containing p53 codons 153-159, 243-250, and 269-275 were prepared, where (Me)C was incorporated at all physiologically methylated CG sites. In each duplex, one of the guanine bases was replaced with [1,7,NH(2)-(15)N(3)-2-(13)C]-guanine, which served as an isotope "tag" to enable specific quantification of guanine lesions originating from that position. After incubation with NNK diazohydroxides, HPLC-ESI(+)-MS/MS analysis was used to determine the yields of NNK adducts at the isotopically labeled guanine and at unlabeled guanine bases elsewhere in the sequence. We found that N7-methyl-2'-deoxyguanosine and N7-[4-oxo-4-(3-pyridyl)but-1-yl]guanine lesions were overproduced at the 3'-guanine bases within polypurine runs, while the formation of O(6)-methyl-2'-deoxyguanosine and O(6)-[4-oxo-4-(3-pyridyl)but-1-yl]-2'-deoxyguanosine adducts was specifically preferred at the 3'-guanine base of 5'-GG and 5'-GGG sequences. In contrast, the presence of 5'-neighboring (Me)C inhibited O(6)-guanine adduct formation. These results indicate that the N7- and O(6)-guanine adducts of NNK are not overproduced at the endogenously methylated CG dinucleotides within the p53 tumor suppressor gene, suggesting that factors other than NNK adduct formation are responsible for mutagenesis at these sites.
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Guan, Jing; Wang, Wanying; Sun, Baolin
2017-01-01
CRISPR-Cas (clustered regularly interspaced short palindromic repeat [CRISPR]-CRISPR-associated protein [Cas]) systems can provide protection against invading genetic elements by using CRISPR RNAs (crRNAs) as a guide to locate and degrade the target DNA. CRISPR-Cas systems have been classified into two classes and five types according to the content of cas genes. Previous studies have indicated that CRISPR-Cas systems can avoid viral infection and block plasmid transfer. Here we show that chromosomal targeting by the Staphylococcus aureus type III-A CRISPR-Cas system can drive large-scale genome deletion and alteration within integrated staphylococcal cassette chromosome mec (SCC mec ). The targeting activity of the CRISPR-Cas system is associated with the complementarity between crRNAs and protospacers, and 10- to 13-nucleotide truncations of spacers partially block CRISPR attack and more than 13-nucleotide truncation can fully abolish targeting, suggesting that a minimal length is required to license cleavage. Avoiding base pairings in the upstream region of protospacers is also necessary for CRISPR targeting. Successive trinucleotide complementarity between the 5' tag of crRNAs and protospacers can disrupt targeting. Our findings reveal that type III-A CRISPR-Cas systems can modulate bacterial genome stability and may serve as a high-efficiency tool for deleting resistance or virulence genes in bacteria. IMPORTANCE Staphylococcus aureus is a pathogen that can cause a wide range of infections in humans. Studies have suggested that CRISPR-Cas systems can drive the loss of integrated mobile genetic elements (MGEs) by chromosomal targeting. Here we demonstrate that CRISPR-mediated cleavage contributes to the partial deletion of integrated SCC mec in methicillin-resistant S. aureus (MRSA), which provides a strategy for the treatment of MRSA infections. The spacer within artificial CRISPR arrays should contain more than 25 nucleotides for immunity, and consecutive trinucleotide pairings between a selected target and the 5' tag of crRNA can block targeting. These findings add to our understanding of the molecular mechanisms of the type III-A CRISPR-Cas system and provide a novel strategy for the exploitation of engineered CRISPR immunity against integrated MGEs in bacteria for clinical and industrial applications.
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Gray, Stephen; Allison, Rachal M; Garcia, Valerie; Goldman, Alastair S H; Neale, Matthew J
2013-07-31
During meiosis, formation and repair of programmed DNA double-strand breaks (DSBs) create genetic exchange between homologous chromosomes-a process that is critical for reductional meiotic chromosome segregation and the production of genetically diverse sexually reproducing populations. Meiotic DSB formation is a complex process, requiring numerous proteins, of which Spo11 is the evolutionarily conserved catalytic subunit. Precisely how Spo11 and its accessory proteins function or are regulated is unclear. Here, we use Saccharomyces cerevisiae to reveal that meiotic DSB formation is modulated by the Mec1(ATR) branch of the DNA damage signalling cascade, promoting DSB formation when Spo11-mediated catalysis is compromised. Activation of the positive feedback pathway correlates with the formation of single-stranded DNA (ssDNA) recombination intermediates and activation of the downstream kinase, Mek1. We show that the requirement for checkpoint activation can be rescued by prolonging meiotic prophase by deleting the NDT80 transcription factor, and that even transient prophase arrest caused by Ndt80 depletion is sufficient to restore meiotic spore viability in checkpoint mutants. Our observations are unexpected given recent reports that the complementary kinase pathway Tel1(ATM) acts to inhibit DSB formation. We propose that such antagonistic regulation of DSB formation by Mec1 and Tel1 creates a regulatory mechanism, where the absolute frequency of DSBs is maintained at a level optimal for genetic exchange and efficient chromosome segregation.
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Kubota, S; Takezawa, T; Mori, Y; Takakuwa, T
1992-09-01
We applied the multicellular spheroids which consist of cholangiocarcinoma cell line (MEC) and human dermal fibroblasts (HDF) to in vitro chemosensitivity test. Five-day multicellular spheroids were incubated with 1.5 micrograms/ml of mitomycin C (MMC) for 24 hrs. Then, cell kinetics of MEC and HDF in a spheroid was determined by flow cytometric analysis. Twenty four hrs after treatment with MMC, both MEC and HDF were accumulated on S phase. Seven-day after treatment, DNA histogram in MEC returned to normal, but that of HDF was disappeared. These results showed that the multicellular assay could be more like on in vivo like chemosensitivity test.
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RB loss contributes to aggressive tumor phenotypes in MYC-driven triple negative breast cancer
Knudsen, Erik S; McClendon, A Kathleen; Franco, Jorge; Ertel, Adam; Fortina, Paolo; Witkiewicz, Agnieszka K
2015-01-01
Triple negative breast cancer (TNBC) is characterized by multiple genetic events occurring in concert to drive pathogenic features of the disease. Here we interrogated the coordinate impact of p53, RB, and MYC in a genetic model of TNBC, in parallel with the analysis of clinical specimens. Primary mouse mammary epithelial cells (mMEC) with defined genetic features were used to delineate the combined action of RB and/or p53 in the genesis of TNBC. In this context, the deletion of either RB or p53 alone and in combination increased the proliferation of mMEC; however, the cells did not have the capacity to invade in matrigel. Gene expression profiling revealed that loss of each tumor suppressor has effects related to proliferation, but RB loss in particular leads to alterations in gene expression associated with the epithelial-to-mesenchymal transition. The overexpression of MYC in combination with p53 loss or combined RB/p53 loss drove rapid cell growth. While the effects of MYC overexpression had a dominant impact on gene expression, loss of RB further enhanced the deregulation of a gene expression signature associated with invasion. Specific RB loss lead to enhanced invasion in boyden chambers assays and gave rise to tumors with minimal epithelial characteristics relative to RB-proficient models. Therapeutic screening revealed that RB-deficient cells were particularly resistant to agents targeting PI3K and MEK pathway. Consistent with the aggressive behavior of the preclinical models of MYC overexpression and RB loss, human TNBC tumors that express high levels of MYC and are devoid of RB have a particularly poor outcome. Together these results underscore the potency of tumor suppressor pathways in specifying the biology of breast cancer. Further, they demonstrate that MYC overexpression in concert with RB can promote a particularly aggressive form of TNBC. PMID:25602521
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Yoo, Seung-Woo; Lee, Inah
2017-01-01
How visual scene memory is processed differentially by the upstream structures of the hippocampus is largely unknown. We sought to dissociate functionally the lateral and medial subdivisions of the entorhinal cortex (LEC and MEC, respectively) in visual scene-dependent tasks by temporarily inactivating the LEC and MEC in the same rat. When the rat made spatial choices in a T-maze using visual scenes displayed on LCD screens, the inactivation of the MEC but not the LEC produced severe deficits in performance. However, when the task required the animal to push a jar or to dig in the sand in the jar using the same scene stimuli, the LEC but not the MEC became important. Our findings suggest that the entorhinal cortex is critical for scene-dependent mnemonic behavior, and the response modality may interact with a sensory modality to determine the involvement of the LEC and MEC in scene-based memory tasks. DOI: http://dx.doi.org/10.7554/eLife.21543.001 PMID:28169828
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Concurrent hydrogen production and phosphorus recovery in dual chamber microbial electrolysis cell.
Almatouq, Abdullah; Babatunde, A O
2017-08-01
Concurrent hydrogen (H 2 ) production and phosphorus (P) recovery were investigated in dual chamber microbial electrolysis cells (MECs). The aim of the study was to explore and understand the influence of applied voltage and influent COD concentration on concurrent H 2 production and P recovery in MEC. P was efficiently precipitated at the cathode chamber and the precipitated crystals were verified as struvite, using X-ray diffraction and scanning electron microscopy analysis. The maximum P precipitation efficiency achieved by the MEC was 95%, and the maximum H 2 production rate was 0.28m 3 -H 2 /m 3 -d. Response surface methodology showed that applied voltage had a great influence on H 2 production and P recovery, while influent COD concentration had a significant effect on P recovery only. The overall energy recovery in the MEC was low and ranged from 25±1 to 37±1.7%. These results confirmed MECs capability for concurrent H 2 production and P recovery. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Bakonyi, Péter; Kumar, Gopalakrishnan; Koók, László; Tóth, Gábor; Rózsenberszki, Tamás; Bélafi-Bakó, Katalin; Nemestóthy, Nándor
2018-03-01
Microbial electrohydrogenesis cells (MECs) are devices that have attracted significant attention from the scientific community to generate hydrogen gas electrochemically with the aid of exoelectrogen microorganisms. It has been demonstrated that MECs are capable to deal with the residual organic materials present in effluents generated along with dark fermentative hydrogen bioproduction (DF). Consequently, MECs stand as attractive post-treatment units to enhance the global H 2 yield as a part of a two-stage, integrated application (DF-MEC). In this review article, it is aimed (i) to assess results communicated in the relevant literature on cascade DF-MEC systems, (ii) describe the characteristics of each steps involved and (iii) discuss the experiences as well as the lessons in order to facilitate knowledge transfer and help the interested readers with the construction of more efficient coupled set-ups, leading eventually to the improvement of overall biohydrogen evolution performances. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Qin, Mohan; Molitor, Hannah; Brazil, Brian; Novak, John T; He, Zhen
2016-01-01
A microbial electrolysis cell (MEC)-forward osmosis (FO) system was previously reported for recovering ammonium and water from synthetic solutions, and here it has been advanced with treating landfill leachate. In the MEC, 65.7±9.1% of ammonium could be recovered in the presence of cathode aeration. Without aeration, the MEC could remove 54.1±10.9% of ammonium from the leachate, but little ammonia was recovered. With 2M NH4HCO3 as the draw solution, the FO process achieved 51% water recovery from the MEC anode effluent in 3.5-h operation, higher than that from the raw leachate. The recovered ammonia was used as a draw solute in the FO for successful water recovery from the treated leachate. Despite the challenges with treating returning solution from the FO, this MEC-FO system has demonstrated the potential for resource recovery from wastes, and provide a new solution for sustainable leachate management. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Grid scale drives the scale and long-term stability of place maps
Mallory, Caitlin S; Hardcastle, Kiah; Bant, Jason S; Giocomo, Lisa M
2018-01-01
Medial entorhinal cortex (MEC) grid cells fire at regular spatial intervals and project to the hippocampus, where place cells are active in spatially restricted locations. One feature of the grid population is the increase in grid spatial scale along the dorsal-ventral MEC axis. However, the difficulty in perturbing grid scale without impacting the properties of other functionally-defined MEC cell types has obscured how grid scale influences hippocampal coding and spatial memory. Here, we use a targeted viral approach to knock out HCN1 channels selectively in MEC, causing grid scale to expand while leaving other MEC spatial and velocity signals intact. Grid scale expansion resulted in place scale expansion in fields located far from environmental boundaries, reduced long-term place field stability and impaired spatial learning. These observations, combined with simulations of a grid-to-place cell model and position decoding of place cells, illuminate how grid scale impacts place coding and spatial memory. PMID:29335607
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2012-01-01
Energy consumption in the U.S. manufacturing sector fell from 21,098 trillion Btu (tBtu) in 2006 to 19,062 tBtu in 2010, a decline of almost 10% , based on preliminary estimates released from the 2010 Manufacturing Energy Consumption Survey (MECS). This decline continues the downward trend in manufacturing energy use since the 1998 MECS report.
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Shim, Eun-Hwa; Kim, Jaisoo; Cho, Kyung-Suk; Ryu, Hee Wook
2006-05-01
This study evaluated the individual and combined removal capacities of benzene, toluene, and xylene (B, T, and X) in the presence and absence of methyl tert-butyl ether (MTBE) in a polyurethane biofilter inoculated with a BTX-degrading microbial consortium, and further examined their interactive effects in various mixtures. In addition, Polymerase chain reaction-denaturing gradient gel electrophoresis and phylogenetic analysis of 16S rRNA gene sequences were used to compare the microbial community structures found in biofilters exposed to the various gases and gas mixtures. The maximum individual elimination capacities (MECs) of B, T, and X were 200, 238, and 400 g m(-3) h(-1), respectively. There was no significant elimination of MTBE alone. Addition of MTBE decreased the MECs of B,T, and X to 75, 100, and 300 g m(-3) h(-1), respectively, indicating that benzene was most strongly inhibited by MTBE. When the three gases were mixed (B + T + X), the removal capacities of individual B, T, and X were 50, 90, and 200 g m(-3) h(-1), respectively. These capacities decreased to 40, 50, and 100 g m(-3) h(-1) when MTBE was added to the mix. The MEC of the three-gas mixture (B + T + X) was 340 g m(-3) h(-1), and that of the four-gas mixture was 200 g m(-3) h(-1). Although MTBE alone was not degraded by the biofilter, it could be co-metabolically degraded in the presence of toluene, benzene, or xylene with the MECs of 34, 23, and 14 g m(-3) h(-1), respectively. The microbial community structure analysis revealed that two large groups could be distinguished based on the presence or absence of MTBE, and many of the dominant bacteria in the consortia were closely related to bacteria isolated from aromatic hydrocarbon-contaminated sites and/ or oil wastewaters. These findings provide important new insights into biofiltration and may be used to improve the rational design of biofilters for remediation of petroleum gas-contaminated airstreams according to composition types of mixed gases.
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Díaz-Murillo, Violeta; Medina-Estrada, Ivan; López-Meza, Joel E; Ochoa-Zarzosa, Alejandra
2016-04-01
β-Defensins are members of the antimicrobial peptide superfamily that are produced in various species from different kingdoms, including plants. Plant defensins exhibit primarily antifungal activities, unlike those from animals that exhibit a broad-spectrum antimicrobial action. Recently, immunomodulatory roles of mammal β-defensins have been observed to regulate inflammation and activate the immune system. Similar roles for plant β-defensins remain unknown. In addition, the regulation of the immune system by mammalian β-defensins has been studied in humans and mice models, particularly in immune cells, but few studies have investigated these peptides in epithelial cells, which are in intimate contact with pathogens. The aim of this work was to evaluate the effect of the chemically synthesized β-defensin γ-thionin from Capsicum chinense on the innate immune response of bovine mammary epithelial cells (bMECs) infected with Staphylococcus aureus, the primary pathogen responsible for bovine mastitis, which is capable of living within bMECs. Our results indicate that γ-thionin at 0.1 μg/ml was able to reduce the internalization of S. aureus into bMECs (∼50%), and it also modulates the innate immune response of these cells by inducing the mRNA expression (∼5-fold) and membrane abundance (∼3-fold) of Toll-like receptor 2 (TLR2), as well as by inducing genes coding for the pro-inflammatory cytokines TNF-α and IL-1β (∼14 and 8-fold, respectively) before and after the bacterial infection. γ-Thionin also induces the expression of the mRNA of anti-inflammatory cytokine IL-10 (∼12-fold). Interestingly, the reduction in bacterial internalization coincides with the production of other antimicrobial products by bMECs, such as NO before infection, and the secretion into the medium of the endogenous antimicrobial peptide DEFB1 after infection. The results from this work support the potential use of β-defensins from plants as immunomodulators of the mammalian innate immune response. Copyright © 2016 Elsevier Inc. All rights reserved.
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Raftopoulos, Harry; Cooper, William; O'Boyle, Erin; Gabrail, Nashat; Boccia, Ralph; Gralla, Richard J
2015-03-01
Subcutaneous APF530 provides controlled sustained release of granisetron to prevent acute (0-24 h) and delayed (24-120 h) chemotherapy-induced nausea and vomiting (CINV). This randomized, double-blind phase 3 trial compared APF530 and palonosetron in preventing acute and delayed CINV after moderately (MEC) or highly emetogenic chemotherapy (HEC). Patients receiving single-day MEC or HEC received single-dose APF530 250 or 500 mg subcutaneously (SC) (granisetron 5 or 10 mg) or intravenous palonosetron 0.25 mg. Primary objectives were to establish APF530 noninferiority to palonosetron for preventing acute CINV following MEC or HEC and delayed CINV following MEC and to determine APF530 superiority to palonosetron for preventing delayed CINV following HEC. The primary efficacy end point was complete response (CR [using CI difference for APF530-palonosetron]). A lower confidence bound greater than -15 % indicated noninferiority. In the modified intent-to-treat population (MEC = 634; HEC = 707), both APF530 doses were noninferior to palonosetron in preventing acute CINV after MEC (CRs 74.8 % [-9.8, 9.3] and 76.9 % [-7.5, 11.4], respectively, vs. 75.0 % palonosetron) and after HEC (CRs 77.7 % [-11.5, 5.5] and 81.3 % [-7.7, 8.7], respectively, vs. 80.7 % palonosetron). APF530 500 mg was noninferior to palonosetron in preventing delayed CINV after MEC (CR 58.5 % [-9.5, 12.1] vs. 57.2 % palonosetron) but not superior in preventing delayed CINV after HEC. Adverse events were generally mild and unrelated to treatment, the most common (excluding injection-site reactions) being constipation. A single subcutaneous APF530 injection offers a convenient alternative to palonosetron for preventing acute and delayed CINV after MEC or HEC.
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Natural DNA transformation is functional in Lactococcus lactis ssp. cremoris KW2.
David, Blandine; Radziejwoski, Amandine; Toussaint, Frédéric; Fontaine, Laetitia; Henry de Frahan, Marie; Patout, Cédric; van Dillen, Sabine; Boyaval, Patrick; Horvath, Philippe; Fremaux, Christophe; Hols, Pascal
2017-06-16
Lactococcus lactis is one of the most commonly used lactic acid bacteria in the dairy industry. Activation of competence for natural DNA transformation in this species would greatly improve the selection of novel strains with desired genetic traits. Here, we investigated the activation of natural transformation in L. lactis ssp. cremoris KW2, a strain of plant origin whose genome encodes the master competence regulator ComX and the complete set of proteins usually required for natural transformation. In the absence of knowledge about competence regulation in this species, we constitutively overproduced ComX in a reporter strain of late competence phase activation and showed, by transcriptomic analyses, a ComX-dependent induction of all key competence genes. We further demonstrated that natural DNA transformation is functional in this strain and requires the competence DNA uptake machinery. Since constitutive ComX overproduction is unstable, we alternatively expressed comX under the control of an endogenous xylose-inducible promoter. This regulated system was used to successfully inactivate the adaptor protein MecA and subunits of the Clp proteolytic complex, which were previously shown to be involved in ComX degradation in streptococci. In the presence of a low amount of ComX, the deletion of mecA , clpC , or clpP genes markedly increased the activation of the late competence phase and transformability. Altogether, our results report the functionality of natural DNA transformation in L. lactis and pave the way for the identification of signaling mechanisms that trigger the competence state in this species. IMPORTANCE Lactococcus lactis is a lactic acid bacterium of major importance, which is used as a starter species for milk fermentation, a host for heterologous protein production, and a delivery platform for therapeutic molecules. Here, we report the functionality of natural transformation in L. lactis ssp. cremoris KW2 by the overproduction of the master competence regulator ComX. The developed procedure enables a flexible approach to modify the chromosome with single point mutation, sequence insertion, or sequence replacement. These results represent an important step for the genetic engineering of L. lactis that will facilitate the design of strains optimized for industrial applications. This will also help to discover natural regulatory mechanisms controlling competence in the genus Lactococcus . Copyright © 2017 American Society for Microbiology.
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Pieri, Fabio A; Vargas, Taise F; Galvão, Newton N; Nogueira, Paulo A; Orlandi, Patrícia P
2016-03-01
The aim of this study was to characterize and compare Staphylococcus spp. isolated from hospitalized patients and beef marketed in the city of Porto Velho-RO, Brazil. The isolates were subjected to antibiogram tests, adherence capacity tests, detection of the mecA gene, and epidemiological investigation by the random amplified polymorphic DNA (RAPD) technique, using the primers M13 and H12. Among the 123 Staphylococcus spp. isolates, 50 were identified as S. aureus and 73 as coagulase-negative Staphylococcus; among the latter, 7 species were identified. It was observed that the coagulase-negative Staphylococcus isolates showed greater adhesion ability than S. aureus. The profile of antimicrobial susceptibility was different among isolates, all of which were susceptible to vancomycin and linezolid, and had high penicillin resistance rates, varying according to the bacterial class and the source. In this study, all strains were negative for mecA gene detection; however, 36% of S. aureus and 17% of coagulase-negative Staphylococcus were resistant to oxacillin. The genetic relationship of these bacteria, analyzed by RAPD, was able to discriminate the species of coagulase-negative Staphylococcus strains of S. aureus along its origin. It was concluded that the isolates of Staphylococcus spp. derived from beef and human infections differ genetically. Thus, it is suggested that isolates from beef, which were grouped within hospital isolates, were probably carried via contact with beef in hospital professionals or patients.
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Gómez, Paula; Lozano, Carmen; Benito, Daniel; Estepa, Vanesa; Tenorio, Carmen; Zarazaga, Myriam; Torres, Carmen
2016-05-01
The objective of this study was to determine the prevalence of Staphylococcus in urban wastewater treatment plants (UWTP) of La Rioja (Spain), and to characterize de obtained isolates. 16 wastewater samples (8 influent, 8 effluent) of six UWTPs were seeded on mannitol-salt-agar and oxacillin-resistance-screening-agar-base for staphylococci and methicillin-resistant Staphylococcus aureus recovery. Antimicrobial susceptibility profile was determined for 16 antibiotics and the presence of 35 antimicrobial resistance genes and 14 virulence genes by PCR. S. aureus was typed by spa, agr, and multilocus-sequence-typing, and the presence of immune-evasion-genes cluster was analyzed. Staphylococcus spp. were detected in 13 of 16 tested wastewater samples (81%), although the number of CFU/mL decreased after treatment. 40 staphylococci were recovered (1-5/sample), and 8 of them were identified as S. aureus being typed as (number of strains): spa-t011/agr-II/ST398 (1), spa-t002/agr-II/ST5 (2), spa-t3262/agr-II/ST5 (1), spa-t605/agr-II/ST126 (3), and spa-t878/agr-III/ST2849 (1). S. aureus ST398 strain was methicillin-resistant and showed a multidrug resistance phenotype. Virulence genes tst, etd, sea, sec, seg, sei, sem, sen, seo, and seu, were detected among S. aureus and only ST5 strains showed genes of immune evasion cluster. Thirty-two coagulase-negative Staphylococcus of 12 different species were recovered (number of strains): Staphylococcus equorum (7), Staphylococcus vitulinus (4), Staphylococcus lentus (4), Staphylococcus sciuri (4), Staphylococcus fleurettii (2), Staphylococcus haemolyticus (2), Staphylococcus hominis (2), Staphylococcus saprophyticus (2), Staphylococcus succinus (2), Staphylococcus capitis (1), Staphylococcus cohnii (1), and Staphylococcus epidermidis (1). Five presented a multidrug resistance phenotype. The following resistance and virulence genes were found: mecA, lnu(A), vga(A), tet(K), erm(C), msr(A)/(B), mph(C), tst, and sem. We found that Staphylococcus spp. are normal contaminants of urban wastewater, including different lineages of S. aureus and a high diversity of coagulase-negative species. The presence of multiple resistance and virulence genes, including mecA, in staphylococci of wastewater can be a concern for the public health. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Takiya, Shigeharu; Tsubota, Takuya; Kimoto, Mai
2016-01-01
The silk gland of the silkworm Bombyx mori is a long tubular organ that is divided into several subparts along its anteroposterior (AP) axis. As a trait of terminal differentiation of the silk gland, several silk protein genes are expressed with unique regional specificities. Most of the Hox and some of the homeobox genes are also expressed in the differentiated silk gland with regional specificities. The expression patterns of Hox genes in the silk gland roughly correspond to those in embryogenesis showing “colinearity”. The central Hox class protein Antennapedia (Antp) directly regulates the expression of several middle silk gland–specific silk genes, whereas the Lin-1/Isl-1/Mec3 (LIM)-homeodomain transcriptional factor Arrowhead (Awh) regulates the expression of posterior silk gland–specific genes for silk fiber proteins. We summarize our results and discuss the usefulness of the silk gland of Bombyx mori for analyzing the function of Hox genes. Further analyses of the regulatory mechanisms underlying the region-specific expression of silk genes will provide novel insights into the molecular bases for target-gene selection and regulation by Hox and homeodomain proteins. PMID:29615585
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Bochniarz, M; Adaszek, Ł; Dzięgiel, B; Nowaczek, A; Wawron, W; Dąbrowski, R; Szczubiał, M; Winiarczyk, S
2016-12-01
The aim of this study was to recognize selected factors of virulence determining the adhesion of Staphylococcus chromogenes to cows' udder tissues in subclinical mastitis and to evaluate the susceptibility of this pathogen to antibiotics. The subjects of the study were 38 isolates of Staph. chromogenes from 335 samples of milk from cows with subclinical coagulase-negative staphylococci mastitis. Somatic cell count ranged between 216,000 and 568,000/mL of milk (average 356,000/mL of milk). We confirmed the ability to produce slime in 24 isolates (63.2%), and the ability to produce protease in 29 isolates (76.3%). In each slime-producing isolate, the bap gene was not found, and the fnbA and eno genes were not detected. In vitro tests showed that ceftiofur had the highest effectiveness against Staph. chromogenes (89.5% of susceptible isolates). Minimum inhibitory concentrations ranged from 0.06 to 2µg/mL for susceptible isolates. The minimum concentrations required to inhibit growth of 90 and 50% of the isolates for ceftiofur were at or below the cutoffs recommended by the Clinical and Laboratory Standards Institute (2 and 0.06µg/mL, respectively). A significant percentage of the isolates were susceptible to other β-lactam antibiotics: amoxicillin with clavulanic acid (84.2%) and ampicillin (81.6%). The lowest effectiveness among β-lactams was for penicillin (73.7% of susceptible isolates), and the minimum inhibitory concentration for penicillin ranged from <0.06 to 8µg/mL. None of the examined isolates had the mecA gene, but β-lactamase was detected in 4 isolates (10.5%). Erythromycin and oxytetracycline exhibited the lowest activity against Staph. chromogenes (71.1 and 63.2% of susceptible isolates, respectively). The genes tetK (6 isolates) and ermA (1 isolate) were also detected. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
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NASA Astrophysics Data System (ADS)
Masud, Salwa F.; Raufer, Stefan; Neely, Stephen T.; Nakajima, Hideko H.
