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The present invention is directed to a rapid and sensitive method for detecting Mycoplasmahominis using M. hominis-specific probes, oligonucleotides or antibodies. In particular a target sequence can be amplified by in vitro nucleic acid amplification techniques, detected by nucleic acid hybridization using the subject probes and oligonucleotides or detected by immunoassay using M. hominis-specific antibodies. M. hominis-specific nucleic acids which do not recognize or hybridize to genomic nucleic acid of other Mycoplasma species are also provided.
DelVecchio, Vito G. (Scranton, PA); Gallia, Gary L. (Philadelphia, PA); McCleskey, Ferne K. (San Antonio, TX)
Cells of two strains of Mycoplasmahominis growing in liquid medium on a glass surface were observed continuously, and cinematographic pictures were taken. Most of the observed structures showed reversible changes of their shape, suggesting the presence of contractile material in membrane or cytoplasma. The frequency and speed of such variations were measured. The deformations seem to be related to multiplication. The mechanisms of these phenomena are unknown. Images
Bredt, W.; Heunert, H. H.; Hofling, K. H.; Milthaler, Brigitte
Cells of two strains of Mycoplasmahominis growing in liquid medium on a glass surface were observed continuously, and cinematographic pictures were taken. Most of the observed structures showed reversible changes of their shape, suggesting the presence of contractile material in membrane or cytoplasma. The frequency and speed of such variations were measured. The deformations seem to be related to multiplication. The mechanisms of these phenomena are unknown. PMID:4570775
Bredt, W; Heunert, H H; Höfling, K H; Milthaler, B
Two cases of Mycoplasmahominis meningitis in the newborn are described. These infants demonstrate the need to consider M. hominis as a cause of neonatal meningitis, especially if the Gram stain is negative, conventional cultures yield no growth, and there is a history of maternal infection. CSF cultures on appropriate medium can quickly confirm the diagnosis.
M Gewitz; R Dinwiddie; L Rees; O Volikas; T Yuille; B OConnell; W C Marshall
In one year Mycoplasmahominis was isolated from 8 out of 250 clinically infected eyes of newborn infants. This infection occurred in only a small proportion of babies whose mothers carried the organism in the vagina. Probably mycoplasma infection of the eye in neonates is commoner than is realized.
Arginine deiminase is a promising anticancer drug active against melanoma, hepatocarcinoma and other tumors. Recombinant strains of Escherichia coli that express arginine deiminase from pathogenic bacteria Mycoplasma have been developed. However, production costs of heterologous arginine deiminase are high due to use of an expensive inducer and extraction buffer, as well as using diluted culture for enzyme induction. We report on a new advanced protocol for Mycoplasmahominis arginine deiminase expression, extraction and renaturation. The main improvements include manipulation with dense suspensions of E. coli, use of lactose instead of isopropyl ?-d-1-thiogalactopyranoside as an inducer and a cheaper but not less efficient buffer for solubilization of arginine deiminase inclusion bodies. In addition, supplementation of the storage culture medium with glucose and substrate (arginine) significantly stabilized the recombinant arginine deiminase producer. Homogenous preparations of recombinant arginine deiminase were obtained using anion-exchange and hydrophobic chromatography. The purified enzyme retained a specific activity of 30-34U/mg for 12 months when stored at 4°C in 20mM sodium phosphate buffer pH 7.2 containing 1M NaCl. PMID:23928331
Both Mycoplasmahominis and Trichomonas vaginalis utilize arginine as an energy source via the arginine dihydrolase (ADH) pathway. It has been previously demonstrated that M. hominis forms a stable intracellular relationship with T. vaginalis; hence, in this study we examined the interaction of two localized ADH pathways by comparing T. vaginalis strain SS22 with the laboratory-generated T. vaginalis strain SS22-MOZ2 infected with M. hominis MOZ2. The presence of M. hominis resulted in an approximately 16-fold increase in intracellular ornithine and a threefold increase in putrescine, compared with control T. vaginalis cultures. No change in the activity of enzymes of the ADH pathway could be demonstrated in SS22-MOZ2 compared with the parent SS22, and the increased production of ornithine could be attributed to the presence of M. hominis. Using metabolic flow analysis it was determined that the elasticity of enzymes of the ADH pathway in SS22-MOZ2 was unchanged compared with the parent SS22; however, the elasticity of ornithine decarboxylase (ODC) in SS22 was small, and it was doubled in SS22-MOZ2 cells. The potential benefit of this relationship to both T. vaginalis and M. hominis is discussed.
Because several reports have suggested that bacterial vaginosis causes premature labour and early rupture of the fetal membranes, the presence of a bacterial flora that causes bacterial vaginosis is thought to be a risk factor for premature labour. The present study investigated two patients with premature delivery and intra-uterine Mycoplasmahominis infection. In microbiological studies, Gram's staining of amniotic fluids revealed numerous neutrophils and epithelial cells but no micro-organisms. Culture of amniotic fluid before antibiotic therapy yielded only M. hominis under anaerobic conditions; aerobic culture was negative. Vaginal discharge taken on the day of delivery yielded no growth in case 1 and M. hominis and Enterococcus faecalis in case 2. Maternal sera showed specific antibodies to M. hominis by ELISA and immunoblotting. As no possible cause of premature labour other than M. hominis infection was detected, it is concluded that the intra-uterine M. hominis infection was associated with premature labour in these patients. PMID:9879962
Shimada, M; Kotani, T; Sameshima, H; Nagamine, Y; Kodama, Y; Ikenoue, T; Kenri, T; Sasaki, T; Ohtaki, S
Mycoplasmahominis is a commensal of the genitourinary tract. It mostly causes infections to associated structures of this system; however,\\u000a occasionally it is a pathogen in nongenitourinary tract infections. Since, M. hominisstrains require special growth conditions and cannot be Gram stained, they may be missed or delay diagnosis. This report\\u000a describes a deep wound infection caused by M. hominis
Matthijs R. Krijnen; Thecla Hekker; Johan Algra; Paul I. J. M. Wuisman; Barend J. Van Royen
Here we examined the involvement of Mycoplasmahominis in the formation of biofilms by uropathogenic Escherichia coli (UPEC) strain CFT073. Initially, we thought that M. hominis does not affect the fitness of UPEC, including the growth and production of signaling molecules, such as autoinducer-2 and indole. We found, however, that the presence of M. hominis significantly decreased the degree of biofilm formation by UPEC CFT073 (approximately a 60% reduction for 10(5) ccu/mL of M. hominis as compared with UPEC alone). We also found that it had a slight effect in inhibiting the attachment and cytotoxicity of UPEC CFT073. These findings are specific to these UPEC strains rather than to enterohemorrhagic E. coli (EHEC) strains, found in normal intestinal flora. In addition, we performed whole-transcriptome profiling and quantitative real-time polymerase chain reaction (qRT-PCR) analysis. This indicated that the PhoPQ system and the anti-termination protein (encoded by ybcQ) were involved in the reduction of biofilm formation by M. hominis (corroborated by qRT-PCR). Furthermore, our results indicate that M. hominis raises the degree of transcription of toxin genes, including hha and pasT. Hence, we suggest a possible role of M. hominis in affecting the formation of biofilms by UPEC in the urinary tract. PMID:24096662
Oh, Sangnam; Go, Gwang-Woong; Choi, Nag-Jin; Oh, Sejong; Kim, Younghoon
The parasite Trichomonas vaginalis causes one of the most common non-viral sexually transmitted infections in humans. Mycoplasmas are frequently found with\\u000a trichomonads but the consequences of this association are not yet known. In the present study, the