2018-05-01
Superior canal dehiscence (SCD) is a hole in the bony wall of the superior semicircular canal, which can cause various auditory and/or vestibular symptoms and can result in wrong and/or delayed diagnosis. Wideband acoustic immittance (WAI) can potentially distinguish various mechanical middle-ear pathologies as well as inner-ear pathologies non-invasively. We found that in patients, SCD was commonly associated with a narrow-band decrease in power reflectance (PR, derived from WAI) near 1 kHz. Because clinical data has large variation across individual ears and because we do not know the individual "normal" state prior to SCD, we measured WAI in five fresh temporal bone specimens to determine the effects of SCD with respect to the normal state. In temporal bone, we measured PR to assess mechanical changes before and after SCD, as well as to assess the effect of an open or closed middle-ear cavity. After SCD, PR had a consistent decrease between 0.48 and 0.76 kHz, and a slight increase between 1.04 and 1.4 kHz in the open cavity condition. However, in several experiments, we observed low PR around 1 kHz in the normal state before SCD, likely due to the specimen's open middle ear cavity (MEC). Because we see effects of both SCD and open MEC around 1 kHz, some of the SCD effect can be masked by the effect of the MEC in the temporal bone specimens. To compensate for this MEC effect, we estimated the effect of SCD in a closed MEC case, but the effect did not differ significantly from the measured open MEC. This study demonstrates the limitation of temporal bone experiments with open MEC when studying inner-ear lesions with WAI.
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NASA Astrophysics Data System (ADS)
Yu, Xiao; Blackmon, Richard L.; Carabas-Hernendez, Patricia; Fuller, Ashley; Troester, Melissa A.; Oldenburg, Amy L.
2016-03-01
Mammary epithelial cell (MEC) organoids in 3D culture recapitulate features of breast ducts in vivo. OCT has the ability to monitor the evolution of MEC organoids non-invasively and longitudinally. The anti-cancer drug Doxorubicin (Dox) is able to inhibit proliferation of cancer cells and has been widely used for chemotherapy of breast cancers; while environmental toxins implicated in breast cancer such as estrogen regulates mammary tumor growth and stimulates the proliferation and metastatic potential of breast cancers. Here we propose a quantitative method for measuring motility of breast cells in 3D cultures based upon OCT speckle fluctuation spectroscopy. The metrics of the inverse power-law exponent (α) and fractional modulation amplitude (M) were extracted from speckle fluctuation spectra. These were used to quantify the responses of MEC organoids to Dox, and estrogen. We investigated MEC organoids comprised of two different MEC lines: MCF10DCIS.com exposed to Dox, and MCF7 exposed to estrogen. We found an increase (p<0.001) in α of MEC along time (t=0, 1 hour, 24 hours, 48 hours and 6 days) at each dose of Dox (0, 1 μM and 10 μM), indicating lower fluctuation intensity at higher frequencies. We also observed a decrease (p<0.001) in M for increasing time. However, both α and M of MCF7 treated with estrogen (0, 1 nM and 10 nM) exhibited the opposite trend along time. This novel technology provides rapid and non-invasive measurements of the effects of toxicants on MEC motility for understanding breast cancer development and assessing anti-cancer drugs.
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Al-Habib, Amro F; Al-Rabie, Abdulkarim; Aleissa, Sami; Albakr, Abdulrahman; Abobotain, Abdulaziz
2017-01-01
This was an interventional human cadaver study and radiological study. Atlas instrumentation is frequently involved in fusion procedures involving the craniocervical junction area. Identification of the entry point at the center of atlas lateral mass (ALM) is challenging because of its rounded posterior surface and the surrounding venous plexus. This report examines using the medial edge of atlas posterior arch (MEC1) as a fixed and reliable anatomic reference to guide the entry point of ALM screws. Fifty, normal, cervical spine computed tomography studies were reviewed. ALM screw trajectories were planned at one point along MEC1 and another point 2 mm lateral to MEC1. Free-hand ALM instrumentation was performed in ten fresh human cadavers using the 2 mm entry point, with a sagittal trajectory parallel to atlas inferior arch (IAC1); three-dimensional imaging was then performed to confirm instrumentation accuracy. The average ALM diameter was 12.35 mm. Inserting a screw using the entry point 2 mm lateral to MEC1 was closer to ALM midpoint than using the entry point along MEC1 ( P < 0.0001). Twenty ALM screws were successfully inserted in the ten cadavers. No encroachments into the spinal canal or foramen transversarium occurred. However, two screws were superiorly directed and violated the occipitocervical joint; they were not parallel to IAC1. MEC1 provides a fixed and reliable landmark for ALM instrumentation. An entry point 2 mm point lateral to MEC1 is close to ALM midpoint. IAC1 also provides a guide for the sagittal trajectory. Attention to anatomic landmarks may help reduce complications associated with atlas instrumentation but should be verified in future clinical studies.
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Zhao, Xiangshan; Malhotra, Gautam K.; Band, Hamid; Band, Vimla
2011-01-01
Introduction: Emerging evidence suggests a direct role of cancer stem cells (CSCs) in the development of breast cancer. In vitro cellular models that recapitulate properties of CSCs are therefore highly desirable. We have previously shown that normal human mammary epithelial cells (hMECs) immortalized with human telomerase reverse transcriptase (hTERT) possess properties of mammary stem / progenitor cells. Materials and Methods: In the present study, we used this cell system to test the idea that other known hMEC-immortalizing oncogenes (RhoA, HPVE6, HPVE7, p53 mutant, and treatment with γ-radiation), share with hTERT, the ability to maintain mammary stem / progenitor cells. Results: The results presented here demonstrate that similar to hMECs immortalized with hTERT, all hMEC cell lines immortalized using various oncogenic strategies express stem / progenitor cell markers. Furthermore, analyses using 2D and 3D culture assays demonstrate that all the immortal cell lines retain their ability to self-renew and to differentiate along the luminal lineage. Remarkably, the stem / progenitor cell lines generated using various oncogenic strategies exhibit a block in differentiation along the myoepithelial lineage, a trait that is retained on hTERT-immortalized stem / progenitors. The inability to differentiate along the myoepithelial lineage could be induced by ectopic mutant p53 expression in hTERT-immortalized hMEC. Conclusions: Our studies demonstrate that stem / progenitor cell characteristics of hMECs are maintained upon immortalization by using various cancer-relevant oncogenic strategies. Oncogene-immortalized hMECs show a block in their ability to differentiate along the myoepithelial lineage. Abrogation of the myoepithelial differentiation potential by a number of distinct oncogenic insults suggests a potential explanation for the predominance of luminal and rarity of myoepithelial breast cancers. PMID:22279424
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Zhao, Xiangshan; Malhotra, Gautam K; Band, Hamid; Band, Vimla
2011-01-01
Emerging evidence suggests a direct role of cancer stem cells (CSCs) in the development of breast cancer. In vitro cellular models that recapitulate properties of CSCs are therefore highly desirable. We have previously shown that normal human mammary epithelial cells (hMECs) immortalized with human telomerase reverse transcriptase (hTERT) possess properties of mammary stem / progenitor cells. In the present study, we used this cell system to test the idea that other known hMEC-immortalizing oncogenes (RhoA, HPVE6, HPVE7, p53 mutant, and treatment with γ-radiation), share with hTERT, the ability to maintain mammary stem / progenitor cells. The results presented here demonstrate that similar to hMECs immortalized with hTERT, all hMEC cell lines immortalized using various oncogenic strategies express stem / progenitor cell markers. Furthermore, analyses using 2D and 3D culture assays demonstrate that all the immortal cell lines retain their ability to self-renew and to differentiate along the luminal lineage. Remarkably, the stem / progenitor cell lines generated using various oncogenic strategies exhibit a block in differentiation along the myoepithelial lineage, a trait that is retained on hTERT-immortalized stem / progenitors. The inability to differentiate along the myoepithelial lineage could be induced by ectopic mutant p53 expression in hTERT-immortalized hMEC. Our studies demonstrate that stem / progenitor cell characteristics of hMECs are maintained upon immortalization by using various cancer-relevant oncogenic strategies. Oncogene-immortalized hMECs show a block in their ability to differentiate along the myoepithelial lineage. Abrogation of the myoepithelial differentiation potential by a number of distinct oncogenic insults suggests a potential explanation for the predominance of luminal and rarity of myoepithelial breast cancers.
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In Vitro Susceptibility Testing Methods for Caspofungin against Aspergillus and Fusarium Isolates
Arikan, Sevtap; Lozano-Chiu, Mario; Paetznick, Victor; Rex, John H.
2001-01-01
We investigated the relevance of prominent reduction in turbidity macroscopically (MIC) and formation of aberrant hyphal tips microscopically (minimum effective concentration; MEC) in measuring the in vitro activity of caspofungin against Aspergillus and Fusarium. Caspofungin generated low MICs and MECs against Aspergillus, but not for Fusarium. While MICs increased inconsistently when the incubation time was prolonged, MEC appeared as a stable and potentially relevant endpoint in testing in vitro caspofungin activity. PMID:11120990
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LaBarge, Nicole; Yilmazel, Yasemin Dilsad; Hong, Pei-Ying; Logan, Bruce E
2017-02-01
Microbial electrolysis cells (MECs) can generate methane by fixing carbon dioxide without using expensive catalysts, but the impact of acclimation procedures on subsequent performance has not been investigated. Granular activated carbon (GAC) was used to pre-enrich electrotrophic methanogenic communities, as GAC has been shown to stimulate direct transfer of electrons between different microbial species. MEC startup times using pre-acclimated GAC were improved compared to controls (without pre-acclimation or without GAC), and after three fed batch cycles methane generation rates were similar (P>0.4) for GAC acclimated to hydrogen (22±9.3nmolcm -3 d -1 ), methanol (25±9.7nmolcm -3 d -1 ), and a volatile fatty acid (VFA) mix (22±11nmolcm -3 d -1 ). However, MECs started with GAC but no pre-acclimation had lower methane generation rates (13±4.1nmolcm -3 d -1 ), and MECs without GAC had the lowest rates (0.7±0.8nmolcm -3 d -1 after cycle 2). Microbes previously found in methanogenic MECs, or previously shown to be capable of exocellular electron transfer, were enriched on the GAC. Pre-acclimation using GAC is therefore a simple approach to enrich electroactive communities, improve methane generation rates, and decrease startup times in MECs. Copyright © 2016 Elsevier B.V. All rights reserved.
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Aboul-Fotouh, Sawsan
2015-03-01
Several studies have pointed to the nicotinic acetylcholine receptor (nAChR) antagonists, such as mecamylamine (MEC), as a potential therapeutic target for the treatment of depression. The present study evaluated the behavioral and neurochemical effects of chronic administration of MEC (1, 2, and 4 mg/kg/day, intraperitoneally (i.p.)) in Wistar rats exposed to chronic restraint stress (CRS, 4 h × 6 W). MEC prevented CRS-induced depressive-like behavior via increasing sucrose preference, body weight, and forced swim test (FST) struggling and swimming while reducing immobility in FST and hypothalamic-pituitary-adrenal (HPA) axis hyperactivity (adrenal gland weight and serum corticosterone). At the same time, MEC amended CRS-induced anxiety as indicated by decreasing central zone duration in open field test and increasing active interaction duration. Additionally, MEC modulated the prefrontal cortex (PFC) level of brain-derived neurotrophic factor (BDNF), 5-hydroxy tryptamine (5-HT), and norepinephrine (NE). In conclusion, the present data suggest that MEC possesses antidepressant and anxiolytic-like activities in rats exposed to CRS. These behavioral effects may be in part mediated by reducing HPA axis hyperactivity and increasing PFC level of BDNF and monoamines. Accordingly, these findings further support the hypothesis that nAChRs blockade might afford a novel promising strategy for pharmacotherapy of depression.
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Li, Jinlong; Li, Desheng; Cui, Yuwei; Xing, Wei; Deng, Shihai
2017-07-01
Nitrogen bioremediation in organic insufficient wastewater generally requires an extra carbon source. In this study, nitrate-contaminated wastewater was treated effectively through simultaneous autotrophic and heterotrophic denitrification based on micro-electrolysis carriers (MECs) and retinervus luffae fructus (RLF), respectively. The average nitrate and total nitrogen removal rates reached 96.3 and 94.0% in the MECs/RLF-based autotrophic and heterotrophic denitrification (MRAHD) system without ammonia and nitrite accumulation. The performance of MRAHD was better than that of MEC-based autotrophic denitrification for the wastewater treatment with low carbon nitrogen (COD/N) ratio. Real-time quantitative polymerase chain reaction (qPCR) revealed that the relative abundance of nirS-type denitrifiers attached to MECs (4.9%) and RLF (5.0%) was similar. Illumina sequencing suggested that the dominant genera were Thiobacillus (7.0%) and Denitratisoma (5.7%), which attached to MECs and RLF, respectively. Sulfuritalea was discovered as the dominant genus in the middle of the reactor. The synergistic interaction between autotrophic and heterotrophic denitrifiers played a vital role in the mixotrophic substrate environment.
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Mec1/ATR, the Program Manager of Nucleic Acids Inc.
Feng, Wenyi
2016-12-28
Eukaryotic cells are equipped with surveillance mechanisms called checkpoints to ensure proper execution of cell cycle events. Among these are the checkpoints that detect DNA damage or replication perturbations and coordinate cellular activities to maintain genome stability. At the forefront of damage sensing is an evolutionarily conserved molecule, known respectively in budding yeast and humans as Mec1 (Mitosis entry checkpoint 1) and ATR (Ataxia telangiectasia and Rad3-related protein). Through phosphorylation, Mec1/ATR activates downstream components of a signaling cascade to maintain nucleotide pool balance, protect replication fork integrity, regulate activation of origins of replication, coordinate DNA repair, and implement cell cycle delay. This list of functions continues to expand as studies have revealed that Mec1/ATR modularly interacts with various protein molecules in response to different cellular cues. Among these newly assigned functions is the regulation of RNA metabolism during checkpoint activation and the coordination of replication-transcription conflicts. In this review, I will highlight some of these new functions of Mec1/ATR with a focus on the yeast model organism.
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Lee, Mi-Young; Kim, Kyoung-Yeol; Yang, Euntae; Kim, In S
2015-01-01
In order to enhance hydrogen production by facilitated proton transport through a forward osmosis (FO) membrane, the FO membrane was integrated into microbial electrolysis cells (MECs). An improved hydrogen production rate was obtained in the FO-MEC (12.5±1.84×10(-3)m(3)H2/m(3)/d) compared to that of the cation exchange membrane (CEM) - MEC (4.42±0.04×10(-3)m(3)H2/m(3)/d) during batch tests (72h). After an internal resistance analysis, it was confirmed that the enhanced hydrogen production in FO-MEC was attributed to the smaller charge transfer resistance than in the CEM-MEC (90.3Ω and 133.4Ω respectively). The calculation of partial internal resistance concluded that the transport resistance can be substantially reduced by replacing a CEM with a FO membrane; decrease of the resistance from 0.069Ωm(2) to 5.99×10(-4)Ωm(2). Copyright © 2015 Elsevier Ltd. All rights reserved.
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Hydrogen production profiles using furans in microbial electrolysis cells.
Catal, Tunc; Gover, Tansu; Yaman, Bugra; Droguetti, Jessica; Yilancioglu, Kaan
2017-06-01
Microbial electrochemical cells including microbial fuel cells (MFCs) and microbial electrolysis cells (MECs) are novel biotechnological tools that can convert organic substances in wastewater or biomass into electricity or hydrogen. Electroactive microbial biofilms used in this technology have ability to transfer electrons from organic compounds to anodes. Evaluation of biofilm formation on anode is crucial for enhancing our understanding of hydrogen generation in terms of substrate utilization by microorganisms. In this study, furfural and hydroxymethylfurfural (HMF) were analyzed for hydrogen generation using single chamber membrane-free MECs (17 mL), and anode biofilms were also examined. MECs were inoculated with mixed bacterial culture enriched using chloroethane sulphonate. Hydrogen was succesfully produced in the presence of HMF, but not furfural. MECs generated similar current densities (5.9 and 6 mA/cm 2 furfural and HMF, respectively). Biofilm samples obtained on the 24th and 40th day of cultivation using aromatic compounds were evaluated by using epi-fluorescent microscope. Our results show a correlation between biofilm density and hydrogen generation in single chamber MECs.
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The improvement of the method of equivalent cross section in HTR
DOE Office of Scientific and Technical Information (OSTI.GOV)
Guo, J.; Li, F.
The Method of Equivalence Cross-Sections (MECS) is a combined transport-diffusion method. By appropriately adjusting the diffusion coefficient of homogenized absorber region, the diffusion theory could yield satisfactory results for the full core model with strong neutron absorber material, for example the control rod in High temperature gas cooled reactor (HTR). Original implementation of MECS based on 1-D cell transport model has some limitation on accuracy and applicability, a new implementation of MECS based on 2-D transport model are proposed and tested in this paper. This improvement can extend the MECS to the calculation of twin small absorber ball system whichmore » have a non-circular boring in graphite reflector and different radial position. A least-square algorithm for the calculation of equivalent diffusion coefficient is adopted, and special treatment for diffusion coefficient for higher energy group is proposed in the case that absorber is absent. Numerical results to adopt MECS into control rod calculation in HTR are encouraging. However, there are some problems left. (authors)« less
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Yang, Xiaojuan; Zhang, Jumei; Yu, Shubo; Wu, Qingping; Guo, Weipeng; Huang, Jiahui; Cai, Shuzhen
2016-01-01
Staphylococcus aureus, particularly methicillin-resistant S.aureus (MRSA), is a life-threatening pathogen in humans, and its presence in food is a public health concern. MRSA has been identified in foods in China, but little information is available regarding MRSA in ready-to-eat (RTE) foods. We aimed to investigate the prevalence of S. aureus and MRSA in Chinese retail RTE foods. All isolated S. aureus were tested for antimicrobial susceptibility, and MRSA isolates were further characterized by multilocus sequence typing (MLST) and staphylococcal cassette chromosome mec (SCCmec) typing. Of the 550 RTE foods collected from 2011 to 2014, 69 (12.5%) were positive for S. aureus. Contamination levels were mostly in the range of 0.3–10 most probable number (MPN)/g, with five samples exceeding 10 MPN/g. Of the 69 S. aureus isolates, seven were identified as MRSA by cefoxitin disc diffusion test. Six isolates were mecA-positive, while no mecC-positive isolates were identified. In total, 75.8% (47/62) of the methicillin-susceptible S. aureus isolates and all of the MRSA isolates were resistant to three or more antibiotics. Amongst the MRSA isolates, four were identified as community-acquired strains (ST59-MRSA-IVa (n = 2), ST338-MRSA-V, ST1-MRSA-V), while one was a livestock-associated strain (ST9, harboring an unreported SCCmec type 2C2). One novel sequence type was identified (ST3239), the SCCmec gene of which could not be typed. Overall, our findings showed that Chinese retail RTE foods are likely vehicles for transmission of multidrug-resistant S. aureus and MRSA lineages. This is a serious public health risk and highlights the need to implement good hygiene practices. PMID:27375562
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Snyder, James W.; Munier, Gina K.; Johnson, Charles L.
2010-01-01
This study compared the BD GeneOhm methicillin-resistant Staphylococcus aureus (MRSA) real-time PCR assay to culture by the use of BBL CHROMagar MRSA for the detection of MRSA in 627 nasal surveillance specimens collected from intensive care unit (ICU) patients. The PCR assay had a sensitivity, specificity, positive predictive value, and negative predictive value of 100%, 96.7%, 70.3%, and 100%, respectively. Nine of 19 false-positive PCR specimens grew methicillin-susceptible S. aureus (MSSA) from broth enrichment culture, of which two demonstrated evidence of mecA gene dropout. Compared to culture by the use of BBL CHROMagar MRSA, the BD GeneOhm MRSA PCR assay demonstrated sensitivity and specificity above 95% for the detection of MRSA nasal colonization and provided shorter turnaround time in generating positive and negative final results. PMID:20181916
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Presence of antibiotic resistance genes in a sewage treatment plant in Thibodaux, Louisiana, USA.
Naquin, Anthony; Shrestha, Arsen; Sherpa, Mingma; Nathaniel, Rajkumar; Boopathy, Raj
2015-01-01
Increasing uses and disposals of antibiotics to the environment have increased emergence of various antibiotic resistance. One of the sources for the spread of antibiotic resistance is wastewater treatment plant, where bacteria and antibiotics can come in contact and can acquire antibiotics resistance. There are very few studies on this subject from a small town sewage treatment plant. Therefore, this study was conducted using raw sewage as well as treated sewage from a sewage treatment plant in Thibodaux in rural southeast Louisiana in USA. Samples were collected monthly from the Thibodaux sewage treatment plant and the presence of antibiotic resistance genes was monitored. The study showed the presence of antibiotic resistance genes in both raw and treated sewage in every month of the study period. The genetic transformation assay showed the successful transformation of methicillin resistant gene, mecA to an antibiotic sensitive Staphylococcus aureus, which became antibiotic resistant within 24h. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Manufacturing Energy Consumption Survey (MECS)
2028-01-01
The Manufacturing Energy Consumption Survey (MECS) is a national sample survey that collects information on the stock of U.S. manufacturing establishment, their energy-related building characteristics, and their energy consumption and expenditures.
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Wang, Dexin; Han, Yuxing; Han, Hongjun; Li, Kun; Xu, Chunyan
2017-08-01
The coupling of micro-electrolysis cell (MEC) with an up-flow anaerobic sludge blanket (UASB) system in pilot scale was established for enhanced treatment of Fischer-Tropsch (F-T) wastewater. The lowest influent pH (4.99±0.10) and reduced alkali addition were accomplished under the assistance of anaerobic effluent recycling of 200% (stage 5). Simultaneously, the optimum COD removal efficiency (93.5±1.6%) and methane production (2.01±0.13m 3 /m 3 ·d) at the lower hydraulic retention time (HRT) were achieved in this stage. In addition, the dissolved iron from MEC could significantly increase the protein content of tightly bound extracellular polymeric substances (TB-EPS), which was beneficial to formation of stable granules. Furthermore, the high-throughput 16S rRNA gene pyrosequencing in this study further confirmed that Geobacter species could utilize iron oxides particles as electron conduit to perform the direct interspecies electron transfer (DIET) with Methanothrix, finally facilitating the syntrophic degradation of propionic acid and butyric acid and contributing completely methane production. Copyright © 2017 Elsevier Ltd. All rights reserved.
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A Visual-Cue-Dependent Memory Circuit for Place Navigation.
Qin, Han; Fu, Ling; Hu, Bo; Liao, Xiang; Lu, Jian; He, Wenjing; Liang, Shanshan; Zhang, Kuan; Li, Ruijie; Yao, Jiwei; Yan, Junan; Chen, Hao; Jia, Hongbo; Zott, Benedikt; Konnerth, Arthur; Chen, Xiaowei
2018-06-05
The ability to remember and to navigate to safe places is necessary for survival. Place navigation is known to involve medial entorhinal cortex (MEC)-hippocampal connections. However, learning-dependent changes in neuronal activity in the distinct circuits remain unknown. Here, by using optic fiber photometry in freely behaving mice, we discovered the experience-dependent induction of a persistent-task-associated (PTA) activity. This PTA activity critically depends on learned visual cues and builds up selectively in the MEC layer II-dentate gyrus, but not in the MEC layer III-CA1 pathway, and its optogenetic suppression disrupts navigation to the target location. The findings suggest that the visual system, the MEC layer II, and the dentate gyrus are essential hubs of a memory circuit for visually guided navigation. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.
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Milheiriço, Catarina; Portelinha, Ana; Krippahl, Ludwig; de Lencastre, Hermínia; Oliveira, Duarte C
2011-04-15
The β-lactamase (bla) locus, which confers resistance to penicillins only, may control the transcription of mecA, the central element of methicillin resistance, which is embedded in a polymorphic heterelogous chromosomal cassette (the SCCmec element). In order to assess the eventual correlation between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes, the allelic variation on the bla locus was evaluated in a representative collection of 54 international epidemic methicillin-resistant Staphylococcus aureus (MRSA) clinical strains and, for comparative purposes, also in 24 diverse methicillin-susceptible S. aureus (MSSA) strains. Internal fragments of blaZ (the β-lactamase structural gene) were sequenced for all strains. A subset of strains, representative of blaZ allotypes, was further characterized by sequencing of internal fragments of the blaZ transcriptional regulators, blaI and blaR1. Thirteen allotypes for blaZ, nine for blaI and 12 for blaR1 were found. In a total of 121 unique single-nucleotide polymorphisms (SNP) detected, no frameshift mutations were identified and only one nonsense mutation within blaZ was found in a MRSA strain. On average, blaZ alleles were more polymorphic among MSSA than in MRSA (14.7 vs 11.4 SNP/allele). Overall, blaR1 was the most polymorphic gene with an average of 24.8 SNP/allele. No correlation could be established between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes. In order to estimate the selection pressure acting on the bla locus, the average dN/dS values were computed. In the three genes and in both collections dN/dS ratios were significantly below 1. The data strongly suggests the existence of a purifying selection to maintain the bla locus fully functional even on MRSA strains. Although, this is in agreement with the notion that in most clinical MRSA strains mecA gene is under the control of the bla regulatory genes, these findings also suggest that the apparently redundant function of blaZ gene for the MRSA resistant phenotype is still important for these strains. In addition, the data shows that the sensor-inducer blaR1 is the primary target for the accumulation of mutations in the bla locus, presumably to modulate the response to the presence of β-lactam antibiotic.
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Takizawa, Nobukazu; Miyauchi, Maki; Yanai, Hiromi; Tateishi, Ryosuke; Shinzawa, Miho; Yoshinaga, Riko; Kurihara, Masaaki; Yasuda, Hisataka; Sakamoto, Reiko; Yoshida, Nobuaki
2016-01-01
Medullary thymic epithelial cells (mTECs) expressing autoimmune regulator (Aire) are critical for preventing the onset of autoimmunity. However, the differentiation program of Aire-expressing mTECs (Aire+ mTECs) is unclear. Here, we describe novel embryonic precursors of Aire+ mTECs. We found the candidate precursors of Aire+ mTECs (pMECs) by monitoring the expression of receptor activator of nuclear factor-κB (RANK), which is required for Aire+ mTEC differentiation. pMECs unexpectedly expressed cortical TEC molecules in addition to the mTEC markers UEA-1 ligand and RANK and differentiated into mTECs in reaggregation thymic organ culture. Introduction of pMECs in the embryonic thymus permitted long-term maintenance of Aire+ mTECs and efficiently suppressed the onset of autoimmunity induced by Aire+ mTEC deficiency. Mechanistically, pMECs differentiated into Aire+ mTECs by tumor necrosis factor receptor-associated factor 6-dependent RANK signaling. Moreover, nonclassical nuclear factor-κB activation triggered by RANK and lymphotoxin-β receptor signaling promoted pMEC induction from progenitors exhibiting lower RANK expression and higher CD24 expression. Thus, our findings identified two novel stages in the differentiation program of Aire+ mTECs. PMID:27401343
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Sim, Junyoung; An, Junyeong; Elbeshbishy, Elsayed; Ryu, Hodon; Lee, Hyung-Sool
2015-11-01
Cathode potential and O2 supply methods were investigated to improve H2O2 synthesis in an electrochemical cell, and optimal cathode conditions were applied for microbial electrochemical cells (MECs). Using aqueous O2 for the cathode significantly improved current density, but H2O2 conversion efficiency was negligible at 0.3-12%. Current density decreased for passive O2 diffusion to the cathode, but H2O2 conversion efficiency increased by 65%. An MEC equipped with a gas diffusion cathode was operated with acetate medium and domestic wastewater, which presented relatively high H2O2 conversion efficiency from 36% to 47%, although cathode overpotential was fluctuated. Due to different current densities, the maximum H2O2 production rate was 141 mg H2O2/L-h in the MEC fed with acetate medium, but it became low at 6 mg H2O2/L-h in the MEC fed with the wastewater. Our study clearly indicates that improving anodic current density and mitigating membrane fouling would be key parameters for large-scale H2O2-MECs. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Neudorf, Kara D; Huang, Yan Nan; Ragush, Colin M; Yost, Christopher K; Jamieson, Rob C; Truelstrup Hansen, Lisbeth
2017-11-15
Domestic wastewater discharges may adversely impact arctic ecosystems and local indigenous people, who rely on being able to hunt and harvest food from their local environment. Therefore, there is a need to develop efficient wastewater treatment plants (WWTPs), which can be operated in remote communities under extreme climatic conditions. WWTPs have been identified as reservoirs of antibiotic resistance genes (ARGs). The objective of this work was to quantify the presence of nine different ARG markers (int1, sul1, sul2, tet(O), erm(B), mecA, bla CTX-M , bla TEM , and qnr(S)) in two passive systems (waste stabilization ponds [WSPs]) and one mechanical filtration plant operating in two smaller and one large community, respectively, in Nunavut, Canada. Measurement of water quality parameters (carbonaceous oxygen demand, ammonia, total suspended solids, Escherichia coli and total coliforms) showed that the WWTPs provided only primary treatment. Low levels of the ARGs (2logcopies/mL) were observed in the effluent, demonstrating that bacteria residing in three northern WWTPs harbour ARGs conferring resistance to multiple clinically-relevant classes of antibiotics. Our results indicate that long-term storage in WSPs benefitted removal of organic material and some ARGs. However, one WSP system showed evidence of the enrichment of sul1, sul2, mecA, tet(O) and qnr(S). Further research is needed to fully understand if these ARG releases pose a risk to human health, especially in the context of traditional hunting and fishing activities. Copyright © 2017 Elsevier B.V. All rights reserved.
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Papadopoulos, Panagiotis; Papadopoulos, Theofilos; Angelidis, Apostolos S; Boukouvala, Evridiki; Zdragas, Antonios; Papa, Anna; Hadjichristodoulou, Christos; Sergelidis, Daniel
2018-02-01
The objective of this study was to estimate the prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in the production chain of dairy products. Of 367 tested samples (36 bulk tank milk (BTM), 19 dairy products, 72 human, 185 animal, 55 equipment), 212 (57.8%) were found positive for S. aureus. Almost all isolates (99.6%) were resistant to at least one antimicrobial and 13.3% were multi-drug resistant (MDR), exhibiting resistance to three or more antibiotic classes. Eleven samples (3%) were found contaminated by MRSA carrying the mecA gene. None of the MRSA isolates carried the mecC or the Pandon-Valentine leucocidin (PVL) genes. Four spa types were identified among the MRSA isolates: t127, t3586, t1773, t4038, with t127 being the most prevalent (7 out of 11). Two of them, t3586 and t1773, were isolated for the first time in Greece. Furthermore, Pulse-Field Gel Electrophoresis (PFGE) analysis indicated clonal circulation through the dairy production chain. The presence of MDR S. aureus, and especially MRSA, in animals and dairy products represents a potential threat for the spread of this pathogen in the community. The results indicated that human, animal and environmental sources could be involved in the contamination of dairy products along their production chain and therefore further investigation of contamination sources is needed to control the dispersion of MRSA in the community. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Warthin Tumor-Like Mucoepidermoid Carcinoma.
Heatley, Nicholas; Harrington, Kevin J; Thway, Khin
2018-02-01
Mucoepidermoid carcinoma (MEC) shows a wide morphologic spectrum, including epithelium with oncocytic or squamous metaplastic changes overlying a prominent cystic architecture, as well as tumor-associated lymphoid tissue. We illustrate a case of MEC of the parotid in a 17-year-old female, in which all these features occurred extensively, such that they accounted for almost the entire neoplasm, and closely mimicked Warthin tumor histologically. This highlights the need for diagnostic awareness of this particular morphologic variant of MEC, as patients could potentially be inappropriately discharged from follow-up if diagnosed with a benign neoplasm.
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Crops Models for Varying Environmental Conditions
NASA Technical Reports Server (NTRS)
Jones, Harry; Cavazzoni, James; Keas, Paul
2001-01-01
New variable environment Modified Energy Cascade (MEC) crop models were developed for all the Advanced Life Support (ALS) candidate crops and implemented in SIMULINK. The MEC models are based on the Volk, Bugbee, and Wheeler Energy Cascade (EC) model and are derived from more recent Top-Level Energy Cascade (TLEC) models. The MEC models simulate crop plant responses to day-to-day changes in photosynthetic photon flux, photoperiod, carbon dioxide level, temperature, and relative humidity. The original EC model allows changes in light energy but uses a less accurate linear approximation. The simulation outputs of the new MEC models for constant nominal environmental conditions are very similar to those of earlier EC models that use parameters produced by the TLEC models. There are a few differences. The new MEC models allow setting the time for seed emergence, have realistic exponential canopy growth, and have corrected harvest dates for potato and tomato. The new MEC models indicate that the maximum edible biomass per meter squared per day is produced at the maximum allowed carbon dioxide level, the nominal temperatures, and the maximum light input. Reducing the carbon dioxide level from the maximum to the minimum allowed in the model reduces crop production significantly. Increasing temperature decreases production more than it decreases the time to harvest, so productivity in edible biomass per meter squared per day is greater at nominal than maximum temperatures, The productivity in edible biomass per meter squared per day is greatest at the maximum light energy input allowed in the model, but the edible biomass produced per light energy input unit is lower than at nominal light levels. Reducing light levels increases light and power use efficiency. The MEC models suggest we can adjust the light energy day-to- day to accommodate power shortages or Lise excess power while monitoring and controlling edible biomass production.
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NASA Astrophysics Data System (ADS)
DuVal, C.; Trembanis, A. C.; Miller, J. K.; Carton, G.
2016-12-01
Munitions and Explosives of Concern (MEC) have been acknowledged globally as a topic of concern. Increasing use of coastal and continental shelf environments for renewable energy development and other activities has and continues to place humans in contact with legacy military munitions. The Bureau of Ocean Energy Management (BOEM) recognized the need to develop guidance concerning methods for MEC detection in the case of offshore energy development. The study was designed to identify the most likely MEC to be encountered in the Atlantic Outer Continental Shelf (OCS) Wind Energy Areas (WEA), review available technologies and develop a process for selecting appropriate technologies and methodologies for their detection. The process for selecting and optimizing technologies and methods for detection of MEC in BOEM OCS WEAs was developed and tested through the synthesis of historical research, physical site characterization, remote sensing technology review, and in-field trials. To test the selected approach, designated personnel were tasked with seeding a portion of the Delaware WEA with munitions surrogates, while a second group of researchers not privy to the surrogate locations, tested and optimized the selected methodology. The effectiveness of a methodology will be related to ease of detection and other associated parameters. The approach for the in-field trial consists of a combination of wide-area assessment surveying by vessel mounted 230/550 kHz Edgetech 6205 Phase Measuring sonar and near-seafloor surveying using a Teledyne Gavia autonomous underwater vehicle (AUV) equipped with high-resolution 900/1800 kHz Marine Sonics side-scan sonar, Geometrics G880-AUV cesium-vapor magnetometer, and 2 megapixel Point Grey color camera. Survey parameters (e.g. track-line spacing, coverage overlap, AUV altitude) were varied to determine the optimal survey methods, as well as simulate MEC burial to test magnetometer range performance. Preliminary results indicate the combination of high-resolution, near-bed side-scan sonar and magnetometry yields promising results for MEC identification, addressing the potential for both surficial and buried MEC.
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Palli, Swetha Rao; Grabner, Michael; Quimbo, Ralph A; Rugo, Hope S
2015-01-01
To determine the incidence of chemotherapy-induced nausea/vomiting (CINV) and chemotherapy treatment delay and adherence among patients receiving palonosetron versus other 5-hydroxytryptamine receptor antagonist (5-HT3 RA) antiemetics. This retrospective claims analysis included adults with primary malignancies who initiated treatment consisting of single-day intravenous highly emetogenic chemotherapy (HEC) or moderately EC (MEC) regimens. Treatment delay was defined as a gap in treatment at least twice the National Comprehensive Cancer Network-specified cycle length, specific to each chemotherapy regimen. Treatment adherence was determined by the percentage of patients who received the regimen-specific recommended number of chemotherapy cycles within the recommended time frame. We identified 1,832 palonosetron and 2,387 other 5-HT3 RA ("other") patients who initiated HEC therapy, and 1,350 palonosetron users and 1,379 patients on other antiemetics who initiated MEC therapy. Fewer patients receiving palonosetron experienced CINV versus other (HEC, 27.5% versus 32.2%, P=0.0011; MEC, 36.1% versus 41.7%, P=0.0026), and fewer treatment delays occurred among patients receiving palonosetron versus other (HEC, 3.2% versus 6.0%, P<0.0001; MEC, 17.0% versus 26.8%, P<0.0001). Compared with the other cohort, patients receiving palonosetron were significantly more adherent to the index chemotherapy regimen with respect to the recommended time frame (HEC, 74.7% versus 69.7%, P=0.0004; MEC, 43.1% versus 37.3%, P=0.0019) and dosage (HEC, 27.3% versus 25.8%, P=0.0004; MEC, 15.0% versus 12.6%, P=0.0019). Palonosetron more effectively reduced occurrence of CINV in patients receiving HEC or MEC compared with other agents in this real-world setting. Additionally, patients receiving palonosetron had better adherence and fewer treatment delays than patients receiving other 5-HT3 RAs.
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Palli, Swetha Rao; Grabner, Michael; Quimbo, Ralph A; Rugo, Hope S
2015-01-01
Purpose To determine the incidence of chemotherapy-induced nausea/vomiting (CINV) and chemotherapy treatment delay and adherence among patients receiving palonosetron versus other 5-hydroxytryptamine receptor antagonist (5-HT3 RA) antiemetics. Materials and methods This retrospective claims analysis included adults with primary malignancies who initiated treatment consisting of single-day intravenous highly emetogenic chemotherapy (HEC) or moderately EC (MEC) regimens. Treatment delay was defined as a gap in treatment at least twice the National Comprehensive Cancer Network-specified cycle length, specific to each chemotherapy regimen. Treatment adherence was determined by the percentage of patients who received the regimen-specific recommended number of chemotherapy cycles within the recommended time frame. Results We identified 1,832 palonosetron and 2,387 other 5-HT3 RA (“other”) patients who initiated HEC therapy, and 1,350 palonosetron users and 1,379 patients on other antiemetics who initiated MEC therapy. Fewer patients receiving palonosetron experienced CINV versus other (HEC, 27.5% versus 32.2%, P=0.0011; MEC, 36.1% versus 41.7%, P=0.0026), and fewer treatment delays occurred among patients receiving palonosetron versus other (HEC, 3.2% versus 6.0%, P<0.0001; MEC, 17.0% versus 26.8%, P<0.0001). Compared with the other cohort, patients receiving palonosetron were significantly more adherent to the index chemotherapy regimen with respect to the recommended time frame (HEC, 74.7% versus 69.7%, P=0.0004; MEC, 43.1% versus 37.3%, P=0.0019) and dosage (HEC, 27.3% versus 25.8%, P=0.0004; MEC, 15.0% versus 12.6%, P=0.0019). Conclusion Palonosetron more effectively reduced occurrence of CINV in patients receiving HEC or MEC compared with other agents in this real-world setting. Additionally, patients receiving palonosetron had better adherence and fewer treatment delays than patients receiving other 5-HT3 RAs. PMID:26124681
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Raftopoulos, Harry; Boccia, Ralph; Cooper, William; O'Boyle, Erin; Gralla, Richard J
2015-09-01
APF530 is a novel sustained-release formulation of granisetron. In a Phase III trial, APF530 500 mg was noninferior to palonosetron 0.25 mg in preventing acute chemotherapy-induced nausea and vomiting (CINV) after moderately (MEC) or highly emetogenic chemotherapy (HEC) and delayed CINV after MEC, but not superior in preventing delayed CINV after HEC. Emetogenicity was classified by Hesketh criteria; this reanalysis uses newer American Society of Clinical Oncology criteria. Complete responses (no emesis or rescue medication) after cycle one were reanalyzed after reclassification of MEC and HEC by American Society of Clinical Oncology criteria. APF530 maintained noninferiority to palonosetron. Single-dose APF530 is a promising alternative to palonosetron for preventing acute and delayed CINV after MEC or HEC. The Clinicaltrials.gov identifier for this study is NCT00343460.
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Wild rodents and shrews are natural hosts of Staphylococcus aureus.
Mrochen, Daniel M; Schulz, Daniel; Fischer, Stefan; Jeske, Kathrin; El Gohary, Heba; Reil, Daniela; Imholt, Christian; Trübe, Patricia; Suchomel, Josef; Tricaud, Emilie; Jacob, Jens; Heroldová, Marta; Bröker, Barbara M; Strommenger, Birgit; Walther, Birgit; Ulrich, Rainer G; Holtfreter, Silva
2017-09-22
Laboratory mice are the most commonly used animal model for Staphylococcus aureus infection studies. We have previously shown that laboratory mice from global vendors are frequently colonized with S. aureus. Laboratory mice originate from wild house mice. Hence, we investigated whether wild rodents, including house mice, as well as shrews are naturally colonized with S. aureus and whether S. aureus adapts to the wild animal host. 295 animals of ten different species were caught in different locations over four years (2012-2015) in Germany, France and the Czech Republic. 45 animals were positive for S. aureus (15.3%). Three animals were co-colonized with two different isolates, resulting in 48 S. aureus isolates in total. Positive animals were found in Germany and the Czech Republic in each studied year. The S. aureus isolates belonged to ten different spa types, which grouped into six lineages (clonal complex (CC) 49, CC88, CC130, CC1956, sequence type (ST) 890, ST3033). CC49 isolates were most abundant (17/48, 35.4%), followed by CC1956 (14/48, 29.2%) and ST890 (9/48, 18.8%). The wild animal isolates lacked certain properties that are common among human isolates, e.g., a phage-encoded immune evasion cluster, superantigen genes on mobile genetic elements and antibiotic resistance genes, which suggests long-term adaptation to the wild animal host. One CC130 isolate contained the mecC gene, implying wild rodents might be both reservoir and vector for methicillin-resistant S. aureus. In conclusion, we demonstrated that wild rodents and shrews are naturally colonized with S. aureus, and that those S. aureus isolates show signs of host adaptation. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.
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Chadwick, Sean G.; Prasad, Aditya; Smith, W. Lamar; Mordechai, Eli; Adelson, Martin E.
2013-01-01
In recent years, the dramatic increase in community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections has become a significant health care challenge. Early detection of CA-MRSA is important because of its increased virulence associated with the arginine catabolic mobile element (ACME), Panton-Valentine leukocidin (PVL), and other toxins that may contribute to disease severity. In particular, the USA300 epidemic clone has emerged and now represents the cause of as much as 98% of CA-MRSA skin and soft tissue infections in the United States. Current diagnostic assays used to identify CA-MRSA strains are based on complex multiplex PCRs targeting the staphylococcal cassette chromosome mec (SCCmec) DNA junction, a multitude of genes, and noncoding DNA fragments or on a number of lengthy sequence-typing methods. Here, two nucleotide polymorphisms, G88A and G2047A, that were found to be in strict linkage disequilibrium in the S. aureus penicillin-binding protein 3 (pbp3) gene were also found to be highly associated with the USA300 clone of CA-MRSA. Clinical isolates that contained this pbp3 allele were also positive for the presence of SCCmec type IV, the ACME, and the PVL toxin gene and matched the t008 or t121 molecular spa types, which are associated specifically with the USA300 CA-MRSA clone. A single allele-specific PCR targeting the G88A polymorphism was developed and was found to be 100% sensitive and specific for the detection of USA300 CA-MRSA and 91.5% sensitive and 100% specific for the detection of all CA-MRSA isolates in this study. PMID:23698534
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Meyer, Wibke; Reich, Margrit; Beier, Silvio; Behrendt, Joachim; Gulyas, Holger; Otterpohl, Ralf
2016-08-01
This study evaluated the impact of secondary municipal effluent discharge on carbamazepine, diclofenac, and metoprolol concentrations in small and medium rivers in northern Germany and compared the measured environmental concentrations (MECs) to the predicted environmental concentrations (PECs) calculated with four well-established models. During a 1-year sampling period, secondary effluent grab samples were collected at four wastewater treatment plants (WWTPs) together with grab samples from the receiving waters upstream and downstream from the wastewater discharge points. The carbamazepine, diclofenac, and metoprolol concentrations were analyzed with high-performance liquid chromatography-tandem mass spectrometry (HPLC/MS-MS) after solid phase extraction. In the secondary effluents, 84-790 ng/L carbamazepine, 395-2100 ng/L diclofenac, and 745-5000 ng/L metoprolol were detected. The carbamazepine, diclofenac, and metoprolol concentrations analyzed in the rivers downstream from the secondary effluent discharge sites ranged from <5 to 68, 370, and 520 ng/L, respectively. Most of the downstream pharmaceutical concentrations were markedly higher than the corresponding upstream concentrations. The impact of wastewater discharge on the MECs in rivers downstream from the WWTPs was clearly demonstrated, but the correlations of the MECs with dilution factors were poor. The smallest rivers exhibited the largest maximum MECs and the widest ranges of MECs downstream from the wastewater discharge point. Three of the four tested models were conservative, as they showed higher PECs than the MECs in the rivers downstream from the WWTPs. However, the most detailed model underestimated the diclofenac concentrations.
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Lakade, Sameer S; Borrull, Francesc; Furton, Kenneth G; Kabir, Abuzar; Marcé, Rosa Maria; Fontanals, Núria
2018-05-01
A novel sample preparation technique named capsule phase microextraction (CPME) is presented here. The technique utilizes a miniaturized microextraction capsule (MEC) as the extraction medium. The MEC consists of two conjoined porous tubular polypropylene membranes, one of which encapsulates the sorbent through sol-gel technology, while the other encapsulates a magnetic metal rod. As such, MEC integrates both the extraction and stirring mechanisms into a single device. The aim of this article is to demonstrate the application potential of CPME as sample preparation technique for the extraction of a group of personal care products (PCPs) from water matrices. Among the different sol-gel sorbent materials (UCON ® , poly(caprolactone-dimethylsiloxane-caprolactone) (PCAP-DMS-CAP) and Carbowax 20M (CW-20M)) evaluated, CW-20M MEC demonstrated the best extraction performance for the selected PCPs. The extraction conditions for sol-gel CW-20M MEC were optimized, including sample pH, stirring speed, addition of salt, extraction time, sample volume, liquid desorption solvent, and time. Under the optimal conditions, sol-gel CW-20M MEC provided recoveries, ranging between 47 and 90% for all analytes, except for ethylparaben, which showed a recovery of 26%. The method based on CPME with sol-gel CW-20M followed by liquid chromatography-tandem mass spectrometry was developed and validated for the extraction of PCPs from river water and effluent wastewater samples. When analyzing different environmental samples, some analytes such as 2,4-dihydroxybenzophenone, 2,2-dihydroxy-4-4 methoxybenzophenone and 3-benzophenone were found at low ng L -1 .
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Paterson, Gavin K; Harrison, Ewan M; Craven, Emily F; Petersen, Andreas; Larsen, Anders Rhod; Ellington, Matthew J; Török, M Estée; Peacock, Sharon J; Parkhill, Julian; Zadoks, Ruth N; Holmes, Mark A
2013-01-01
We sought to determine the prevalence of nasal colonisation with methicillin-resistant Staphylococcus aureus among cattle veterinarians in the UK. There was particular interest in examining the frequency of colonisation with MRSA harbouring mecC, as strains with this mecA homologue were originally identified in bovine milk and may represent a zoonotic risk to those in contact with dairy livestock. Three hundred and seven delegates at the British Cattle Veterinarian Association (BCVA) Congress 2011 in Southport, UK were screening for nasal colonisation with MRSA. Isolates were characterised by whole genome sequencing and antimicrobial susceptibility testing. Eight out of three hundred and seven delegates (2.6%) were positive for nasal colonisation with MRSA. All strains were positive for mecA and none possessed mecC. The time since a delegate's last visit to a farm was significantly shorter in the MRSA-positive group than in MRSA-negative counterparts. BCVA delegates have an increased risk of MRSA colonisation compared to the general population but their frequency of colonisation is lower than that reported from other types of veterinarian conference, and from that seen in human healthcare workers. The results indicate that recent visitation to a farm is a risk factor for MRSA colonisation and that mecC-MRSA are rare among BCVA delegates (<1% based on sample size). Contact with livestock, including dairy cattle, may still be a risk factor for human colonisation with mecC-MRSA but occurs at a rate below the lower limit of detection available in this study.
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Shahid, Muhammad; Gao, Jian; Zhou, Yanan; Liu, Gang; Ali, Tariq; Deng, Youtian; Sabir, Naveed; Su, Jingliang; Han, Bo
2017-01-01
Bovine protothecal mastitis results in considerable economic losses worldwide. However, Prototheca zopfii induced morphological alterations and oxidative stress in bovine mammary epithelial cells (bMECs) is not comprehensively studied yet. Therefore, the aim of this current study was to investigate the P. zopfii induced pathomorphological changes, oxidative stress and apoptosis in bMECs. Oxidative stress was assessed by evaluating catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA) contents and lactate dehydrogenase (LDH) activity, while ROS generation and apoptosis was measured by confocal laser scanning microscopy. The results revealed that infection of P. zopfii genotype II (GTII) significantly changed bMECs morphology, increased apoptotic rate and MDA contents at 12 h (p < 0.05) and 24 h (p < 0.01) in comparison with control group, in time-dependent manner. LDH activity and ROS generation was also increased (p < 0.01) at 12 h and 24 h. However, SOD and CAT contents in bMECs infected with GTII were decreased (p < 0.05) at 12 h, while GPx (p < 0.01), SOD (p < 0.05) and CAT (p < 0.01) levels were reduced at 24 h. In case of GTI, only CAT and GPx activities were significantly decreased when the duration prolonged to 24 h but lesser than GTII. This suggested that GTII has more devastating pathogenic effects in bMECs, and the findings of this study concluded that GTII induced apoptosis and oxidative stress in bMECs via the imbalance of oxidant and antioxidant defenses as well as the production of intracellular ROS. PMID:28404882
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Shahid, Muhammad; Gao, Jian; Zhou, Yanan; Liu, Gang; Ali, Tariq; Deng, Youtian; Sabir, Naveed; Su, Jingliang; Han, Bo
2017-05-09
Bovine protothecal mastitis results in considerable economic losses worldwide. However, Prototheca zopfii induced morphological alterations and oxidative stress in bovine mammary epithelial cells (bMECs) is not comprehensively studied yet. Therefore, the aim of this current study was to investigate the P. zopfii induced pathomorphological changes, oxidative stress and apoptosis in bMECs. Oxidative stress was assessed by evaluating catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA) contents and lactate dehydrogenase (LDH) activity, while ROS generation and apoptosis was measured by confocal laser scanning microscopy. The results revealed that infection of P. zopfii genotype II (GTII) significantly changed bMECs morphology, increased apoptotic rate and MDA contents at 12 h (p < 0.05) and 24 h (p < 0.01) in comparison with control group, in time-dependent manner. LDH activity and ROS generation was also increased (p < 0.01) at 12 h and 24 h. However, SOD and CAT contents in bMECs infected with GTII were decreased (p < 0.05) at 12 h, while GPx (p < 0.01), SOD (p < 0.05) and CAT (p < 0.01) levels were reduced at 24 h. In case of GTI, only CAT and GPx activities were significantly decreased when the duration prolonged to 24 h but lesser than GTII. This suggested that GTII has more devastating pathogenic effects in bMECs, and the findings of this study concluded that GTII induced apoptosis and oxidative stress in bMECs via the imbalance of oxidant and antioxidant defenses as well as the production of intracellular ROS.
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Pu, WanXia; Su, Yang; Li, JianXi; Li, ChunHui; Yang, ZhiQiang; Deng, HaiPing; Ni, ChunXia
2014-01-01
Staphylococcus aureus is a main cause of bovine mastitis and a major pathogen affecting human health. The emergence and spread of methicillin-resistant Staphylococcus aureus (MRSA) has become a significant concern for both animal health and public health. This study investigated the incidence of MRSA in milk samples collected from dairy cows with clinical mastitis and characterized the MRSA isolates using antimicrobial susceptibility tests and genetic typing methods. In total, 103 S. aureus isolates were obtained from dairy farms in 4 different provinces in China, including Gansu, Shanghai, Sichuan, and Guizhou. Antimicrobial susceptibility testing of these isolates revealed that the resistance rates to penicillin and sulfamethoxazole were high, while the resistance rates to ciprofloxacin and vancomycin were low. Among the 103 isolates, 49 (47.6%) were found to be mecA-positive, indicating the high incidence of MRSA. However, 37 of the 49 mecA-positive isolates were susceptible to oxacillin as determined by antimicrobial susceptibility assays and were thus classified as oxacillin-susceptible mecA-positive S. aureus (OS-MRSA). These isolates could be misclassified as methicillin susceptible Staphylococcus aureus (MSSA) if genetic detection of mecA was not performed. Molecular characterization of selected mecA-positive isolates showed that they were all negative with Panton-Valentine leukocidin (PVL), but belonged to different spa types and SCCmec types. These results indicate that OS-MRSA is common in bovine mastitis in China and underscore the need for genetic methods (in addition to phenotypic tests) to accurately identify MRSA.
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Perl, Alexander E; Kasner, Margaret T; Tsai, Donald E; Vogl, Dan T; Loren, Alison W; Schuster, Stephen J; Porter, David L; Stadtmauer, Edward A; Goldstein, Steven C; Frey, Noelle V; Nasta, Sunita D; Hexner, Elizabeth O; Dierov, Jamil K; Swider, Cezary R; Bagg, Adam; Gewirtz, Alan M; Carroll, Martin; Luger, Selina M
2009-11-01
Inhibiting mammalian target of rapamycin (mTOR) signaling in acute myelogenous leukemia (AML) blasts and leukemic stem cells may enhance their sensitivity to cytotoxic agents. We sought to determine the safety and describe the toxicity of this approach by adding the mTOR inhibitor, sirolimus (rapamycin), to intensive AML induction chemotherapy. We performed a phase I dose escalation study of sirolimus with the chemotherapy regimen MEC (mitoxantrone, etoposide, and cytarabine) in patients with relapsed, refractory, or untreated secondary AML. Twenty-nine subjects received sirolimus and MEC across five dose levels. Dose-limiting toxicities were irreversible marrow aplasia and multiorgan failure. The maximum tolerated dose (MTD) of sirolimus was determined to be a 12 mg loading dose on day 1 followed by 4 mg/d on days 2 to 7, concurrent with MEC chemotherapy. Complete or partial remissions occurred in 6 (22%) of the 27 subjects who completed chemotherapy, including 3 (25%) of the 12 subjects treated at the MTD. At the MTD, measured rapamycin trough levels were within the therapeutic range for solid organ transplantation. However, direct measurement of the mTOR target p70 S6 kinase phosphorylation in marrow blasts from these subjects only showed definite target inhibition in one of five evaluable samples. Sirolimus and MEC is an active and feasible regimen. However, as administered in this study, the synergy between MEC and sirolimus was not confirmed. Future studies are planned with different schedules to clarify the clinical and biochemical effects of sirolimus in AML and to determine whether target inhibition predicts chemotherapy response.
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Sukhanova, Maria V; D'Herin, Claudine; Boiteux, Serge; Lavrik, Olga I
2014-10-01
To characterize proteins that interact with single-stranded/double-stranded (ss/ds) DNA junctions in whole cell free extracts of Saccharomyces cerevisiae, we used [(32)P]-labeled photoreactive partial DNA duplexes containing a 3'-ss/ds-junction (3'-junction) or a 5'-ss/ds-junction (5'-junction). Identification of labeled proteins was achieved by MALDI-TOF mass spectrometry peptide mass fingerprinting and genetic analysis. In wild-type extract, one of the components of the Ddc1-Rad17-Mec3 complex, Ddc1, was found to be preferentially photocrosslinked at a 3'-junction. On the other hand, RPAp70, the large subunit of the replication protein A (RPA), was the predominant crosslinking product at a 5'-junction. Interestingly, ddc1Δ extracts did not display photocrosslinking of RPAp70 at a 5'-junction. The results show that RPAp70 crosslinked to DNA with a 5'-junction is subject to limited proteolysis in ddc1Δ extracts, whereas it is stable in WT, rad17Δ, mec3Δ and mec1Δ extracts. The degradation of the RPAp70-DNA adduct in ddc1Δ extract is strongly reduced in the presence of the proteasome inhibitor MG 132. We also addressed the question of the stability of free RPA, using anti-RPA antibodies. The results show that RPAp70 is also subject to proteolysis without photocrosslinking to DNA upon incubation in ddc1Δ extract. The data point to a novel property of Ddc1, modulating the turnover of DNA binding proteins such as RPAp70 by the proteasome. Copyright © 2014 Elsevier B.V. All rights reserved.
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Kaspar, Ursula; de Haro Sautto, Jorge A; Molinaro, Sonja; Peters, Georg; Idelevich, Evgeny A; Becker, Karsten
2018-05-07
Livestock-associated, methicillin-resistant Staphylococcus aureus (LA-MRSA) are increasingly migrating from livestock into human and animal health care settings. Alternative substances are wanted to overcome the drawbacks of currently available drugs applied for MRSA eradication. The recombinant bacteriophage endolysin HY-133 has been proven as an active agent against S. aureus Here, the in vitro activity of HY-133 was studied against a large collection of genetically diverse LA-MRSA revealing its high activity against mecA, mecB and mecC LA-MRSA. Copyright © 2018 American Society for Microbiology.
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Use of an emergency medical pictorial communication book during simulated disaster conditions.
Behar, Solomon; Benson Ii, Richard; Kurzweil, Ami; Azen, Colleen; Nager, Alan L
2013-10-01
During disasters, the needs of victims outstrip available resources. Rapid assessment of patients must be performed; however, language barriers can be an impediment to efficient patient assessment, especially if interpreter resources are limited. Dependency on interpretive services requiring technology such a telephones, cell phones, and video conferencing may be inefficient, as they may be unavailable during disaster conditions. A low-tech, portable tool that aids in communication with non-English speakers would be beneficial. The medical emergency communication (MEC) book, developed at Children's Hospital Los Angeles, has the potential to be a useful tool in this capacity. The goal of this pilot study was to compare the accuracy of a newly developed disaster-focused medical history obtained from Spanish-speaking patients or caregivers using the MEC book, compared to a control group with whom no book was used. Our hypothesis was that use of the MEC book improves accuracy of medical history taking between English-only speaking health care workers and Spanish-speaking patients better than a monolingual clinician trying to take a medical history without it. We anticipated a higher overall score in the group of subjects whose histories were taken using the MEC book than in the control group. Patient satisfaction with the MEC book also was measured.
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Bjornson, Marta; Balcke, Gerd Ulrich; Xiao, Yanmei; de Souza, Amancio; Wang, Jin-Zheng; Zhabinskaya, Dina; Tagkopoulos, Ilias; Tissier, Alain; Dehesh, Katayoon
2017-07-01
To maintain homeostasis in the face of intrinsic and extrinsic insults, cells have evolved elaborate quality control networks to resolve damage at multiple levels. Interorganellar communication is a key requirement for this maintenance, however the underlying mechanisms of this communication have remained an enigma. Here we integrate the outcome of transcriptomic, proteomic, and metabolomics analyses of genotypes including ceh1, a mutant with constitutively elevated levels of both the stress-specific plastidial retrograde signaling metabolite methyl-erythritol cyclodiphosphate (MEcPP) and the defense hormone salicylic acid (SA), as well as the high MEcPP but SA deficient genotype ceh1/eds16, along with corresponding controls. Integration of multi-omic analyses enabled us to delineate the function of MEcPP from SA, and expose the compartmentalized role of this retrograde signaling metabolite in induction of distinct but interdependent signaling cascades instrumental in adaptive responses. Specifically, here we identify strata of MEcPP-sensitive stress-response cascades, among which we focus on selected pathways including organelle-specific regulation of jasmonate biosynthesis; simultaneous induction of synthesis and breakdown of SA; and MEcPP-mediated alteration of cellular redox status in particular glutathione redox balance. Collectively, these integrated multi-omic analyses provided a vehicle to gain an in-depth knowledge of genome-metabolism interactions, and to further probe the extent of these interactions and delineate their functional contributions. Through this approach we were able to pinpoint stress-mediated transcriptional and metabolic signatures and identify the downstream processes modulated by the independent or overlapping functions of MEcPP and SA in adaptive responses. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.
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2014-01-01
The methylation of cytosine to 5-methylcytosine (5-meC) is an important epigenetic DNA modification in many bacteria, plants, and mammals, but its relevance for important model organisms, including Caenorhabditis elegans and Drosophila melanogaster, is still equivocal. By reporting the presence of 5-meC in a broad variety of wild, laboratory, and industrial yeasts, a recent study also challenged the dogma about the absence of DNA methylation in yeast species. We would like to bring to attention that the protocol used for gas chromatography/mass spectrometry involved hydrolysis of the DNA preparations. As this process separates cytosine and 5-meC from the sugar phosphate backbone, this method is unable to distinguish DNA- from RNA-derived 5-meC. We employed an alternative LC–MS/MS protocol where by targeting 5-methyldeoxycytidine moieties after enzymatic digestion, only 5-meC specifically derived from DNA is quantified. This technique unambiguously identified cytosine DNA methylation in Arabidopsis thaliana (14.0% of cytosines methylated), Mus musculus (7.6%), and Escherichia coli (2.3%). Despite achieving a detection limit at 250 attomoles (corresponding to <0.00002 methylated cytosines per nonmethylated cytosine), we could not confirm any cytosine DNA methylation in laboratory and industrial strains of Saccharomyces cerevisiae, Schizosaccharomyces pombe, Saccharomyces boulardii, Saccharomyces paradoxus, or Pichia pastoris. The protocol however unequivocally confirmed DNA methylation in adult Drosophila melanogaster at a value (0.034%) that is up to 2 orders of magnitude below the detection limit of bisulphite sequencing. Thus, 5-meC is a rare DNA modification in drosophila but absent in yeast. PMID:24640988
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Harastani, Houda H.; Tokajian, Sima T.
2014-01-01
Background The emergence of community-associated methicillin resistant Staphylococcus aureus (CA-MRSA) has caused a change in MRSA epidemiology worldwide. In the Middle East, the persistent spread of CA-MRSA isolates that were associated with multilocus sequence type (MLST) clonal complex 80 and with staphylococcal cassette chromosome mec (SCCmec) type IV (CC80-MRSA-IV), calls for novel approaches for infection control that would limit its spread. Methodology/Principal Findings In this study, the epidemiology of CC80-MRSA-IV was investigated in Jordan and Lebanon retrospectively covering the period from 2000 to 2011. Ninety-four S. aureus isolates, 63 (67%) collected from Lebanon and 31 (33%) collected from Jordan were included in this study. More than half of the isolates (56%) were associated with skin and soft tissue infections (SSTIs), and 73 (78%) were Panton-Valentine Leukocidin (PVL) positive. Majority of the isolates (84%) carried the gene for exofoliative toxin d (etd), 19% had the Toxic Shock Syndrome Toxin-1 gene (tst), and seven isolates from Jordan had a rare combination being positive for both tst and PVL genes. spa typing showed the prevalence of type t044 (85%) and pulsed-field gel electrophoresis (PFGE) recognized 21 different patterns. Antimicrobial susceptibility testing showed the prevalence (36%) of a unique resistant profile, which included resistance to streptomycin, kanamycin, and fusidic acid (SKF profile). Conclusions The genetic diversity among the CC80 isolates observed in this study poses an additional challenge to infection control of CA-MRSA epidemics. CA-MRSA related to ST80 in the Middle East was distinguished in this study from the ones described in other countries. Genetic diversity observed, which may be due to mutations and differences in the antibiotic regimens between countries may have led to the development of heterogeneous strains. Hence, it is difficult to maintain “the European CA-MRSA clone” as a uniform clone and it is better to designate as CC80-MRSA-IV isolates. PMID:25078407
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Singh, Rashmi Bhat; Amare Kadam, Pratibha S
2013-10-01
To investigate the most recurrent deletion loci on 3p12-p26 by deletion mapping studies by PCR-LOH and BAC array-FISH in sporadic conventional renal cell carcinoma (cRCC) and further, to evaluate the their clinicopathologic significance in cRCC. Comparative allelotyping studies in cRCC and major epithelial carcinomas (MEC) such as lung, breast, and bladder tumors were also carried out to investigate the specificity of the targeted loci in cRCC. A total of 40 c-RCC patients were enrolled in this study, categorized in to 2 groups: group I comprises of patients of stages I and II and group II includes patients at stages III and IV. Loss of heterozygosity (LOH) studies were performed by PCR using 15 microsatellite markers of region 3p12-p26 on paired normal-tumor tissues. The recurrent LOH loci found in 27 cRCC tumors were further validated by BAC array-FISH using 23 serially mapped BAC clones. Simultaneously, the allelic deletion status of fragile histidine triad (FHIT) gene was studied by FISH in cRCC and major epithelial carcinoma (MEC) tumors. The numerical aberrations of chromosome 3 were also studied using the centromere enumeration probe (CEP) probe for chromosome 3 to validate the observed allelic losses by BAC array-FISH in cRCC as well as MECs. Our study revealed 3 affected regions of LOH on 3p in cRCC: 3p12.2-p14.1, 3p14.2-p21.1, and 3p24.2-p26.1 in both group I (stages I and II) and group II (stage III and IV). Comparative allelotyping studies revealed that except for LOH loci D3S2406 (20%), D3S1766 (14%), and D3S1560 (20%), remaining affected loci revealed retention of heterozygosity (ROH) in breast carcinomas. Lung and bladder tumors revealed ROH at all affected LOH loci. FISH with FHIT gene probe revealed deletions in cRCC (88%), breast (30%), and lung tumors (10%). FHIT gene deletions frequency was almost equal in both groups I and II (>70%), whereas a locus 3p13 (D3S2454) revealed the highest LOH in group II (83%) patients in comparison to group I (16%). BAC array-FISH studies in cRCC identified 15 recurrent deletion loci at crucial regions, 3p12.2, 3p14.2, 3p21.3, and 3p24.2-p26 with long continuous deletion of 3p14.1-p26.1 exclusively in patients of stages III and IV. Validation of LOH loci in breast carcinomas by BAC array-FISH with BAC clones mapped at these loci revealed comparatively lower deletion frequency for RP11-59E22 (3p12.2) (30%), RP11-759B7(3p21.1) (12%), and RP11-57D6 (3p25.2, proximal to VHL) (15%) than cRCC. Molecular cytogenetic studies by BAC array-FISH was found to be more sensitive over LOH. Deletion patterns on 3p explored that deletion of FHIT and flanking loci may occur as an initiating event followed by deletions at 3p12.2, 3p21.31-3p21.32, and 3p24.2-3p26.1 in the initial stage of development of disease, while continuous large deletions of 3p21.3-3p26.1 and 3p14.1-3p26.1 occur as progressive deletion due to genetic instability. Lack of VHL along with flanking loci in 50% cRCC patients that included both groups I and II supported the hypothesis of both VHL dependent and VHL independent pathways in cRCC tumorigenesis. Comparative allelotyping studies in cRCC and MECs indicated association of specific targeted loci including VHL in cRCC. Further expansion of these studies with characterization of the genes at targeted loci and correlation with clinical outcome will explore the prognostic significance and also provide an insight into the mechanisms of tumor suppressive pathways in genitourinary cancers such as CRCC. Copyright © 2013 Elsevier Inc. All rights reserved.
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Berg, Matthew D; Genereaux, Julie; Karagiannis, Jim; Brandl, Christopher J
2018-05-31
Tra1 is an essential component of the SAGA/SLIK and NuA4 complexes in S. cerevisiae , recruiting these co-activator complexes to specific promoters. As a PIKK family member, Tra1 is characterized by a C-terminal phosphoinositide 3-kinase (PI3K) domain. Unlike other PIKK family members ( e.g. , Tor1, Tor2, Mec1, Tel1), Tra1 has no demonstrable kinase activity. We identified three conserved arginine residues in Tra1 that reside proximal or within the cleft between the N- and C-terminal subdomains of the PI3K domain. To establish a function for Tra1's PI3K domain and specifically the cleft region, we characterized a tra1 allele where these three arginine residues are mutated to glutamine. The half-life of the Tra1[Formula: see text] protein is reduced but its steady state level is maintained at near wild-type levels by a transcriptional feedback mechanism. The tra1 [Formula: see text] allele results in slow growth under stress and alters the expression of genes also regulated by other components of the SAGA complex. Tra1[Formula: see text] is less efficiently transported to the nucleus than the wild-type protein. Likely related to this, Tra1[Formula: see text] associates poorly with SAGA/SLIK and NuA4. The ratio of Spt7 SLIK to Spt7 SAGA increases in the tra1 [Formula: see text] strain and truncated forms of Spt20 become apparent upon isolation of SAGA/SLIK. Intragenic suppressor mutations of tra1 [Formula: see text] map to the cleft region further emphasizing its importance. We propose that the PI3K domain of Tra1 is directly or indirectly important for incorporating Tra1 into SAGA and NuA4 and thus the biosynthesis and/or stability of the intact complexes. Copyright © 2018 Berg et al.
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Cerrillo, Míriam; Viñas, Marc; Bonmatí, August
2016-09-01
The combination of the anaerobic digestion (AD) process with a microbial electrolysis cell (MEC) coupled to an ammonia stripping unit as a post-treatment was assessed both in series operation, to improve the quality of the effluent, and in loop configuration recirculating the effluent, to increase the AD robustness. The MEC allowed maintaining the chemical oxygen demand removal of the whole system of 46±5% despite the AD destabilization after doubling the organic and nitrogen loads, while recovering 40±3% of ammonia. The AD-MEC system, in loop configuration, helped to recover the AD (55% increase in methane productivity) and attained a more stable and robust operation. The microbial population assessment revealed an enhancement of AD methanogenic archaea numbers and a shift in eubacterial population. The AD-MEC combined system is a promising strategy for stabilizing AD against organic and nitrogen overloads, while improving the quality of the effluent and recovering nutrients for their reutilization. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Shehab, Noura A; Ortiz-Medina, Juan F; Katuri, Krishna P; Hari, Ananda Rao; Amy, Gary; Logan, Bruce E; Saikaly, Pascal E
2017-09-01
Applying microbial electrochemical technologies for the treatment of highly saline or thermophilic solutions is challenging due to the lack of proper inocula to enrich for efficient exoelectrogens. Brine pools from three different locations (Valdivia, Atlantis II and Kebrit) in the Red Sea were investigated as potential inocula sources for enriching exoelectrogens in microbial electrolysis cells (MECs) under thermophilic (70°C) and hypersaline (25% salinity) conditions. Of these, only the Valdivia brine pool produced high and consistent current 6.8±2.1A/m 2 -anode in MECs operated at a set anode potential of +0.2V vs. Ag/AgCl (+0.405V vs. standard hydrogen electrode). These results show that exoelectrogens are present in these extreme environments and can be used to startup MEC under thermophilic and hypersaline conditions. Bacteroides was enriched on the anode of the Valdivia MEC, but it was not detected in the open circuit voltage reactor seeded with the Valdivia brine pool. Copyright © 2017 Elsevier Ltd. All rights reserved.
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NASA Astrophysics Data System (ADS)
Mitov, M.; Chorbadzhiyska, E.; Nalbandian, L.; Hubenova, Y.
2017-07-01
The development of cost-effective cathodes, operating at neutral pH and ambient temperatures, is a crucial challenge for the practical application of microbial electrolysis cell (MEC) technology. In this study, NiW and NiMo co-deposits produced by electroplating on Ni-foam are explored as cathodes in MEC. The fabricated electrodes exhibit higher corrosion stability and enhanced electrocatalytic activity towards hydrogen evolution reaction in neutral electrolyte compared to the bare Ni-foam. NiW/Ni-foam electrodes possess six times higher intrinsic catalytic activity, estimated from data obtained by linear voltammetry and chronoamperometry. The newly developed electrodes are applied as cathodes in single-chamber membrane-free MEC reactors, inoculated with wastewater and activated sludge from a municipal wastewater treatment plant. Cathodic hydrogen recovery of 79% and 89% by using NiW and NiMo cathodes, respectively, is achieved at applied voltage of 0.6 V. The obtained results reveal potential for practical application of used catalysts in MEC.
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Milanović, Vesna; Osimani, Andrea; Pasquini, Marina; Aquilanti, Lucia; Garofalo, Cristiana; Taccari, Manuela; Cardinali, Federica; Riolo, Paola; Clementi, Francesca
2016-06-16
This study was aimed at investigating the occurrence of 11 transferable antibiotic resistance (AR) genes [erm(A), erm(B), erm(C), vanA, vanB, tet(M), tet(O), tet(S), tet(K), mecA, blaZ] in 11 species of marketed edible insects (small crickets powder, small crickets, locusts, mealworm larvae, giant waterbugs, black ants, winged termite alates, rhino beetles, mole crickets, silkworm pupae, and black scorpions) in order to provide a first baseline for risk assessment. Among the AR genes under study, tet(K) occurred with the highest frequency, followed by erm(B), tet(S) and blaZ. A high variability was seen among the samples, in terms of occurrence of different AR determinants. Cluster Analysis and Principal Coordinates Analysis allowed the 11 samples to be grouped in two main clusters, one including all but one samples produced in Thailand and the other including those produced in the Netherlands. Copyright © 2016 Elsevier B.V. All rights reserved.
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Targeting Aberrant p70S6K Activation for Estrogen Receptor-Negative Breast Cancer Prevention.
Wang, Xiao; Yao, Jun; Wang, Jinyang; Zhang, Qingling; Brady, Samuel W; Arun, Banu; Seewaldt, Victoria L; Yu, Dihua
2017-11-01
The prevention of estrogen receptor-negative (ER-) breast cancer remains a major challenge in the cancer prevention field, although antiestrogen and aromatase inhibitors have shown adequate efficacy in preventing estrogen receptor-positive (ER + ) breast cancer. Lack of commonly expressed, druggable targets is a major obstacle for meeting this challenge. Previously, we detected the activation of Akt signaling pathway in atypical hyperplasic early-stage lesions of patients. In the current study, we found that Akt and the downstream 70 kDa ribosomal protein S6 kinase (p70S6K) signaling pathway was highly activated in ER - premalignant breast lesions and ER - breast cancer. In addition, p70S6K activation induced transformation of ER - human mammary epithelial cells (hMEC). Therefore, we explored the potential of targeting Akt/p70S6K in the p70S6K activated, ER - hMEC models and mouse mammary tumor models for the prevention of ER - breast cancer. We found that a clinically applicable Akt/p70S6K dual inhibitor, LY2780301, drastically decreased proliferation of hMECs with ErbB2-induced p70S6K activation via Cyclin B1 inhibition and cell-cycle blockade at G 0 -G 1 phase, while it did not significantly reverse the abnormal acinar morphology of these hMECs. In addition, a brief treatment of LY2780301 in MMTV- neu mice that developed atypical hyperplasia (ADH) and mammary intraepithelial neoplasia (MIN) lesions with activated p70S6K was sufficient to suppress S6 phosphorylation and decrease cell proliferation in hyperplasic MECs. In summary, targeting the aberrant Akt/p70S6K activation in ER - hMEC models in vitro and in the MMTV- neu transgenic mouse model in vivo effectively inhibited Akt/S6K signaling and reduced proliferation of hMECs in vitro and ADH/MIN lesions in vivo , indicating its potential in prevention of p70S6K activated ER - breast cancer. Cancer Prev Res; 10(11); 641-50. ©2017 AACR . ©2017 American Association for Cancer Research.
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Liu, Yuan; Xu, Xiaofang; Guan, Sumin; Wu, Junzheng; Liu, Yanpu
2013-01-01
Background Multidrug resistance-related protein 1 (MRP1/ABCC1) and multidrug resistance protein 1 (MDR1/P-glycoprotein/ABCB1) are both membrane-bound drug transporters. In contrast to MDR1, MRP1 also transports glutathione (GSH) and drugs conjugated to GSH. Due to its extraordinary transport properties, MRP1/ABCC1 contributes to several physiological functions and pathophysiological incidents. We previously found that nuclear translocation of MRP1 contributes to multidrug-resistance (MDR) of mucoepidermoid carcinoma (MEC). The present study investigated how MRP1 contributes to MDR in the nuclei of MEC cells. Methods Western blot and RT-PCR was carried out to investigate the change of multidrug-resistance protein 1 (MDR1) in MC3/5FU cells after MRP1 was downregulated through RNA interference (RNAi). Immunohistochemistry (IHC) staining of 127 cases of MEC tissues was scored with the expression index (EI). The EI of MDR1 and MRP1 (or nuclear MRP1) was analyzed with Spearman's rank correlation analysis. Using multiple tumor tissue assays, the location of MRP1 in other tissues was checked by HIC. Luciferase reporter assays of MDR1 promoter was carried out to check the connection between MRP1 and MDR1 promoter. Results MRP1 downregulation led to a decreased MDR1 expression in MC3/5FU cells which was caused by decreased activity of MDR1 promoter. IHC study of 127 cases of MEC tissues demonstrated a strong positive correlation between nuclear MRP1 expression and MDR1 expression. Furthermore, IHC study of multiple tumor tissue array sections showed that although nuclear MRP1 widely existed in MEC tissues, it was not found in normal tissues or other tumor tissues. Conclusions Our findings indicate that nuclear MRP1 contributes to MDR mainly through regulating MDR1 expression in MEC. And the unique location of MRP1 made it an available target in identifying MEC from other tumors. PMID:24013781
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Methicillin resistant S. aureus in human and bovine mastitis.
Holmes, Mark A; Zadoks, Ruth N
2011-12-01
Staphylococcus aureus is a ubiquitous organism that causes a variety of diseases including mastitis in cattle and humans. High-level resistance of S. aureus to β-lactams conferred by a mecA gene encoding a modified penicillin binding protein (PBP2a) was first observed in the early 1960's. These methicillin resistant S. aureus (MRSA) have been responsible for both hospital acquired infections (HA-MRSA) and, more recently, community acquired MRSA (CA-MRSA). A small number of human MRSA mastitis cases and outbreaks in maternity or neonatal units have been reported which are generally the result of CA-MRSA. The establishment of the sequence type 398 (ST398) in farm animals, primarily pigs, in the early 2000's has provided a reservoir of infection for humans and dairy cattle, particularly in continental Europe, described as livestock-associated MRSA (LA-MRSA). Prior to the emergence of ST398 there were sporadic reports of MRSA in bovine milk and cases of mastitis, often caused by strains from human associated lineages. Subsequently, there have been several reports describing bovine udder infections caused by ST-398 MRSA. Recently, another group of LA-MRSA strains was discovered in humans and dairy cattle in Europe. This group carries a divergent mecA gene and includes a number of S. aureus lineages (CC130, ST425, and CC1943) that were hitherto thought to be bovine-specific but are now also found as carriage or clinical isolates in humans. The emergence of MRSA in dairy cattle may be associated with contact with other host species, as in the case of ST398, or with the exchange of genetic material between S. aureus and coagulase negative Staphylococcus species, which are the most common species associated with bovine intramammary infections and commonly carry antimicrobial resistance determinants.
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Raeisi, Javad; Saifi, Mahnaz; Pourshafie, Mohammad Reza; Asadi Karam, Mohammad Reza; Mohajerani, Hamid Reza
2015-01-01
Background: Methicillin-Resistant Staphylococcus aureus (MRSA) is a major pathogen in the hospital and community settings. Rapid methods to diagnose S. aureus infections are sought by many researchers worldwide. The current study aimed to utilize a phenotypic method of turanose fermentation to identify methicillin-susceptible and resistant S. aureus. Objectives: The current study aimed to assay the turanose metabolism at different dilutions as a rapid phenotypic method to identify MRSA isolates. Materials and Methods: A total of 150 Staphylococcus isolates were collected from Tehran health centers. Staphylococcus aureus isolates were identified based on cultural characteristics, biochemical reactions and positive tube coagulase test. Methicillin resistance was determined by the disk diffusion method. The Polymerase Chain Reaction amplification was used to detect the mecA gene in MRSA isolates. All the methicillin-resistant and susceptible isolates were evaluated for turanose metabolism with 1%, 0.7% and 0.5% dilutions using the microplate method. Results: Out of the 150 staphylococcal isolates, 80 were identified as S. aureus. Among which 40 (50%) of the isolates were MRSA. The mecA gene was present in all S. aureus isolates resistant to methicillin. A considerable difference was also observed between susceptible and resistant isolates of S. aureus at a 0.7% dilution of turanose. Conclusions: Since it is highly important to rapidly detect MRSA isolates, especially in nosocomial infections, phenotypic methods may certainly be useful for this purpose. Resistance to methicillin in S. aureus shows a substantially increased ability in turanose metabolism. It is concluded that fermentation of turanose at 0.7% dilution could be a rapid detection method for primary screening of MRSA isolates. PMID:26495105
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Oliveira, C J B; Tiao, N; de Sousa, F G C; de Moura, J F P; Santos Filho, L; Gebreyes, W A
2016-03-01
The aim of this study was to investigate the phenotypic and genotypic diversity and anti-microbial resistance among staphylococci of dairy herds that originated from Paraiba State, north-eastern Brazil, a region where such studies are rare. Milk samples (n = 552) were collected from 15 dairy farms. Isolates were evaluated for anti-microbial susceptibility by Kirby-Bauer disc diffusion method. Confirmation of methicillin-resistant Staphylococcus aureus (MRSA) was performed using multiplex PCR targeting mecA and nuc genes in addition to phenotypic assay based on PBP-2a latex agglutination. Clonal relatedness of isolates was determined by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) genotyping. Staphylococci were detected in 269 (49%) of the samples. Among these, 65 (24%) were S. aureus. The remaining 204 isolates were either coagulase-negative staphylococci (n = 188; 70%) or coagulase positive other than S. aureus (n = 16; 6%). Staphylococci were cultured in seven (35%) of the 20 hand swab samples, from which five isolates were S. aureus. The isolates were most commonly resistant against penicillin (43%), ampicillin (38%) and oxacillin (27%). The gene mecA was detected in 21 S. aureus from milk and in one isolate from a milker's hand. None of the isolates were resistant to vancomycin. PFGE findings showed high clonal diversity among the isolates. Based on MLST, we identified a total of 11 different sequence types (STs 1, 5, 6, 83, 97, 126, 1583, 1622, 1623, 1624 and 1625) with four novel STs (ST1622-ST1625). The findings show that MRSA is prevalent in milk from semi-extensive dairy cows in north-eastern Brazil, and further investigation on its extent in various types of milk production systems and the farm-to-table continuum is warranted. © 2015 Blackwell Verlag GmbH.
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Characterization of tumor cells and stem cells by differential nuclear methylation imaging
NASA Astrophysics Data System (ADS)
Tajbakhsh, Jian; Wawrowsky, Kolja A.; Gertych, Arkadiusz; Bar-Nur, Ori; Vishnevsky, Eugene; Lindsley, Erik H.; Farkas, Daniel L.
2008-02-01
DNA methylation plays a key role in cellular differentiation. Aberrant global methylation patterns are associated with several cancer types, as a result of changes in long-term activation status of up to 50% of genes, including oncogenes and tumor-suppressor genes, which are regulated by methylation and demethylation of promoter region CpG dinucleotides (CpG islands). Furthermore, DNA methylation also occurs in nonisland CpG sites (> 95% of the genome), present once per 80 dinucleotides on average. Nuclear DNA methylation increases during the course of cellular differentiation while cancer cells usually show a net loss in methylation. Given the large dynamic range in DNA methylation load, the methylation pattern of a cell can provide a valuable distinction as to its status during differentiation versus the disease state. By applying immunofluorescence, confocal microscopy and 3D image analysis we assessed the potential of differential nuclear distribution of methylated DNA to be utilized as a biomarker to characterize cells during development and when diseased. There are two major fields that may immediately benefit from this development: (1) the search for factors that contribute to pluripotency and cell fate in human embryonic stem cell expansion and differentiation, and (2) the characterization of tumor cells with regard to their heterogeneity in molecular composition and behavior. We performed topological analysis of the distribution of methylated CpG-sites (MeC) versus heterochromatin. This innovative approach revealed significant differences in colocalization patterns of MeC and heterochromatin-derived signals between undifferentiated and differentiated human embryonic stem cells, as well as untreated AtT20 mouse pituitary tumor cells compared to a subpopulation of these cells treated with 5-azacytidine for 48 hours.
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Chamon, Raiane Cardoso; Iorio, Natalia Lopes Pontes; Cavalcante, Fernanda Sampaio; Teodoro, Cristiane R S; de Oliveira, Ana Paula Chaves; Maia, Fernanda; dos Santos, Kátia Regina Netto
2014-12-01
In this work, the molecular and phenotypic antimicrobial resistance and clonal diversity of 10 linezolid-resistant Staphylococcus spp. isolates were investigated. The 7 Staphylococcus haemolyticus isolates presented Staphylococcal cassete chromosome mec (SCCmec) V and belonged to the same pulsed-field gel electrophoresis pulsotype. Their MICs for oxacillin, vancomycin, and linezolid were ≥ 256 μg/mL, 1-4 μg/mL, and 8-16 μg/mL, respectively. The 3 S. hominis presented MIC values 32 to >256 μg/mL, 2-4 μg/mL, and 12-24 μg/mL, and all carried the nontypeable SCCmec (ccr1 + mecA class) and belonged to 2 different genotypes. The cfr gene was not found, but the mutation G2603T was detected in S. haemolyticus and C2190T and G2603T in Staphylococcus hominis in 23S rRNA. This study demonstrates the spread of a linezolid-resistant S. haemolyticus genotype and, for the first time, describes the mutation C2190T among S. hominis isolates with a double mutation in Brazil. Copyright © 2014 Elsevier Inc. All rights reserved.
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Weisser, Maja; Schoenfelder, Sonja M K; Orasch, Christina; Arber, Caroline; Gratwohl, Alois; Frei, Reno; Eckart, Martin; Flückiger, Ursula; Ziebuhr, Wilma
2010-07-01
We report on a leukemic patient who suffered from a persistent, generalized, and eventually fatal Staphylococcus epidermidis infection during prolonged aplasia. Over a 6-week period, we isolated a genetically and phenotypically unstable S. epidermidis strain related to an epidemic clone associated with hospital infections worldwide. Strikingly, the strain showed a remarkable degree of variability, with evidence of selection and increasing predominance of biofilm-producing and oxacillin-resistant variants over time. Thus, in the early stages of the infection, the strain was found to generate subpopulations which had spontaneously lost the biofilm-mediating ica locus along with the oxacillin resistance-conferring mecA gene. These deletion mutants were obviously outcompeted by the ica- and mecA-positive wild-type genotype, with the selection and predominance of strongly biofilm-forming and oxacillin-resistant variants in the later stages of the infection. Also, a switch from protein- to polysaccharide intercellular adhesin/poly-N-acetylglucosamine (PIA/PNAG)-mediated-biofilm production was detected among ica-positive variants in the course of the infection. The data highlight the impact of distinct S. epidermidis clonal lineages as serious nosocomial pathogens that, through the generation and selection of highly pathogenic variants, may critically determine disease progression and outcome.
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DOE Office of Scientific and Technical Information (OSTI.GOV)
Shott, Gregory
This special analysis (SA) evaluates whether the Materials and Energy Corporation (M&EC) Sealed Source waste stream (PERM000000036, Revision 0) is suitable for shallow land burial (SLB) at the Area 5 Radioactive Waste Management Site (RWMS) on the Nevada National Security Site (NNSS). Disposal of the M&EC Sealed Source waste meets all U.S. Department of Energy (DOE) Manual DOE M 435.1-1, “Radioactive Waste Management Manual,” Chapter IV, Section P performance objectives (DOE 1999). The M&EC Sealed Source waste stream is recommended for acceptance without conditions.
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Consensus sediment quality guidelines for polycyclic aromatic hydrocarbon mixtures
DOE Office of Scientific and Technical Information (OSTI.GOV)
Swartz, R.C.
1999-04-01
Sediment quality guidelines (SQGs) for polycyclic aromatic hydrocarbons (PAHs) have been derived from a variety of laboratory, field, and theoretical foundations. They include the screening level concentration, effects ranges-low and -median, equilibrium partitioning concentrations, apparent effects threshold, {Sigma}PAH model, and threshold and probable effects levels. The resolution of controversial differences among the PAH SQGs lies in an understanding of the effects of mixtures. Polycyclic aromatic hydrocarbons virtually always occur in field-collected sediment as a complex mixture of covarying compounds. When expressed as a mixture concentration, that is, total PAH (TPAH), the guidelines form three clusters that were intended in theirmore » original derivations to represent threshold (TEC = 290 {micro}g/g organic carbon [OC]), median (MEC = 1,800 {micro}g/g OC), and extreme (EEC = 10,000 {micro}g/g OC) effects concentrations. The TEC/MEC/EEC consensus guidelines provide a unifying synthesis of other SQGs, reflect causal rather than correlative effects, account for mixtures, and predict sediment toxicity and benthic community perturbations at sites of PAH contamination. The TEC offers the most useful SQG because PAH mixtures are unlikely to cause adverse effects on benthic ecosystems below the TEC.« less
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Li, Ming; Pan, Yuzhen; Huang, Liping; Zhang, Yong; Yang, Jinhui
2017-03-01
A self-driven microbial fuel cell (MFC) - microbial electrolysis cell (MEC) system, where electricity generated from MFCs is in situ utilized for powering MECs, has been previously reported for recovering Cr(VI), Cu(II) and Cd(II) with individual metals fed in different units of the system in batch operation. Here it was advanced with treating synthetic mixed metals' solution at appropriately adjusting composites in fed-batch and continuous flow operations for complete separation of Cr(VI), Cu(II) and Cd(II) from each other. Under an optimal condition of hydraulic residence time of 4 h, matching of two serially connected MFCs with one MEC, and fed with a composite of either 5 mg L -1 Cr(VI), 1 mg L -1 Cu(II) and 5 mg L -1 Cd(II), or 1 mg L -1 Cr(VI), 5 mg L -1 Cu(II) and 5 mg L -1 Cd(II), the self-driven MFC-MEC system can completely and sequentially recover Cu(II), Cr(VI) and Cd(II) from mixed metals. This study provides a true sustainable and zero-energy-consumed approach of using bioelectrochemical systems for completely recovering and separating Cr(VI), Cu(II) and Cd(II) from each other or from wastes or contaminated sites.
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Cathelicidin production and release by mammary epithelial cells during infectious mastitis.
Cubeddu, Tiziana; Cacciotto, Carla; Pisanu, Salvatore; Tedde, Vittorio; Alberti, Alberto; Pittau, Marco; Dore, Simone; Cannas, Agnese; Uzzau, Sergio; Rocca, Stefano; Addis, Maria Filippa
2017-07-01
Cathelicidins are well-characterized antimicrobial peptides (AMPs) that are present in significant amounts in mastitic milk. Neutrophils are believed to be the main producers of these AMPs, while the role of mammary epithelial cells (MECs) in their production and release is still unclear. In this work, cathelicidin production patterns were investigated in mammary tissues of ewes infected by Staphylococcus aureus, Streptococcus uberis, or Mycoplasma agalactiae, with a combined approach including immunohistochemistry, immune-colocalization, and fluorescent in situ hybridization. Our results confirm that MECs produce and release cathelicidins in response to different mastitis pathogens. As opposed to neutrophils, however, MECs do not seem to store the preformed protein precursor in their cytoplasm, but appear to synthesize and release it only upon exposure to the microorganisms. Cathelicidin production by MECs appears to occur before leukocyte influx in the milk, suggesting a role for these cells in the initial response of the mammary epithelium to microbial infection. Once in the milk, infiltrating neutrophils release massive amounts of cathelicidin by degranulation and production of neutrophil extracellular traps, acting as the main contributor for cathelicidin abundance in mastitic milk. Taken together, our results support the active contribution of MECs to cathelicidin production and release, and reinforce the value of cathelicidins as sensitive and pathogen-independent mastitis markers. Copyright © 2017 Elsevier B.V. All rights reserved.
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Rivera, Isaac; Bakonyi, Péter; Cuautle-Marín, Manuel Alejandro; Buitrón, Germán
2017-05-01
In this study single-chamber microbial electrolysis cells (MECs) were applied to treat cheese whey (CW), an industrial by-product, and recover H 2 gas. Firstly, this substrate was fed directly to the MEC to get the initial feedback about its H 2 generation potential. The results indicated that the direct application of CW requires an adequate pH control to realize bioelectrohydrogenesis and avoid operational failure due to the loss of bioanode activity. In the second part of the study, the effluents of anaerobic (methanogenic) digester and hydrogenogenic (dark fermentative H 2 -producing) reactor utilizing the CW were tested in the MEC process (representing the concept of a two-stage technology). It turned out that the residue of the methanogenic reactor - with its relatively lower carbohydrate- and higher volatile fatty acid contents - was more suitable to produce hydrogen bioelectrochemically. The MEC operated with the dark fermentation effluent, containing a high portion of carbohydrates and low amount of organic acids, produced significant amount of undesired methane simultaneously with H 2 . Overall, the best MEC behavior was attained using the effluent of the methanogenic reactor and therefore, considering a two-stage system, methanogenesis is an advisable pretreatment step for the acidic CW to enhance the H 2 formation in complementary microbial electrohydrogenesis. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Straub, Jürg Oliver
2016-04-01
Sulfamethoxazole (SMX) is an old sulfonamide antibiotic that was launched first in combination with trimethoprim in 1969 by F.Hoffmann-La Roche. Although sales figures for SMX have been declining over the past 20 yr, the compound is still widely used; moreover, many measured environmental concentrations (MECs) are available from Europe, the United States, Asia, Australia, and Africa. To assess aquatic risks of SMX in Europe, the exposure of European surface waters was predicted based on actual sales figures from IMS Health, incorporating environmental fate data on one side, and based on collated MECs representing more than 5500 single measurements in Europe on the other. Environmental effects were assessed using chronic and subchronic ecotoxicity data for 16 groups of aquatic organisms, from periphyton communities to cyanobacteria, algae, higher plants, various invertebrates, and vertebrates. Predicted no-effect concentrations (PNECs) were derived using both deterministic and probabilistic methodology. The predicted environmental concentration (PEC)/PNEC and MEC/PNEC comparisons overall showed no appreciable risk, except in a low incidence (<0.55%) of cases in which exceptionally high MECs led to MEC/PNEC risk characterization ratios greater than 1. The PNECs derived in the present study can be used to extend aquatic environmental risk assessment for SMX to other continents. No risk appears for indirect human exposure to SMX via the environment. © 2015 SETAC.
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Persistence of Cytosine Methylation of DNA following Fertilisation in the Mouse
Li, Yan; O'Neill, Chris
2012-01-01
Normal development of the mammalian embryo requires epigenetic reprogramming of the genome. The level of cytosine methylation of CpG-rich (5meC) regions of the genome is a major epigenetic regulator and active global demethylation of 5meC throughout the genome is reported to occur within the first cell-cycle following fertilization. An enzyme or mechanism capable of catalysing such rapid global demethylation has not been identified. The mouse is a widely used model for studying developmental epigenetics. We have reassessed the evidence for this phenomenon of genome-wide demethylation following fertilisation in the mouse. We found when using conventional methods of immunolocalization that 5meC showed a progressive acid-resistant antigenic masking during zygotic maturation which gave the appearance of demethylation. Changing the unmasking strategy by also performing tryptic digestion revealed a persistence of a methylated state. Analysis of methyl binding domain 1 protein (MBD1) binding confirmed that the genome remained methylated following fertilisation. The maintenance of this methylated state over the first several cell-cycles required the actions of DNA methyltransferase activity. The study shows that any 5meC remodelling that occurs during early development is not explained by a global active loss of 5meC staining during the cleavage stage of development and global loss of methylation following fertilization is not a major component of epigenetic reprogramming in the mouse zygote. PMID:22292019
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Granisetron Extended-Release Injection: A Review in Chemotherapy-Induced Nausea and Vomiting.
Deeks, Emma D
2016-12-01
An extended-release (ER) subcutaneously injectable formulation of the first-generation 5-HT 3 receptor antagonist granisetron is now available in the USA (Sustol ® ), where it is indicated for the prevention of acute and delayed chemotherapy-induced nausea and vomiting (CINV) following moderately emetogenic chemotherapy (MEC) or anthracycline and cyclophosphamide combination chemotherapy regimens in adults. Granisetron ER is administered as a single subcutaneous injection and uses an erosion-controlled drug-delivery system to allow prolonged granisetron release. Primary endpoint data from phase III studies after an initial cycle of chemotherapy indicate that, when used as part of an antiemetic regimen, granisetron ER injection is more effective than intravenous ondansetron in preventing delayed CINV following highly emetogenic chemotherapy (HEC); is noninferior to intravenous palonosetron in preventing both acute CINV following MEC or HEC and delayed CINV following MEC; and is similar, but not superior, to palonosetron in preventing delayed CINV following HEC. The benefits of granisetron ER were seen in various patient subgroups, including those receiving anthracycline plus cyclophosphamide-based HEC, and (in an extension of one of the studies) over multiple MEC or HEC cycles. Granisetron ER injection is generally well tolerated, with an adverse event profile similar to that of ondansetron or palonosetron. Thus, granisetron ER injection expands the options for preventing both acute and delayed CINV in adults with cancer receiving MEC or anthracycline plus cyclophosphamide-based HEC.
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Pinheiro, Luiza; Brito, Carla Ivo; Pereira, Valéria Cataneli; Oliveira, Adilson; Bartolomeu, Ariane Rocha; Camargo, Carlos Henrique; Cunha, Maria Lourdes Ribeiro Souza
2016-06-01
The aim of this study was to evaluate the antimicrobial susceptibility profile of 85 Staphylococcus epidermidis and 84 Staphylococcus haemolyticus strains isolated from blood cultures to oxacillin, vancomycin, tigecycline, linezolid, daptomycin, and quinupristin/dalfopristin over a period of 12 years. S. epidermidis and S. haemolyticus isolated from blood cultures of inpatients, attended at a teaching hospital, were analyzed for the presence of the mecA gene and by SCCmec typing. The minimum inhibitory concentration (MIC) values of tigecycline, linezolid, daptomycin, quinupristin/dalfopristin, and vancomycin were determined. Isolates exhibiting vancomycin MICs of ≥2 μg/ml were typed by pulsed-field gel electrophoresis (PFGE). The rate of mecA positivity was 92.9% and 100% in S. epidermidis and S. haemolyticus, respectively. The most frequent SCCmec types were type III (53.2%) in S. epidermidis and type I (32.1%) in S. haemolyticus. All isolates were susceptible to linezolid and daptomycin, but 7.1% of S. haemolyticus and 2.3% of S. epidermidis isolates were resistant to tigecycline, and 1.2% each of S. haemolyticus and S. epidermidis were resistant and intermediately resistant to quinupristin/dalfopristin, respectively. S. epidermidis exhibited higher vancomycin MICs (40% with MIC of ≥2 μg/ml). Clonal typing of strains with vancomycin MIC of ≥2 μg/ml revealed the presence of different PFGE types of S. epidermidis and S. haemolyticus over a period of up to 4 years (2002-2004, 2005-2008, 2006-2009, 2010-2011). Despite the observation of a high prevalence of mecA, the clinical strains were fully susceptible to vancomycin and to the new drugs linezolid, daptomycin, tigecycline, and quinupristin/dalfopristin. The PFGE types with vancomycin MIC of ≥2 μg/ml exhibited a great diversity of SCCmec cassettes, demonstrating that S. epidermidis and S. haemolyticus may easily acquire these resistance-conferring genetic elements.
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Zuway, Khaled Y; Smith, Jamie P; Foster, Christopher W; Kapur, Nikil; Banks, Craig E; Sutcliffe, Oliver B
2015-09-21
The global increase in the production and abuse of cathinone-derived New Psychoactive Substances (NPSs) has developed the requirement for rapid, selective and sensitive protocols for their separation and detection. Electrochemical sensing of these compounds has been demonstrated to be an effective method for the in-field detection of these substances, either in their pure form or in the presence of common adulterants, however, the technique is limited in its ability to discriminate between structurally related cathinone-derivatives (for example: (±)-4′-methylmethcathinone (4-MMC, 2a) and (±)-4′-methyl-N-ethylmethcathinone (4-MEC, 2b) when they are both present in a mixture. In this paper we demonstrate, for the first time, the combination of HPLC-UV with amperometric detection (HPLC-AD) for the qualitative and quantitative analysis of 4-MMC and 4-MEC using either a commercially available impinging jet (LC-FC-A) or custom-made iCell channel (LC-FC-B) flow-cell system incorporating embedded graphite screen-printed macroelectrodes. The protocol offers a cost-effective, reproducible and reliable sensor platform for the simultaneous HPLC-UV and amperometric detection of the target analytes. The two systems have similar limits of detection, in terms of amperometric detection [LC-FC-A: 14.66 μg mL(-1) (2a) and 9.35 μg mL(-1) (2b); LC-FC-B: 57.92 μg mL(-1) (2a) and 26.91 μg mL(-1) (2b)], to the previously reported oxidative electrochemical protocol [39.8 μg mL(-1) (2a) and 84.2 μg mL(-1) (2b)], for two synthetic cathinones, prevalent on the recreational drugs market. Though not as sensitive as standard HPLC-UV detection, both flow cells show a good agreement, between the quantitative electroanalytical data, thereby making them suitable for the detection and quantification of 4-MMC and 4-MEC, either in their pure form or within complex mixtures. Additionally, the simultaneous HPLC-UV and amperometric detection protocol detailed herein shows a marked improvement and advantage over previously reported electroanalytical methods, which were either unable to selectively discriminate between structurally related synthetic cathinones (e.g. 4-MMC and 4-MEC) or utilised harmful and restrictive materials in their design.
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Duong, MyLinh T.; Akli, Said; Wei, Caimiao; Wingate, Hannah F.; Liu, Wenbin; Lu, Yiling; Yi, Min; Mills, Gordon B.; Hunt, Kelly K.; Keyomarsi, Khandan
2012-01-01
Elastase-mediated cleavage of cyclin E generates low molecular weight cyclin E (LMW-E) isoforms exhibiting enhanced CDK2–associated kinase activity and resistance to inhibition by CDK inhibitors p21 and p27. Approximately 27% of breast cancers express high LMW-E protein levels, which significantly correlates with poor survival. The objective of this study was to identify the signaling pathway(s) deregulated by LMW-E expression in breast cancer patients and to identify pharmaceutical agents to effectively target this pathway. Ectopic LMW-E expression in nontumorigenic human mammary epithelial cells (hMECs) was sufficient to generate xenografts with greater tumorigenic potential than full-length cyclin E, and the tumorigenicity was augmented by in vivo passaging. However, cyclin E mutants unable to interact with CDK2 protected hMECs from tumor development. When hMECs were cultured on Matrigel, LMW-E mediated aberrant acinar morphogenesis, including enlargement of acinar structures and formation of multi-acinar complexes, as denoted by reduced BIM and elevated Ki67 expression. Similarly, inducible expression of LMW-E in transgenic mice generated hyper-proliferative terminal end buds resulting in enhanced mammary tumor development. Reverse-phase protein array assay of 276 breast tumor patient samples and cells cultured on monolayer and in three-dimensional Matrigel demonstrated that, in terms of protein expression profile, hMECs cultured in Matrigel more closely resembled patient tissues than did cells cultured on monolayer. Additionally, the b-Raf-ERK1/2-mTOR pathway was activated in LMW-E–expressing patient samples, and activation of this pathway was associated with poor disease-specific survival. Combination treatment using roscovitine (CDK inhibitor) plus either rapamycin (mTOR inhibitor) or sorafenib (a pan kinase inhibitor targeting b-Raf) effectively prevented aberrant acinar formation in LMW-E–expressing cells by inducing G1/S cell cycle arrest. LMW-E requires CDK2–associated kinase activity to induce mammary tumor formation by disrupting acinar development. The b-Raf-ERK1/2-mTOR signaling pathway is aberrantly activated in breast cancer and can be suppressed by combination treatment with roscovitine plus either rapamycin or sorafenib. PMID:22479189
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Mucoepidermoid carcinoma of the posterior-lateral border of tongue: a rare presentation.
Mathew, Lisha; Janardhanan, Mahija; Suresh, Rakesh; Savithri, Vindhya
2017-10-24
Mucoepidermoid carcinoma (MEC) is the most common malignant tumour of the major and minor salivary glands. Minor salivary glands are scattered in different areas of the oral cavity such as palate, retromolar area, floor of the mouth, buccal mucosa, lips and tongue, but so far, only a few lingual MEC cases have been documented in the literature and most of the studies have shown a predilection for base and dorsum of the tongue. We report a rare case of MEC involving the posterior-lateral border of the tongue. © BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.
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Jiménez, Judy Natalia; Ocampo, Ana María; Vanegas, Johanna Marcela; Rodríguez, Erika Andrea; Garcés, Carlos Guillermo; Patiño, Luz Adriana; Ospina, Sigifredo; Correa, Margarita María
2011-12-01
Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric patients between zero-14 years. We identified the genes encoding virulence factors, which included Panton-Valentine leucocidine (PVL), staphylococcal enterotoxins A-E, exfoliative toxins A and B and toxic shock syndrome toxin 1. Typing of the staphylococcal chromosome cassette mec (SCCmec) was performed in MRSA strains. The virulence genes were more diverse and frequent in MSSA than in MRSA isolates (83% vs. 73%). MRSA strains harboured SCCmec types IVc (60%), I (30%), IVa (7%) and V (3%). SCCmec type IVc isolates frequently carried the PVL encoding genes and harboured virulence determinants resembling susceptible strains while SCCmec type I isolates were often negative. PVL was not exclusive to skin and soft tissue infections. As previously suggested, these differences in the distribution of virulence factor genes may be due to the fitness cost associated with methicillin resistance.
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Lewis, Alex J.; Borole, Abhijeet P.
2016-06-16
We investigated the effect of flow rate and recycle on the conversion of a biomass-derived pyrolysis aqueous phase in amicrobial electrolysis cell (MEC) to demonstrate production of renewable hydrogen in biorefinery. A continuous MEC operation was investigated under one-pass and recycle conditions usingthe complex, biomass-derived, fermentable, mixed substrate feed at a constant concentration of 0.026 g/L,while testing flow rates ranging from 0.19 to 3.6 mL/min. This corresponds to an organic loading rate (OLR) of 0.54₋10 g/L-day. Mass transfer issues observed at low flow rates were alleviated using high flow rates.Increasing the flow rate to 3.6 mL/min (3.7 min HRT) duringmore » one-pass operation increased the hydrogen productivity 3-fold, but anode conversion efficiency (ACE) decreased from 57.9% to 9.9%. Recycle of the anode liquid helped to alleviate kinetic limitations and the ACE increased by 1.8-fold and the hydrogen productivity by 1.2-fold compared to the one-pass condition at the flow rate of 3.6 mL/min (10 g/L-d OLR). High COD removal was also achieved under recycle conditions, reaching 74.2 1.1%, with hydrogen production rate of 2.92 ± 0.51 L/L-day. This study demonstrates the advantages of combining faster flow rates with a recycle process to improve rate of hydrogen production from a switchgrass-derived stream in the biorefinery.« less
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ERIC Educational Resources Information Center
Berksteiner, Earl J.
2013-01-01
The purpose of this quantitative descriptive correlational study was to determine if associations existed between middle- and early-college (MEC) principals' leadership styles, teacher motivation, and teacher satisfaction. MEC programs were programs designed to assist high school students who were not served well in a traditional setting (Middle…
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Growing Our Workforce through Business and Education
ERIC Educational Resources Information Center
Pauley, Douglas R.; Davidchik, Daniel
2010-01-01
In 2004, Central Community College (CCC) established the Mechatronics Education Center (MEC), a regional center of excellence, to help the state address the shortage of skilled technicians in the area of industrial automation. The MEC addresses the needs of the current and future workforce through the implementation of its three main components:…
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Enhanced succinic acid production from corncob hydrolysate by microbial electrolysis cells.
Zhao, Yan; Cao, Weijia; Wang, Zhen; Zhang, Bowen; Chen, Kequan; Ouyang, Pingkai
2016-02-01
In this study, Actinobacillus succinogenes NJ113 microbial electrolysis cells (MECs) were used to enhance the reducing power responsible for succinic acid production from corncob hydrolysate. During corncob hydrolysate fermentation, electric MECs resulted in a 1.31-fold increase in succinic acid production and a 1.33-fold increase in the reducing power compared with those in non-electric MECs. When the hydrolysate was detoxified by combining Ca(OH)2, NaOH, and activated carbon, succinic acid production increased from 3.47 to 6.95 g/l. Using a constant potential of -1.8 V further increased succinic acid production to 7.18 g/l. A total of 18.09 g/l of succinic acid and a yield of 0.60 g/g total sugar were obtained after a 60-h fermentation when NaOH was used as a pH regulator. The improved succinic acid yield from corncob hydrolysate fermentation using A. succinogenes NJ113 in electric MECs demonstrates the great potential of using biomass as a feedstock to cost-effectively produce succinate. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Li, Xiaohu; Zhang, Ruizhe; Qian, Yawei; Angelidaki, Irini; Zhang, Yifeng
2017-07-01
The impact of different anode acclimation methods for enhancing hydrogen production in microbial electrolysis cell (MEC) was investigated in this study. The anodes were first acclimated in microbial fuel cells using acetate, butyrate and corn stalk fermentation effluent (CSFE) as substrate before moving into MECs, respectively. Subsequently, CSFE was used as feedstock in all the three MECs. The maximum hydrogen yield with the anode pre-acclimated with butyrate (5.21±0.24L H 2 /L CSFE) was higher than that pre-acclimated with acetate (4.22±0.19L H 2 /L CSFE) and CSFE (4.55±0.14L H 2 /L CSFE). The current density (480±11A/m 3 ) and hydrogen production rate (4.52±0.13m 3 /m 3 /d) with the anode pre-acclimated with butyrate were also higher that another two reactors. These results demonstrated that the anode biofilm pre-acclimated with butyrate has significant advantages in CSFE treatment and could improve the performance of hydrogen production in MEC. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Mehta, Sanjay R; Estrada, Jasmine; Ybarra, Juan; Fierer, Joshua
2017-04-01
Variation in MRSA genotypes may affect the sensitivity of molecular assays to detect this organism. We compared 2 commonly used screening assays, the Cepheid™ Xpert® MRSA and the BD MAX™ MRSA XT on consecutively obtained nasal swabs from 479 subjects. Specimens giving discordant results were subjected to additional microbiologic and molecular testing. Six hundred forty-two (97.6%) of the 658 test results were concordant. Of the 16 discordant results from 12 subjects, additional results suggested that 9 (60%) of the 15 MRSA XT assays were likely correct, and 6 (40%) of the 15 Xpert® assays were likely correct. One discordant result could not be resolved. A mecA dropout and novel mec right-extremity junction (MREJ) sites led to false-positive and negative results by Xpert®. While both assays performed well, continued vigilance is needed to monitor for Staphylococcus aureus with novel MREJ sites, mecA dropouts, and mecC, leading to inaccurate results in screening assays. Published by Elsevier Inc.
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de Souza, Lélia Batista; de Oliveira, Lucileide Castro; Nonaka, Cassiano Francisco Weege; Lopes, Maria Luiza Diniz de Sousa; Pinto, Leão Pereira; Queiroz, Lélia Maria Guedes
2017-06-01
This study aimed to evaluate and compare the immunoexpression of glucose transporter-1 (GLUT-1) and angiogenic index between pleomorphic adenomas (PAs), adenoid cystic carcinomas (ACCs), and mucoepidermoid carcinomas (MECs) of the salivary glands, and establish associations with the respective subtype/histological grade. Twenty PAs, 20 ACCs, and 10 MECs were submitted to morphological and immunohistochemical analysis. GLUT-1 expression was semi-quantitatively evaluated and angiogenic index was assessed by microvessel counts using anti-CD34 antibody. Higher GLUT-1 immunoexpression was observed in the MECs compared to PAs and ACCs (p = 0.022). Mean number of microvessels was 66.5 in MECs, 40.4 in PAs, and 21.2 in ACCs (p < 0.001). GLUT-1 expression and angiogenic index showed no significant correlation in the tumors studied. Results suggest that differences in biological behavior of the studied tumors are related to GLUT-1. Benign and malignant salivary gland tumors differ in the angiogenic index; however, angiogenesis may be independent of the tumor cell's metabolic demand.
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Evaluation of pharmaceuticals in surface water: reliability of PECs compared to MECs.
Celle-Jeanton, Hélène; Schemberg, Dimitri; Mohammed, Nabaz; Huneau, Frédéric; Bertrand, Guillaume; Lavastre, Véronique; Le Coustumer, Philippe
2014-12-01
Due to the current analytical processes that are not able to measure all the pharmaceutical molecules and to the high costs and the consumption of time to sample and analyze PhACs, models to calculate Predicted Environmental Concentrations (PECs) have been developed. However a comparison between MECs and PECs, taking into account the methods of calculations and peculiarly the parameters included in the calculation (consumption data, pharmacokinetic parameters, elimination rate in STPs and in the environment), is necessary to assess the validity of PECs. MEC variations of sixteen target PhACs [acetaminophen (ACE), amlodipine (AML), atenolol (ATE), caffeine (CAF), carbamazepine (CAR), doxycycline (DOX), epoxycarbamazepine (EPO), fluvoxamine (FLU), furosemide (FUR), hydrochlorothiazide (HYD), ifosfamide (IFO), losartan (LOS), pravastatin (PRA), progesterone (PROG), ramipril (RAM), trimetazidine (TRI)] have been evaluated during one hydrological cycle, from October 2011 to October 2012 and compared to PECs calculated by using an adaptation of the models proposed by Heberer and Feldmann (2005) and EMEA (2006). Comparison of PECs and MECS has been achieved for six molecules: ATE, CAR, DOX, FUR, HYD and PRA. DOX, FUR and HYD present differences between PECs and MECs on an annual basis but their temporal evolutions follow the same trends. PEC evaluation for these PhACs could then be possible but need some adjustments of consumption patterns, pharmacokinetic parameters and/or mechanisms of (bio)degradation. ATE, CAR and PRA are well modeled; PECs can then be used as reliable estimation of concentrations without any reserve. Copyright © 2014 Elsevier Ltd. All rights reserved.
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Cerrillo, Míriam; Viñas, Marc; Bonmatí, August
2017-03-01
Thermophilic anaerobic digestion (AD) of pig slurry coupled to a microbial electrolysis cell (MEC) with a recirculation loop was studied at lab-scale as a strategy to increase AD stability when submitted to organic and nitrogen overloads. The system performance was studied, with the recirculation loop both connected and disconnected, in terms of AD methane production, chemical oxygen demand removal (COD) and volatile fatty acid (VFA) concentrations. Furthermore, the microbial population was quantitatively and qualitatively assessed through DNA and RNA-based qPCR and high throughput sequencing (MiSeq), respectively to identify the RNA-based active microbial populations from the total DNA-based microbial community composition both in the AD and MEC reactors under different operational conditions. Suppression of the recirculation loop reduced the AD COD removal efficiency (from 40% to 22%) and the methane production (from 0.32 to 0.03 m 3 m -3 d -1 ). Restoring the recirculation loop led to a methane production of 0.55 m 3 m -3 d -1 concomitant with maximum MEC COD and ammonium removal efficiencies of 29% and 34%, respectively. Regarding microbial analysis, the composition of the AD and MEC anode populations differed from really active microorganisms. Desulfuromonadaceae was revealed as the most active family in the MEC (18%-19% of the RNA relative abundance), while hydrogenotrophic methanogens (Methanobacteriaceae) dominated the AD biomass. Copyright © 2016 Elsevier Ltd. All rights reserved.
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Saunders, Leslie J; Mazumder, Asit; Lowe, Christopher J
2016-04-01
Pharmaceuticals and personal care products (PPCPs) are emerging chemicals of concern detected in surface waters globally. Recent reviews advocate that PPCP occurrence, fate, and exposure need to be better predicted and characterized. The use of pharmaceutical prescription rates to estimate PPCP concentrations in the environment has been suggested. Concentrations of 7 pharmaceuticals (acetylsalicylic acid, diclofenac, fenoprofen, gemfibrozil, ibuprofen, ketoprofen, and naproxen) were measured in municipal wastewater using gas chromatography/ion trap-tandem mass spectroscopy (GC/IT-MS/MS). Subregional pharmaceutical prescription data were investigated to determine whether they could predict measured effluent concentrations (MECs) in wastewaters. Predicted effluent concentrations (PECs) for 5 of the 7 pharmaceuticals were within 2-fold agreement of the MECs when the fraction of parent pharmaceutical excreted was not considered. When the fraction of parent pharmaceutical excreted was considered, the respective PECs decreased, and most were within an order of magnitude of the MECs. Regression relationships of monthly PECs versus MECs were statistically significant (p < 0.05) but weak (R(2) = 0.18-0.56) for all pharmaceuticals except ketoprofen. This suggests high variability in the data and may be the result of factors influencing MECs such as the analytical methods used, wastewater sampling frequency, and methodology. The PECs were based solely on prescription rates and did not account for inputs of pharmaceuticals that had a significant over-the-counter component or were from other sources (e.g., hospitals). © 2015 SETAC.
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Ishibashi, Kenichiro; Ito, Yohei; Masaki, Ayako; Fujii, Kana; Beppu, Shintaro; Sakakibara, Takeo; Takino, Hisashi; Takase, Hiroshi; Ijichi, Kei; Shimozato, Kazuo; Inagaki, Hiroshi
2015-11-01
There has been some debate as to whether a subset of metaplastic Warthin tumors (mWTs) harbor the mucoepidermoid carcinoma (MEC)-associated CRTC1-MAML2 fusion. We analyzed 15 tumors originally diagnosed as mWT (mWT-like tumors), 2 of which had concurrent MECs. We looked for the CRTC1/3-MAML2 fusion transcripts and performed immunohistochemistry for p63 and fluorescence in situ hybridization (FISH) for the MAML2 split. To localize MAML2 split-positive cells at the cellular level, whole tumor tissue sections were digitalized (whole-slide imaging [WSI]). The CRTC1-MAML2, but not CRTC3-MAML2 was detected in 5/15 mWT-like tumors. FISH-WSI results showed that all epithelial cells harbored the MAML2 split in fusion-positive mWT-like tumors and were totally negative in fusion-negative mWT-like tumors. A review of the hematoxylin and eosin-stained slides showed that morphology of the "metaplastic" epithelium was virtually indistinguishable between fusion-positive and fusion-negative tumors. However, oncocytic bilayered tumor epithelium, characteristic to typical WT, was always found somewhere in the fusion-negative tumors but not in the fusion-positive tumors. This distinguishing histologic finding enabled 5 pathologists to easily differentiate the 2 tumor groups with 100% accuracy. The age and sex distribution of fusion-positive mWT-like tumor cases was similar to that of fusion-positive MEC cases and significantly different from those of fusion-negative mWT-like tumor and typical WT cases. In addition, only fusion-positive mWT-like tumors possessed concurrent low-grade MECs. In conclusion, a subset of mWT-like tumors were positive for the CRTC1-MAML2 fusion and had many features that are more in accord with MEC than with WT. The term Warthin-like MEC should be considered for fusion-positive mWT-like tumors.
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Zamalloa, Carlos; Arends, Jan B A; Boon, Nico; Verstraete, Willy
2013-06-25
Septic tanks are used for the removal of organic particulates in wastewaters by physical accumulation instead of through the biological production of biogas. Improved biogas production in septic tanks is crucial to increase the potential of this system for both energy generation and organic matter removal. In this study, the effect on the biogas production and biogas quality of coupling a 20 L lab-scale septic tank with a microbial electrolysis cell (MEC) was investigated and compared with a standard septic tank. Both reactors were operated at a volumetric organic loading rate of 0.5gCOD/Ld and a hydraulic retention time between 20 and 40 days using black water as an input under mesophilic conditions for a period of 3 months. The MEC-septic tank was operated at an applied voltage of 2.0±0.1V and the current experienced ranged from 40 mA (0.9A/m(2) projected electrode area) to 180 mA (5A/m(2) projected electrode area). The COD removal was of the order of 85% and the concentration of residual COD was not different between both reactors. Yet, the total phosphorous in the output was on average 39% lower in the MEC-septic tank. Moreover, the biogas production rate in the MEC-septic tank was a factor of 5 higher than in the control reactor and the H2S concentration in the biogas was a factor of 2.5 lower. The extra electricity supplied to the MEC-septic tank was recovered as extra biogas produced. Overall, it appears that the combination of MEC and a septic tank offers perspectives in terms of lower discharge of phosphorus and H2S, nutrient recuperation and a more reliable supply of biogas. Copyright © 2013 Elsevier B.V. All rights reserved.
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Nousia, C S; Gouveris, H; Giatromanolaki, A; Katotomichelakis, M; Ypsilantis, P; Riga, M; Sivridis, E; Watelet, J B; van Cauwenberge, P; Danielides, V
2013-06-01
Extracellular matrix (ECM) proteins such as fibronectin and collagen III, enzymes such as matrix metalloproteinases and macrophages have been demonstrated to intervene in nasal and paranasal sinuses wound healing. To compare concentration of ECM proteins, enzymes and the recruitment of macrophages during wound repair after monopolar electrocautery in contrast with ultrasound submucosal surgical tissue reduction of inferior nasal turbinate (INT) tested in sheep. Prospective controlled study in sheep. Immunostaining for collagen III, fibronectin, CD68 and matrix metalloproteinase-9 (MMP9) was applied in tissue specimens of INT mucosa after monopolar electrocoagulation (MEC) and ultrasound tissue reduction (UTR). Twelve INTs were studied 1, 3 and 8 weeks post-operatively in each interventional group (MEC and UTR) and 5 INTs were studied in animals of the control group (without surgery). The immunoreactivity was quantitatively graded between 0% to 100% immunoreactivity by a blinded senior pathologist. At the end of the study period collagen III, fibronectin and MMP9 were increased in both groups compared to the levels of the control group. When compared to control group, CD68 immunoreactivity was found higher in MEC group but not in UTR group. Fibronectin subepithelial immunoreactivity exhibited a substantial negative correlation with mucosal epithelial cell necrosis, a substantial positive correlation with fibrosis in MEC-treated specimens and a significant positive correlation with sinusoid engorgement in UTR-treated specimens. Collagen III tissue immunoreactivity showed a particularly significant negative correlation with sinusoid engorgement in MEC-treated specimens. Correlation of fibronectin and collagen III immunoreactivity to histopathologic findings suggests different ECM repair processes between MEC and UTR turbinate tissue reduction. The use of CD68 and MMP9 provides additional clues to the mode of actions of these techniques and to the molecular and cellular events of the nasal mucosa wound healing process.
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Hydrogen production by geobacter species and a mixed consortium in a microbial electrolysis cell.
Call, Douglas F; Wagner, Rachel C; Logan, Bruce E
2009-12-01
A hydrogen utilizing exoelectrogenic bacterium (Geobacter sulfurreducens) was compared to both a nonhydrogen oxidizer (Geobacter metallireducens) and a mixed consortium in order to compare the hydrogen production rates and hydrogen recoveries of pure and mixed cultures in microbial electrolysis cells (MECs). At an applied voltage of 0.7 V, both G. sulfurreducens and the mixed culture generated similar current densities (ca. 160 A/m3), resulting in hydrogen production rates of ca. 1.9 m(3) H2/m3/day, whereas G. metallireducens exhibited lower current densities and production rates of 110 +/- 7 A/m3 and 1.3 +/- 0.1 m3 H2/m3/day, respectively. Before methane was detected in the mixed-culture MEC, the mixed consortium achieved the highest overall energy recovery (relative to both electricity and substrate energy inputs) of 82% +/- 8% compared to G. sulfurreducens (77% +/- 2%) and G. metallireducens (78% +/- 5%), due to the higher coulombic efficiency of the mixed consortium. At an applied voltage of 0.4 V, methane production increased in the mixed-culture MEC and, as a result, the hydrogen recovery decreased and the overall energy recovery dropped to 38% +/- 16% compared to 80% +/- 5% for G. sulfurreducens and 76% +/- 0% for G. metallireducens. Internal hydrogen recycling was confirmed since the mixed culture generated a stable current density of 31 +/- 0 A/m3 when fed hydrogen gas, whereas G. sulfurreducens exhibited a steady decrease in current production. Community analysis suggested that G. sulfurreducens was predominant in the mixed-culture MEC (72% of clones) despite its relative absence in the mixed-culture inoculum obtained from a microbial fuel cell reactor (2% of clones). These results demonstrate that Geobacter species are capable of obtaining similar hydrogen production rates and energy recoveries as mixed cultures in an MEC and that high coulombic efficiencies in mixed culture MECs can be attributed in part to the recycling of hydrogen into current.
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Gatti, Matteo; Dosso, Paolo; Maurino, Marco; Merli, Maria Clara; Bernizzoni, Fabio; José Pirez, Facundo; Platè, Bonfiglio; Bertuzzi, Gian Carlo; Poni, Stefano
2016-01-01
Ground-based proximal sensing of vineyard features is gaining interest due to its ability to serve in even quite small plots with the advantage of being conducted concurrently with normal vineyard practices (i.e., spraying, pruning or soil tilling) with no dependence upon weather conditions, external services or law-imposed limitations. The purpose of the present work was to test performance of the new terrestrial multi-sensor MECS-VINE® in terms of reliability and degree of correlation with several canopy growth and yield parameters in the grapevine. MECS-VINE®, once conveniently positioned in front of the tractor, can provide simultaneous assessment of growth features and microclimate of specific canopy sections of the two adjacent row sides. MECS-VINE® integrates a series of microclimate sensors (air relative humidity, air and surface temperature) with two (left and right) matrix-based optical RGB imaging sensors and a related algorithm, termed Canoyct). MECS-VINE® was run five times along the season in a mature cv. Barbera vineyard and a Canopy Index (CI, pure number varying from 0 to 1000), calculated through its built-in algorithm, validated vs. canopy structure parameters (i.e., leaf layer number, fractions of canopy gaps and interior leaves) derived from point quadrat analysis. Results showed that CI was highly correlated vs. any canopy parameter at any date, although the closest relationships were found for CI vs. fraction of canopy gaps (R2 = 0.97) and leaf layer number (R2 = 0.97) for data pooled over 24 test vines. While correlations against canopy light interception and total lateral leaf area were still unsatisfactory, a good correlation was found vs. cluster and berry weight (R2 = 0.76 and 0.71, respectively) suggesting a good potential also for yield estimates. Besides the quite satisfactory calibration provided, main improvements of MECS-VINE® usage versus other current equipment are: (i) MECS-VINE® delivers a segmented evaluation of the canopy up to 15 different sectors, therefore allowing to differentiate canopy structure and density at specific and crucial canopy segments (i.e., basal part where clusters are located) and (ii) the sensor is optimized to work at any time of the day with any weather condition without the need of any supplemental lighting system. PMID:27898049
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Hönigsperger, Christoph; Nigro, Maximiliano J.
2016-01-01
Key points Kv2 channels underlie delayed‐rectifier potassium currents in various neurons, although their physiological roles often remain elusive. Almost nothing is known about Kv2 channel functions in medial entorhinal cortex (mEC) neurons, which are involved in representing space, memory formation, epilepsy and dementia.Stellate cells in layer II of the mEC project to the hippocampus and are considered to be space‐representing grid cells. We used the new Kv2 blocker Guangxitoxin‐1E (GTx) to study Kv2 functions in these neurons.Voltage clamp recordings from mEC stellate cells in rat brain slices showed that GTx inhibited delayed‐rectifier K+ current but not transient A‐type current.In current clamp, GTx had multiple effects: (i) increasing excitability and bursting at moderate spike rates but reducing firing at high rates; (ii) enhancing after‐depolarizations; (iii) reducing the fast and medium after‐hyperpolarizations; (iv) broadening action potentials; and (v) reducing spike clustering.GTx is a useful tool for studying Kv2 channels and their functions in neurons. Abstract The medial entorhinal cortex (mEC) is strongly involved in spatial navigation, memory, dementia and epilepsy. Although potassium channels shape neuronal activity, their roles in mEC are largely unknown. We used the new Kv2 blocker Guangxitoxin‐1E (GTx; 10–100 nm) in rat brain slices to investigate Kv2 channel functions in mEC layer II stellate cells (SCs). These neurons project to the hippocampus and are considered to be grid cells representing space. Voltage clamp recordings from SCs nucleated patches showed that GTx inhibited a delayed rectifier K+ current activating beyond –30 mV but not transient A‐type current. In current clamp, GTx (i) had almost no effect on the first action potential but markedly slowed repolarization of late spikes during repetitive firing; (ii) enhanced the after‐depolarization (ADP); (iii) reduced fast and medium after‐hyperpolarizations (AHPs); (iv) strongly enhanced burst firing and increased excitability at moderate spike rates but reduced spiking at high rates; and (v) reduced spike clustering and rebound potentials. The changes in bursting and excitability were related to the altered ADPs and AHPs. Kv2 channels strongly shape the activity of mEC SCs by affecting spike repolarization, after‐potentials, excitability and spike patterns. GTx is a useful tool and may serve to further clarify Kv2 channel functions in neurons. We conclude that Kv2 channels in mEC SCs are important determinants of intrinsic properties that allow these neurons to produce spatial representation. The results of the present study may also be important for the accurate modelling of grid cells. PMID:27562026
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NASA Astrophysics Data System (ADS)
de Luca, N.; Ferraz-Mello, S.
Contents: Os princípios da mec@OSC@@CCF@anica celeste. O problema de 2-corpos. O sistema de equações canônicas. A teoria das perturbações. O movimento de um satélite artifical. A teoria lunar e o movimento de outros satélites naturais. A teoria planetária. Procedimentos numéricos.
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Background: There is an increased interest in the application of microbial electrochemical cell (MEC) for the recovery of value-added products such as hydrogen gas and hydrogen peroxide (H2O2) from wastewater. H2O2 has strong oxidation capability and produces hydroxyl radicals wh...
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Karthikeyan, Rengasamy; Cheng, Ka Yu; Selvam, Ammaiyappan; Bose, Arpita; Wong, Jonathan W C
2017-11-01
Microbial electrolysis cells (MECs) are a promising technology for biological hydrogen production. Compared to abiotic water electrolysis, a much lower electrical voltage (~0.2V) is required for hydrogen production in MECs. It is also an attractive waste treatment technology as a variety of biodegradable substances can be used as the process feedstock. Underpinning this technology is a recently discovered bioelectrochemical pathway known as "bioelectrohydrogenesis". However, little is known about the mechanism of this pathway, and numerous hurdles are yet to be addressed to maximize hydrogen yield and purity. Here, we review various aspects including reactor configurations, microorganisms, substrates, electrode materials, and inhibitors of methanogenesis in order to improve hydrogen generation in MECs. Copyright © 2017 Elsevier Inc. All rights reserved.
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Goudarzi, Mehdi; Bahramian, Mahnaz; Satarzadeh Tabrizi, Mahboobeh; Udo, Edet E; Figueiredo, Agnes Marie Sá; Fazeli, Maryam; Goudarzi, Hossein
2017-04-01
Methicillin-resistant Staphylococcus aureus (MRSA) as a major cause of infection in health care, hospital and community settings is a global health concern. The purpose of this study was to determine the antibiotic susceptibility pattern and distribution of circulating molecular types of MRSA in a burn hospital in Tehran, the capital of Iran. During a 10-month study period, 106 Staphylococcus aureus isolates were assessed. Isolates were subjected to susceptibility testing using the disk diffusion method and Polymerase Chain Reaction (PCR) for detection of mecA, fem and nuc genes. The presence of PVL and tst encoding genes were determined by PCR method. All the MRSA isolates were genotyped by multilocus sequence typing (MLST), spa typing, SCCmec typing and agr typing. The presence of mecA gene was confirmed in all the Staphylococcus aureus isolates. Antimicrobial susceptibility testing revealed a high resistance rate (90.6%) to ampicillin, tetracycline, and erythromycin. The rates of resistance to remaining antibiotics tested varied between 18.9% and 84.9%. The high- level of resistance to mupirocin was confirmed in 19.8% of MRSA strains isolated from burn patients. Multi-drug resistance was observed in 90.6% of isolates. Sixteen of the 106 MRSA isolates (15.1%) harbored PVL-encoding genes. The majority of our MRSA strains carried SCCmec III (71.7%). ST239-SCCmec III/t037 (34%) was the most common genotype followed by ST239-SCCmec III/t030 (24.5%), ST15-SCCmec IV/t084 (15.1%), ST22-SCCmec IV/t790 (13.2%), and ST239-SCCmec III/t631 (13.2%). Mupirocin resistant MRSA isolates belonged to ST15-SCCmec IV/t084 (40%), ST22-SCCmec IV/t790 (23.3%), ST239-SCCmec III/t631 (20%), and ST239-SCCmec III/t030 (16.7%) clones. The results showed that genetically diverse strains of MRSA are circulating in our burn hospitals with relatively high prevalence of ST239-SCCmec III/t037 clone. The findings support the need for regular surveillance of MRSA to determine the distribution of existing MRSA clones and to detect the emergence of new MRSA clones. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Buchan, Blake W.; Ginocchio, Christine C.; Manii, Ryhana; Cavagnolo, Robert; Pancholi, Preeti; Swyers, Lettie; Thomson, Richard B.; Anderson, Christopher; Kaul, Karen; Ledeboer, Nathan A.
2013-01-01
Background A multicenter study was conducted to evaluate the diagnostic accuracy (sensitivity and specificity) of the Verigene Gram-Positive Blood Culture Test (BC-GP) test to identify 12 Gram-positive bacterial gene targets and three genetic resistance determinants directly from positive blood culture broths containing Gram-positive bacteria. Methods and Findings 1,252 blood cultures containing Gram-positive bacteria were prospectively collected and tested at five clinical centers between April, 2011 and January, 2012. An additional 387 contrived blood cultures containing uncommon targets (e.g., Listeria spp., S. lugdunensis, vanB-positive Enterococci) were included to fully evaluate the performance of the BC-GP test. Sensitivity and specificity for the 12 specific genus or species targets identified by the BC-GP test ranged from 92.6%–100% and 95.4%–100%, respectively. Identification of the mecA gene in 599 cultures containing S. aureus or S. epidermidis was 98.6% sensitive and 94.3% specific compared to cefoxitin disk method. Identification of the vanA gene in 81 cultures containing Enterococcus faecium or E. faecalis was 100% sensitive and specific. Approximately 7.5% (87/1,157) of single-organism cultures contained Gram-positive bacteria not present on the BC-GP test panel. In 95 cultures containing multiple organisms the BC-GP test was in 71.6% (68/95) agreement with culture results. Retrospective analysis of 107 separate blood cultures demonstrated that identification of methicillin resistant S. aureus and vancomycin resistant Enterococcus spp. was completed an average of 41.8 to 42.4 h earlier using the BC-GP test compared to routine culture methods. The BC-GP test was unable to assign mecA to a specific organism in cultures containing more than one Staphylococcus isolate and does not identify common blood culture contaminants such as Micrococcus, Corynebacterium, and Bacillus. Conclusions The BC-GP test is a multiplex test capable of detecting most leading causes of Gram-positive bacterial blood stream infections as well as genetic markers of methicillin and vancomycin resistance directly from positive blood cultures. Please see later in the article for the Editors' Summary PMID:23843749
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Sudagidan, Mert; Aydin, Ali
2010-04-15
In this study, three Panton-Valentine Leukocidin gene carrying methicillin-susceptible Staphylococcus aureus (MSSA) strains (M1-AAG42B, PY30C-b and YF1B-b) were isolated from different food samples in Kesan-Edirne, Turkey. These strains were characterized on the basis of MLST type, spa type, virulence factor gene contents, antibiotic susceptibilities against 21 antibiotics and biofilm formation. The genetic relatedness of the strains was determined by PFGE. In addition, the complete gene sequences of lukS-PV and lukF-PV were also investigated. All strains were found to be susceptible to tested antibiotics and they were mecA negative. Three strains showed the same PFGE band pattern, ST152 clonal type and t355 spa type. In the detection of virulence factor genes, sea, seb, sec, sed, see, seg, seh, sei, sej, sek, sel, sem, sen, seo, sep, seq, seu, eta, etb, set1, geh and tst genes were not detected. All strains showed the positive results for alpha- and beta-haemolysin genes (hla and hlb), protease encoding genes (sspA, sspB and aur), lukE and lukD leukocidin genes (lukED). The strains were found to be non-biofilm formers. By this study, the virulence properties of the strains were described and this is one of the first reports regarding PVL-positive MSSA strains from food. (c) 2010 Elsevier B.V. All rights reserved.
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Staphylococcus aureus from the German general population is highly diverse.
Becker, Karsten; Schaumburg, Frieder; Fegeler, Christian; Friedrich, Alexander W; Köck, Robin
2017-01-01
This prospective cohort study evaluates colonization dynamics and molecular characteristics of methicillin-susceptible and - resistant Staphylococcus aureus (MSSA/MRSA) in a German general population. Nasal swabs of 1878 non-hospitalized adults were screened for S. aureus. Participants were screened thrice in intervals of 6-8 months. Isolates were characterized by spa and agr typing, mecA and mecC possession, respectively, and PCRs targeting virulence factors. 40.9% of all participants carried S. aureus at least once while 0.7% of the participants carried MRSA (mainly spa t011). MSSA isolates (n=1359) were associated with 331 different spa types; t084 (7.7%), t091 (6.1%) and t012 (71, 5.2%) were predominant. Of 206 participants carrying S. aureus at all three sampling time points, 14.1% carried the same spa type continuously; 5.3% carried different spa types with similar repeat patterns, but 80.6% carried S. aureus with unrelated spa types. MSSA isolates frequently harboured genes encoding enterotoxins (sec: 16.6%, seg: 63.1%, sei: 64.5%) and toxic shock syndrome toxin (tst: 17.5%), but rarely Panton-Valentine leukocidin (lukS-PV/lukF-PV: 0.2%). MSSA colonizing human nares in the community are clonally highly diverse. Among those constantly carrying S. aureus, clonal lineages changed over time. The proportion of persistent S. aureus carriers was lower than reported elsewhere. Copyright © 2016 Elsevier GmbH. All rights reserved.
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Baba, Yoshifumi; Baba, Hideo; Yamamoto, Sachiko; Shimada, Hideaki; Shibata, Tomotaka; Miyazaki, Tatsuya; Yoshikawa, Takaki; Nakajima, Yasuaki; Tsuji, Yasushi; Shimokawa, Mototsugu; Kitagawa, Yuko; Aiba, Keisuke
2017-02-01
Chemotherapy is an indispensable therapeutic approach for esophageal cancer. Although chemotherapy-induced nausea and vomiting (CINV) is one of the most crucial adverse events, the current state of CINV in patients with esophageal cancer remains unclear. This multicenter prospective observational study analyzed data for 192 patents with esophageal cancer who underwent moderately emetogenic chemotherapy (MEC) or highly emetogenic chemotherapy (HEC). The patients recorded their CINV incidence and severity daily for 7 days after receiving chemotherapy, using visual analog scales (VAS). Of the 192 patients, 181 received HEC including cisplatin, and 11 patients received MEC including nedaplatin. Approximately 81% of HEC and 82% of MEC patients received antiemetic therapy in compliance with guidelines. Although CINV was controlled relatively well in the early phase (days 1-4), it was not fully controlled in late phase (days 5-7) for both the HEC and MEC groups. Female sex was a major risk factor for delayed vomiting (P=0.034). Multivariate logistic regression analysis for VAS revealed that motion sickness, age, and use of other antiemetics were risk factors for delayed nausea. Adherence to antiemetic guidelines effectively controls vomiting but is less effective against delayed CINV in both HEC and MEC patients. Identification of individual risk factors, such as female sex, will help develop personalized treatments for CINV. In the clinical setting for esophageal cancer, regimens that include nedaplatin might need to be treated as HEC. © 2016 International Society for Diseases of the Esophagus.
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Grid cells form a global representation of connected environments.
Carpenter, Francis; Manson, Daniel; Jeffery, Kate; Burgess, Neil; Barry, Caswell
2015-05-04
The firing patterns of grid cells in medial entorhinal cortex (mEC) and associated brain areas form triangular arrays that tessellate the environment [1, 2] and maintain constant spatial offsets to each other between environments [3, 4]. These cells are thought to provide an efficient metric for navigation in large-scale space [5-8]. However, an accurate and universal metric requires grid cell firing patterns to uniformly cover the space to be navigated, in contrast to recent demonstrations that environmental features such as boundaries can distort [9-11] and fragment [12] grid patterns. To establish whether grid firing is determined by local environmental cues, or provides a coherent global representation, we recorded mEC grid cells in rats foraging in an environment containing two perceptually identical compartments connected via a corridor. During initial exposures to the multicompartment environment, grid firing patterns were dominated by local environmental cues, replicating between the two compartments. However, with prolonged experience, grid cell firing patterns formed a single, continuous representation that spanned both compartments. Thus, we provide the first evidence that in a complex environment, grid cell firing can form the coherent global pattern necessary for them to act as a metric capable of supporting large-scale spatial navigation. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.
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Grid Cells Form a Global Representation of Connected Environments
Carpenter, Francis; Manson, Daniel; Jeffery, Kate; Burgess, Neil; Barry, Caswell
2015-01-01
Summary The firing patterns of grid cells in medial entorhinal cortex (mEC) and associated brain areas form triangular arrays that tessellate the environment [1, 2] and maintain constant spatial offsets to each other between environments [3, 4]. These cells are thought to provide an efficient metric for navigation in large-scale space [5–8]. However, an accurate and universal metric requires grid cell firing patterns to uniformly cover the space to be navigated, in contrast to recent demonstrations that environmental features such as boundaries can distort [9–11] and fragment [12] grid patterns. To establish whether grid firing is determined by local environmental cues, or provides a coherent global representation, we recorded mEC grid cells in rats foraging in an environment containing two perceptually identical compartments connected via a corridor. During initial exposures to the multicompartment environment, grid firing patterns were dominated by local environmental cues, replicating between the two compartments. However, with prolonged experience, grid cell firing patterns formed a single, continuous representation that spanned both compartments. Thus, we provide the first evidence that in a complex environment, grid cell firing can form the coherent global pattern necessary for them to act as a metric capable of supporting large-scale spatial navigation. PMID:25913404
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USDA-ARS?s Scientific Manuscript database
We evaluated whether the efficacy of various insecticides for codling moth, Cydia pomonella (L.), could be improved with the addition of a microencapsulated formulation of pear ester, ethyl (2E, 4Z)-2,4-decadienoate (PE-MEC, 5% AI), in field trials from 2005 to 2009. The addition of PE-MEC (< 3.0 g ...
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Liu, Wei; Niu, Xiaojun; Chen, Weiyi; An, Shaorong; Sheng, Hong
2017-04-01
Phosphine (PH 3 ) emission from conventional biological wastewater treatment is very inefficient (ng-μg m -3 ). In this work, we investigated the feasibility of promoting PH 3 formation from inorganic phosphorus (IP) or organic phosphorus (OP) containing synthetic wastewater treatment by Microbial Electrolysis Cell (MEC) for the first time. Positive effect of applied potential on PH 3 production was observed after methanogens was inhibited. The highest production of PH 3 (1103.10 ± 72.02 ng m -3 ) was obtained in IP-fed MEC operated at -0.6 V, which was about 5-fold and 2-fold compared to that in open circuit experiment and OP-fed MEC, respectively. Meanwhile, PH 3 formation corresponded positively with current density and alkaline phosphatase activity. This result showed that suitable potential could enhance the activity of relevant enzymes and boost the biosynthesis of PH 3 . Bacterial communities analysis based on high-throughput sequencing revealed that applied potential was conductive to the enrichment of phosphate-reducing organisms in contrast to the control test. These results provide a new idea for resource utilization of phosphorus in wastewater. Copyright © 2017 Elsevier Ltd. All rights reserved.
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Incorporating advanced EMI technologies in operational munitions characterization surveys
NASA Astrophysics Data System (ADS)
Miller, Jonathan S.; Shubiditze, Fridon; Pasion, Leonard; Schultz, Gregory; Chung, Heesoo
2011-06-01
The presence of unexploded ordnance (UXO), discarded military munitions (DMM), and munitions constituents (MC) at both active and formerly used defense sites (FUDS) has created a necessity for production-level efforts to remove these munitions and explosives of concern (MEC). Ordnance and explosives (OE) and UXO removal operations typically employ electromagnetic induction (EMI) or magnetometer surveys to identify potential MEC hazards in previously determined areas of interest. A major cost factor in these operations is the significant allocation of resources for the excavation of harmless objects associated with fragmentation, scrap, or geological clutter. Recent advances in classification and discrimination methodologies, as well as the development of sensor technologies that fully exploit physics-based analysis, have demonstrated promise for significantly reducing the false alarm rate due to MEC related clutter. This paper identifies some of the considerations for and the challenges associated with implementing these discrimination methodologies and advanced sensor technologies in production-level surveys. Specifically, we evaluate the implications of deploying an advanced multi-axis EMI sensor at a variety of MEC sites, the discrimination methodologies that leverage the data produced by this sensor, and the potential for productivity increase that could be realized by incorporating this advanced technology as part of production protocol.
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Nagasawa, Yuya; Kiku, Yoshio; Sugawara, Kazue; Tanabe, Fuyuko; Hayashi, Tomohito
2018-01-01
The exfoliation rate of mammary epithelial cells (MECs) in milk is affected by physiological, breeding and environmental factors. Little is known about the relationship between the MEC exfoliation into milk and mammary-infected Staphylococcus aureus (S. aureus) load on bovine mastitis caused by S. aureus. The aim of this study was to investigate the relationship between S. aureus load and the proportion of MEC exfoliation in milk using five substantial bovine mastitis models. In 64 randomly extracted milk samples from udders at 3-21 days after S. aureus infusion, there were various samples with different numbers of S. aureus counts and somatic cell counts. No significant correlations were found between the S. aureus counts and somatic cell count (r = 0.338). In contrast, a significant correlation was noted between S. aureus counts and the proportion of cytokeratin-positive cells in the milk from the infused udders (r = 0.734, P < 0.01). In conclusion, the increasing MEC exfoliation rate in milk from mastitis udders caused by S. aureus may contribute to reduced milk yield. © 2017 Japanese Society of Animal Science.
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Functional Division of Hippocampal Area CA1 Via Modulatory Gating of Entorhinal Cortical Inputs
Ito, Hiroshi T.; Schuman, Erin M.
2013-01-01
The hippocampus receives two streams of information, spatial and nonspatial, via major afferent inputs from the medial (MEC) and lateral entorhinal cortexes (LEC). The MEC and LEC projections in the temporoammonic pathway are topographically organized along the transverse-axis of area CA1. The potential for functional segregation of area CA1, however, remains relatively unexplored. Here, we demonstrated differential novelty-induced c-Fos expression along the transverse-axis of area CA1 corresponding to topographic projections of MEC and LEC inputs. We found that, while novel place exposure induced a uniform c-Fos expression along the transverse-axis of area CA1, novel object exposure primarily activated the distal half of CA1 neurons. In hippocampal slices, we observed distinct presynaptic properties between LEC and MEC terminals, and application of either DA or NE produced a largely selective influence on one set of inputs (LEC). Finally, we demonstrated that differential c-Fos expression along the transverse axis of area CA1 was largely abolished by an antagonist of neuromodulatory receptors, clozapine. Our results suggest that neuromodulators can control topographic TA projections allowing the hippocampus to differentially encode new information along the transverse axis of area CA1. PMID:21240920
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The performance of 3-D graphite doped anodes in microbial electrolysis cells
NASA Astrophysics Data System (ADS)
Yasri, Nael G.; Nakhla, George
2017-02-01
This study investigated the use of granular activated carbon (GAC) as high surface area 3-dimensional (3-D) anode in MECs systems. The interfacial anodes' charge transfer resistance of the doped GAC did not impact the overall performance of MECs. Based on our finding, the 3-D anode packed with GAC-doped with nonconductive calcium sulfide (CaS) outperformed the more conductive iron (II) sulfide (FeS), magnetite (Fe3O4), or GAC without doping. The results showed higher current densities for 3-D CaS (40.1 A/m3), as compared with 3-D FeS (34.4 A/m3), 3-D Fe3O4 (29.8 A/m3), and 3-D GAC (23.1 A/m3). The higher current density in the 3-D CaS translated to higher coulombic efficiency (96.7%), hydrogen yield (3.6 mol H2/mol acetate), and attached biomass per anode mass (54.01 mg COD biomass/g GAC). Although the 3-D MEC achieved similar hydrogen yield, hydrogen recovery efficiency, and COD removal rate to a conventional sandwich type MEC, the current density, coulombic efficiency, and overall energy efficiency were higher.
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Molecular cloning, structure, and chromosomal localization of the mouse LIM/homeobox gene Lhx5
DOE Office of Scientific and Technical Information (OSTI.GOV)
Bertuzzi, S.; Sheng, Hui Z.; Westphal, H.
1996-09-01
Lhx5, the mouse ortholog of the Xenopus Xlim-5, is a LIM/homeobox gene expressed in the central nervous system during both embryonic development and adulthood. During development its domain of expression is mainly localized at the most anterior portion of the neural tube, and it precedes the morphological differentiation of the forebrain; for this reason we believe that Lhx5 could play an important role in forebrain patterning. Here we present the structural organization and the chromosomal localization of the Lhx5 gene. The gene is composed of five exons spanning more than 10 kb of genomic sequence. The first and second LIMmore » domains are encoded by the first and second exon, while the codons of the homeobox are split between the third and the fourth exons. The structure of Lhx5 is similar to that of other LIM/homeodomain proteins, Lxh1/lim1 and Lhx3/lim3, but differs from that of other LIM genes, such as mec3 and LMO1/Rbtn1, in which the codons for the LIM domains are interrupted by introns. We have mapped Lhx5 to the central region of mouse chromosome 5. 38 refs., 4 figs.« less
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Dhar, Bipro Ranjan; Elbeshbishy, Elsayed; Hafez, Hisham; Lee, Hyung-Sool
2015-12-01
An integrated dark fermentation and microbial electrochemical cell (MEC) process was evaluated for hydrogen production from sugar beet juice. Different substrate to inoculum (S/X) ratios were tested for dark fermentation, and the maximum hydrogen yield was 13% of initial COD at the S/X ratio of 2 and 4 for dark fermentation. Hydrogen yield was 12% of initial COD in the MEC using fermentation liquid end products as substrate, and butyrate only accumulated in the MEC. The overall hydrogen production from the integrated biohydrogen process was 25% of initial COD (equivalent to 6 mol H2/mol hexoseadded), and the energy recovery from sugar beet juice was 57% using the combined biohydrogen. Copyright © 2015 Elsevier Ltd. All rights reserved.
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The combinatorial control of alternative splicing in C. elegans
2017-01-01
Normal development requires the right splice variants to be made in the right tissues at the right time. The core splicing machinery is engaged in all splicing events, but which precise splice variant is made requires the choice between alternative splice sites—for this to occur, a set of splicing factors (SFs) must recognize and bind to short RNA motifs in the pre-mRNA. In C. elegans, there is known to be extensive variation in splicing patterns across development, but little is known about the targets of each SF or how multiple SFs combine to regulate splicing. Here we combine RNA-seq with in vitro binding assays to study how 4 different C. elegans SFs, ASD-1, FOX-1, MEC-8, and EXC-7, regulate splicing. The 4 SFs chosen all have well-characterised biology and well-studied loss-of-function genetic alleles, and all contain RRM domains. Intriguingly, while the SFs we examined have varied roles in C. elegans development, they show an unexpectedly high overlap in their targets. We also find that binding sites for these SFs occur on the same pre-mRNAs more frequently than expected suggesting extensive combinatorial control of splicing. We confirm that regulation of splicing by multiple SFs is often combinatorial and show that this is functionally significant. We also find that SFs appear to combine to affect splicing in two modes—they either bind in close proximity within the same intron or they appear to bind to separate regions of the intron in a conserved order. Finally, we find that the genes whose splicing are regulated by multiple SFs are highly enriched for genes involved in the cytoskeleton and in ion channels that are key for neurotransmission. Together, this shows that specific classes of genes have complex combinatorial regulation of splicing and that this combinatorial regulation is critical for normal development to occur. PMID:29121637
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Shore, Anna C.; Corcoran, Suzanne; Tecklenborg, Sarah; Coleman, David C.; O'Connell, Brian
2012-01-01
Sequence type 22 (ST22) methicillin-resistant Staphylococcus aureus (MRSA) harboring staphylococcal cassette chromosome mec (SCCmec) IV (ST22-MRSA-IV) has predominated in Irish hospitals since the late 1990s. Six distinct clones of community-associated MRSA (CA-MRSA) have also been identified in Ireland. A new strain of CA-MRSA, ST772-MRSA-V, has recently emerged and become widespread in India and has spread into hospitals. In the present study, highly similar MRSA isolates were recovered from seven colonized neonates in a neonatal intensive care unit (NICU) in a maternity hospital in Ireland during 2010 and 2011, two colonized NICU staff, one of their colonized children, and a NICU environmental site. The isolates exhibited multiantibiotic resistance, spa type t657, and were assigned to ST772-MRSA-V by DNA microarray profiling. All isolates encoded resistance to macrolides [msr(A) and mpb(BM)] and aminoglycosides (aacA-aphD and aphA3) and harbored the Panton-Valentine leukocidin toxin genes (lukF-PV and lukS-PV), enterotoxin genes (sea, sec, sel, and egc), and one of the immune evasion complex genes (scn). One of the NICU staff colonized by ST772-MRSA-V was identified as the probable index case, based on recent travel to India. Seven additional hospital and CA-ST772-MRSA-V isolates recovered from skin and soft tissue infections in Ireland between 2009 and 2011 exhibiting highly similar phenotypic and genotypic characteristics to the NICU isolates were also identified. The clinical details of four of these patients revealed connections with India through ethnic background or travel. Our study indicates that hospital-acquired and CA-ST772-MRSA-V is currently emerging in Ireland and may have been imported from India on several occasions. PMID:22189119
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High current density of 10.0-14.6 A/m2 and COD removal up to 96% were obtained in a microbial electrochemical cell (MEC) fed with digestate at hydraulic retention time (HRT) of 4d and 8d. Volatile fatty acids became undetectable in MEC effluent (HRT 8d), except for trivial acetat...
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Chen, Wei; Liu, Yongxia; Zhang, Limei; Gu, Xiaolong; Liu, Gang; Shahid, Muhammad; Gao, Jian; Ali, Tariq; Han, Bo
2017-01-01
Nocardia is one of the causing agents of bovine mastitis and increasing prevalence of nocardial mastitis in shape of serious outbreaks has been reported from many countries. However, the mechanisms by which this pathogen damages the bovine mammary epithelial cells (bMECs) is not yet studied. Therefore, this study was designed with the aim to evaluate the apoptotic effects elicited by Nocardia and to investigate the pathway by which the Nocardia induce apoptosis in bMECs. Clinical Nocardia cyriacigeorgica strain from bovine mastitis was used to infect the bMECs for different time intervals, viz . 1, 3, 6, 12, and 18 h, and then the induced effects on bMECs were studied using adhesion and invasion assays, release of lactate dehydrogenase (LDH), apoptosis analysis by annexin V and propidium iodide (PI) double staining, morphological, and ultrastructural observations under scanning electron microscope (SEM) and transmission electron microscope (TEM), mitochondrial transmembrane potential (ΔΨm) assay using flow cytometry, and the protein quantification of mitochondrial cytochrome c and caspase-9 and caspase-3 by western blotting. The results of this study showed that N. cyriacigeorgica possessed the abilities of adhesion and invasion to bMECs. N. cyriacigeorgica was found to collapse mitochondrial transmembrane potential, significantly ( p < 0.05) release mitochondrial cytochrome c and ultimately induce cell apoptosis. Additionally, it promoted casepase-9 ( p < 0.01) and casepase-3 ( p < 0.05) levels, significantly ( p < 0.01) increased the release of LDH and promoted DNA fragmentation which further confirmed the apoptosis. Furthermore, N. cyriacigeorgica induced apoptosis/necrosis manifested specific ultrastructure features under TEM, such as swollen endoplasmic reticulum, cristae degeneration, and swelling of mitochondria, vesicle formation on the cell surface, rupturing of cell membrane and nuclear membrane, clumping, fragmentation, and margination of chromatin. The present study is the first comprehensive insight into patho-morphological ultrastructural features of apoptosis/necrosis induced by N. cyriacigeorgica , which concluded that the clinical N. cyriacigeorgica induced apoptotic changes in the bMECs through mitochondrial-caspase dependent apoptotic pathway.
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Hydrogen Production by Geobacter Species and a Mixed Consortium in a Microbial Electrolysis Cell▿
Call, Douglas F.; Wagner, Rachel C.; Logan, Bruce E.
2009-01-01
A hydrogen utilizing exoelectrogenic bacterium (Geobacter sulfurreducens) was compared to both a nonhydrogen oxidizer (Geobacter metallireducens) and a mixed consortium in order to compare the hydrogen production rates and hydrogen recoveries of pure and mixed cultures in microbial electrolysis cells (MECs). At an applied voltage of 0.7 V, both G. sulfurreducens and the mixed culture generated similar current densities (ca. 160 A/m3), resulting in hydrogen production rates of ca. 1.9 m3 H2/m3/day, whereas G. metallireducens exhibited lower current densities and production rates of 110 ± 7 A/m3 and 1.3 ± 0.1 m3 H2/m3/day, respectively. Before methane was detected in the mixed-culture MEC, the mixed consortium achieved the highest overall energy recovery (relative to both electricity and substrate energy inputs) of 82% ± 8% compared to G. sulfurreducens (77% ± 2%) and G. metallireducens (78% ± 5%), due to the higher coulombic efficiency of the mixed consortium. At an applied voltage of 0.4 V, methane production increased in the mixed-culture MEC and, as a result, the hydrogen recovery decreased and the overall energy recovery dropped to 38% ± 16% compared to 80% ± 5% for G. sulfurreducens and 76% ± 0% for G. metallireducens. Internal hydrogen recycling was confirmed since the mixed culture generated a stable current density of 31 ± 0 A/m3 when fed hydrogen gas, whereas G. sulfurreducens exhibited a steady decrease in current production. Community analysis suggested that G. sulfurreducens was predominant in the mixed-culture MEC (72% of clones) despite its relative absence in the mixed-culture inoculum obtained from a microbial fuel cell reactor (2% of clones). These results demonstrate that Geobacter species are capable of obtaining similar hydrogen production rates and energy recoveries as mixed cultures in an MEC and that high coulombic efficiencies in mixed culture MECs can be attributed in part to the recycling of hydrogen into current. PMID:19820150
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Stelinski, L L; McGhee, P; Haas, M; Il'ichev, A L; Gut, L J
2007-08-01
Several application parameters of microencapsulated (MEC) sex pheromone formulations were manipulated to determine their impact on efficacy of disruption for codling moth, Cydia pomonella (L.); oriental fruit moth, Grapholita molesta (Busck); obliquebanded leafroller, Choristoneura rosaceana (Harris); and redbanded leafroller, Argyrotaenia velutinana (Walker). Depending on the experiment, the formulations evaluated were those formerly manufactured by 3M Canada (London, ON, Canada) or those that are currently available from Suterra LLC (Bend, OR). The efficacy of MEC formulations applied by air-blast sprayer evenly throughout the entire canopy of 2-3-m-tall apple (Malus spp.) trees was equivalent to treatments in which targeted applications of MECs were made to the lower or upper 1.5 m of the canopy (at equivalent overall rates) for oriental fruit moth and both leafroller species. The realized distribution of deposited microcapsules within the tree canopy corresponded well with the intended heights of application within the canopy. The additional coapplication of the pine resin sticker Nu-Film 17 increased efficacy but not longevity of MEC formulations for oriental fruit moth; this adjuvant had no added effects for codling moth or leafroller formulations. Increasing the rate of active ingredient (AI) per hectare by 20-30-fold (range 2.5-75.0 g/ha) did not improve the disruption efficacy of MECs for codling moth or either leafroller species when both low and high rates were applied at equivalent frequencies per season. A low-rate, high-frequency (nine applications per season) application protocol was compared with a standard protocol in which two to three applications were made per season, once before each moth generation for each species. The low-rate, high-frequency protocol resulted in equivalent or better disruption efficacy for each moth species, despite using two-fold less total AI per hectare per season with the former treatment. The low-rate, frequent-application protocol should make the use of MEC formulations of synthetic pheromone more economical and perhaps more effective.
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Taherikalani, Morovat; Mohammadzad, Mohammad Reza; Soroush, Setareh; Maleki, Mohammad Hossein; Azizi-Jalilian, Farid; Pakzad, Iraj; Sadeghifard, Nourkhoda; Asadollahi, Parisa; Emaneini, Mohammad; Monjezi, Aazam; Alikhani, Mohammad Yousef
2016-04-01
Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important pathogens worldwide and compared to other staphylococcal species that are associated with higher mortality rate. A total of 500 Staphylococcus spp. was collected from selected hospitals in Ilam, Kermanshah, Khorram Abad and Hamadan cities and, via phenotypic and genotypic methods, was assessed to find MRSA. The presence or absence of prevalent antibiotic resistance genes and virulence genes was evaluated among MRSA isolates, using polymerase chain reaction (PCR) method, and then the SCCmec typing of these isolates was assayed by multiplex PCR. A total of 372 (74.4%) Stapylococcus spp. isolates were identified as S. aureus, among which 200 (53.8%) possessed the mecA gene and were distinguished as MRSA. All of MRSA isolates contained blaZ gene. The frequency of ermA and ermC genes among erythromycin-resistant MRSA isolates was 21.6% and 66.7%, respectively. The frequency of the virulence genes eta, hla and sea among MRSA isolates was 10%, 80.5% and 100%, respectively. SCCmec type IV accounted for 30.6% of the MRSA isolates and SCCmec type III, SCCmec type II and SCCmec type I accounted for 30%, 22% and 17.5% of the isolates, respectively. The antibiotic resistance genes and the virulence genes of blaZ, hla, sea, eta and ermC had high frequencies among the MRSA isolates. This study showed that the antibiotic resistance genes had higher frequencies among SCCmec types I and IV, which confirms the previous reports in this field.
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McKee, Sherry A; Weinberger, Andrea H; Harrison, Emily L R; Coppola, Sabrina; George, Tony P
2009-12-01
Individuals with schizophrenia have higher plasma nicotine levels in comparison to non-psychiatric smokers, even when differences in smoking are equated. This difference may be related to how intensely cigarettes are smoked but this has not been well studied. Mecamylamine (MEC), a non-competitive nicotinic acetylcholine receptor (nAChR) antagonist, which has been shown to increase ad-lib smoking and to affect smoking topography, was used in the current study as a pharmacological probe to increase our understanding of smoking behavior, smoking topography, and resulting nicotine levels in smokers with schizophrenia. This preliminary study used a within-subject, placebo-controlled design in smokers with schizophrenia (n=6) and healthy control smokers (n=8) to examine the effects of MEC (10mg/day) on ad-lib smoking behavior, topography, nicotine levels, and tobacco craving across two smoking deprivation conditions (no deprivation and 12-h deprivation). MEC, compared to placebo, increased the number of cigarettes smoked and plasma nicotine levels. MEC increased smoking intensity and resulted in greater plasma nicotine levels in smokers with schizophrenia compared to controls, although these results were not consistent across deprivation conditions. MEC also increased tobacco craving in smokers with schizophrenia but not in control smokers. Our results suggest that antagonism of high-affinity nAChRs in smokers with schizophrenia may prompt compensatory smoking, increasing the intensity of smoking and nicotine exposure without alleviating craving. Further work is needed to assess whether nicotine levels are directly mediated by how intensely the cigarettes are smoked, and to confirm whether this effect is more pronounced in smokers with schizophrenia.
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Lippert, Frank
2012-10-01
This study investigated the effects of lesion baseline characteristics and different strontium (Sr) to calcium (Ca) ratios in plaque fluid-like solutions (PF) on lesion de- and remineralization. Caries lesions were formed in enamel using three protocols: methylcellulose acid gel (MeC) and partially saturated lactic acid solutions containing carboxymethylcellulose (CMC) or not (SOLN). Lesions were exposed to PF with four distinct Sr:Ca molar ratios (0:1/3:1:3), but otherwise identical composition and total Sr+Ca molarity, for seven days. Lesions were characterized using transverse microradiography (TMR) at baseline and post-treatment. At baseline, MeC and CMC had similar integrated mineral loss values, whereas SOLN lesions were more demineralized. All lesions showed significant differences in their mineral distributions, with CMC and SOLN having lower R values (integrated mineral loss to lesion depth ratio) than MeC. Post-PF exposure, no interaction was found between lesion type and Sr:Ca ratio. Within lesion type, MeC demineralized, whereas CMC and SOLN exhibited some remineralization, with the differences between MeC and the other lesion types being of statistical significance. Within Sr:Ca ratio, the 1:3 ratio exhibited some remineralization whereas other groups tended to demineralize. Only the difference between groups SrCa1/3 and SrCa0 was of statistical significance. In summary, both lesion baseline characteristics and Sr:Ca ratio were shown to effect lesion de- and remineralization. Under the conditions of the study, high-R lesions are more prone to demineralize under PF-like conditions than low-R lesions. In addition, partial Sr substitution for Ca in PF was shown to enhance lesion remineralization. Copyright © 2012 Elsevier Ltd. All rights reserved.
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Chen, Melissa J; Gaffield, Mary E; Kiarie, James
2016-09-28
Given recent updates to the postpartum contraception recommendations in the fifth edition of the Medical Eligibility Criteria for Contraceptive Use (MEC), published by the World Health Organization (WHO), the purpose of this qualitative study was to assess the extent to which national family planning policies in sub-Saharan African countries are in agreement with the WHO MEC, particularly with regard to postpartum contraceptive use. WHO headquarters sent questionnaires to country-level focal points to complete with their Ministry of Health counterparts. Between February and May 2016, 23 of 32 (72%) surveys were completed. All respondents reported that their countries had used the MEC document in the past, with most reporting that they had used the guidance as a reference (n = 20, 87%), for training purposes (n = 19, 83%), to change clinical practices (n = 17, 74%), and to develop national policies (n = 16, 70%). While many respondents (16, 70%) indicated their countries already include immediate postpartum intrauterine device insertion among breastfeeding women in their family planning policies, few reported currently allowing use of progestogen-only pills (n = 8, 35%) or implants (n = 8, 35%) during the immediate postpartum period (i.e., less than 48 hours after delivery) for breastfeeding women. A higher percentage of respondents indicated their countries allowed breastfeeding women the option of progestogen-only pills (n = 16, 70%) and implants (n = 13, 57%) between 48 hours and 6 weeks postpartum. Findings from this baseline assessment suggest that many countries may benefit from training and policy formulation support to adapt both new WHO MEC updates as well as existing recommendations from previous MEC revisions into national family planning guidelines. © Chen et al.
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Yu, Shiying; Burke, Thomas A; Chan, Alexandre; Kim, Hoon-Kyo; Hsieh, Ruey Kuen; Hu, Xichun; Liang, Jin-Tung; Baños, Ana; Spiteri, Carmel; Keefe, Dorothy M K
2015-01-01
This paper reports prescribing patterns for prophylaxis of chemotherapy-induced nausea and vomiting (CINV) after highly or moderately emetogenic chemotherapy (HEC or MEC) for cancer in six Asia Pacific countries. In a prospective noninterventional study, 31 sites in Australia, China, India, Singapore, South Korea, and Taiwan recorded details of CINV prophylaxis for the acute phase (first 24 h) and delayed phase (days 2-5) after single-day HEC or MEC for adult patients. Additional information on CINV prophylactic medications was collected from 6-day patient diaries. Primary antiemetic therapies were defined as corticosteroids, the 5-hydroxytryptamine-3 receptor antagonists (5HT3-RAs), and neurokinin-1 receptor antagonists (NK1-RAs). Evaluable patients in cycle 1 numbered 648 (318 [49%] HEC and 330 [51%] MEC) of mean (SD) age of 56 (12) years, including 58% women. For the acute phase after HEC, overall (and country range), 96% (91-100%) of patients received a 5HT3-RA, 87% (70-100%) a corticosteroid, and 43% (0-91%) an NK1-RA. CINV prophylaxis for the HEC delayed phase was more variable: including 22% (7-65%) 5HT3-RA, 52% (12-93%) corticosteroid, and 46% (0-88%) NK1-RA. For the MEC acute phase, 97% (87-100%) of patients received 5HT3-RA and 86% (73-97%) a corticosteroid. For the MEC delayed phase, 201 patients (61%) received a primary antiemetic, including 5HT3-RA (41%), corticosteroid (37%), and/or NK1-RA (4%). The 5HT3-RAs were prescribed consistently in all countries, while prescribing of other antiemetic therapies was variable, and corticosteroids were under-prescribed for CINV prophylaxis, particularly in the delayed phase.
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Lee, Jaewoong; Park, Yeon Joon; Park, Dong Jin; Park, Kang Gyun; Lee, Hae Kyung
2017-01-01
We evaluated the performance of the BD MAX StaphSR Assay (SR assay; BD, USA) for direct detection of Staphylococcus aureus and methicillin resistance not only in S. aureus but also in coagulase-negative Staphylococci (CNS) from positive blood cultures. From 228 blood culture bottles, 103 S. aureus [45 methicillin-resistant S. aureus (MRSA), 55 methicillin-susceptible S. aureus (MSSA), 3 mixed infections (1 MRSA+Enterococcus faecalis, 1 MSSA+MRCNS, 1 MSSA+MSCNS)], and 125 CNS (102 MRCNS, 23 MSCNS) were identified by Vitek 2. For further analysis, we obtained the cycle threshold (Ct) values from the BD MAX system software to determine an appropriate cutoff value. For discrepancy analysis, conventional mecA/mecC PCR and oxacillin minimum inhibitory concentrations (MICs) were determined. Compared to Vitek 2, the SR assay identified all 103 S. aureus isolates correctly but failed to detect methicillin resistance in three MRSA isolates. All 55 MSSA isolates were correctly identified by the SR assay. In the concordant cases, the highest Ct values for nuc, mecA, and mec right-extremity junction (MREJ) were 25.6, 22, and 22.2, respectively. Therefore, we selected Ct values from 0-27 as a range of positivity, and applying this cutoff, the sensitivity/specificity of the SR assay were 100%/100% for detecting S. aureus, and 97.9%/98.1% and 99.0%/95.8% for detecting methicillin resistance in S. aureus and CNS, respectively. We propose a Ct cutoff value for nuc/mec assay without considering MREJ because mixed cultures of MSSA and MRCNS were very rare (0.4%) in the positive blood cultures.
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Cilz, Nicholas I; Lei, Saobo
2017-05-01
In the brain, histamine (HA) serves as a neuromodulator and a neurotransmitter released from the tuberomammillary nucleus (TMN). HA is involved in wakefulness, thermoregulation, energy homeostasis, nociception, and learning and memory. The medial entorhinal cortex (MEC) receives inputs from the TMN and expresses HA receptors (H 1 , H 2 , and H 3 ). We investigated the effects of HA on GABAergic transmission in the MEC and found that HA significantly increased the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) with an EC 50 of 1.3 µM, but failed to significantly alter sIPSC amplitude. HA-induced increases in sIPSC frequency were sensitive to tetrodotoxin (TTX), required extracellular Ca 2+ , and persisted when GDP-β-S, a G-protein inactivator, was applied postsynaptically via the recording pipettes, indicating that HA increased GABA release by facilitating the excitability of GABAergic interneurons in the MEC. Recordings from local MEC interneurons revealed that HA significantly increased their excitability as determined by membrane depolarization, generation of an inward current at -65 mV, and augmentation of action potential firing frequency. Both H 1 and H 2 receptors were involved in HA-induced increases in sIPSCs and interneuron excitability. Immunohistochemical staining showed that both H 1 and H 2 receptors are expressed on GABAergic interneurons in the MEC. HA-induced depolarization of interneurons involved a mixed ionic mechanism including activation of a Na + -permeable cation channel and inhibition of a cesium-sensitive inward rectifier K + channel, although HA also inhibited the delayed rectifier K + channels. Our results may provide a cellular mechanism, at least partially, to explain the roles of HA in the brain. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.
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Hoopes, Andrea J; Simmons, Katharine B; Godfrey, Emily M; Sucato, Gina S
2017-04-01
The US Medical Eligibility Criteria for Contraceptive Use (MEC) and US Selected Practice Recommendations for Contraceptive Use (SPR) provide evidence-based guidance to safely provide contraception counseling and services. Both documents were updated in 2016 and are endorsed by the North American Society for Pediatric and Adolescent Gynecology. The purpose of this mini-review is to highlight updates to the US MEC and US SPR that are most relevant to health care providers of adolescents to support dissemination and implementation of these evidence-based best practices. This document is intended to highlight these changes and to complement, not replace, the detailed practice guidance within the US MEC and US SPR. Copyright © 2017 North American Society for Pediatric and Adolescent Gynecology. Published by Elsevier Inc. All rights reserved.
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Richter, Sandra S.; Heilmann, Kristopher P.; Dohrn, Cassie L.; Riahi, Fathollah; Costello, Andrew J.; Kroeger, Jennifer S.; Biek, Donald; Critchley, Ian A.; Diekema, Daniel J.; Doern, Gary V.
2011-01-01
A Staphylococcus aureus surveillance program was initiated in the United States to examine the in vitro activity of ceftaroline and epidemiologic trends. Susceptibility testing by Clinical and Laboratory Standards Institute broth microdilution was performed on 4,210 clinically significant isolates collected in 2009 from 43 medical centers. All isolates were screened for mecA by PCR and evaluated by pulsed-field gel electrophoresis. Methicillin-resistant S. aureus (MRSA) were analyzed for Panton-Valentine leukocidin (PVL) genes and the staphylococcal cassette chromosome mec (SCCmec) type. All isolates had ceftaroline MICs of ≤2 μg/ml with an MIC50 of 0.5 and an MIC90 of 1 μg/ml. The overall resistance rates, expressed as the percentages of isolates that were intermediate and resistant (or nonsusceptible), were as follows: ceftaroline, 1.0%; clindamycin, 30.2% (17.4% MIC ≥ 4 μg/ml; 12.8% inducible); daptomycin, 0.2%; erythromycin, 65.5%; levofloxacin, 39.9%; linezolid, 0.02%; oxacillin, 53.4%; tetracycline, 4.4%; tigecycline, 0%; trimethoprim-sulfamethoxazole, 1.6%; vancomycin, 0%; and high-level mupirocin, 2.2%. The mecA PCR was positive for 53.4% of the isolates. The ceftaroline MIC90s were 0.25 μg/ml for methicillin-susceptible S. aureus and 1 μg/ml for MRSA. Among the 2,247 MRSA isolates, 51% were USA300 (96.9% PVL positive, 99.7% SCCmec type IV) and 17% were USA100 (93.4% SCCmec type II). The resistance rates for the 1,137 USA300 MRSA isolates were as follows: erythromycin, 90.9%; levofloxacin, 49.1%; clindamycin, 7.6% (6.2% MIC ≥ 4 μg/ml; 1.4% inducible); tetracycline, 3.3%; trimethoprim-sulfamethoxazole, 0.8%; high-level mupirocin, 2.7%; daptomycin, 0.4%; and ceftaroline and linezolid, 0%. USA300 is the dominant clone causing MRSA infections in the United States. Ceftaroline demonstrated potent in vitro activity against recent S. aureus clinical isolates, including MRSA, daptomycin-nonsusceptible, and linezolid-resistant strains. PMID:21709080
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Sued, Bruna Pinto Ribeiro; Pereira, Paula Marcele Afonso; Faria, Yuri Vieira; Ramos, Juliana Nunes; Binatti, Vanessa Batista; Santos, Kátia Regina Netto Dos; Seabra, Sérgio Henrique; Hirata, Raphael; Vieira, Verônica Viana; Mattos-Guaraldi, Ana Luíza; Pereira, José Augusto Adler
2017-03-01
The association between Staphylococcus haemolyticus and severe nosocomial infections is increasing. However, the extent to which fomites contribute to the dissemination of this pathogen through patients and hospital wards remains unknown. In the present study, sphygmomanometers and thermometers were evaluated as potential fomites of oxacillin-resistant S. haemolyticus (ORSH). The influence of oxacillin and vancomycin on biofilm formation by ORSH strains isolated from fomites was also investigated. The presence of ORSH on swabs taken from fomite surfaces in a Brazilian hospital was assessed using standard microbiological procedures. Antibiotic susceptibility profiles were determined by the disk diffusion method, and clonal distribution was assessed in pulsed-field gel electrophoresis (PFGE) assays. Minimum inhibitory concentrations (MICs) of oxacillin and vancomycin were evaluated via the broth microdilution method. Polymerase chain reaction (PCR) assays were performed to detect the mecA and icaAD genes. ORSH strains grown in media containing 1/4 MIC of vancomycin or oxacillin were investigated for slime production and biofilm formation on glass, polystyrene and polyurethane catheter surfaces. ORSH strains comprising five distinct PFGE types were isolated from sphygmomanometers (n = 5) and a thermometer (n = 1) used in intensive care units and surgical wards. ORSH strains isolated from fomites showed susceptibility to only linezolid and vancomycin and were characterised as multi-drug resistant (MDR). Slime production, biofilm formation and the survival of sessile bacteria differed and were independent of the presence of the icaAD and mecA genes, PFGE type and subtype. Vancomycin and oxacillin did not inhibit biofilm formation by vancomycin-susceptible ORSH strains on abiotic surfaces, including on the catheter surface. Enhanced biofilm formation was observed in some situations. Moreover, a sub-lethal dose of vancomycin induced biofilm formation by an ORSH strain on polystyrene. Sphygmomanometers and thermometers are fomites for the transmission of ORSH. A sub-lethal dose of vancomycin may favor biofilm formation by ORSH on fomites and catheter surfaces.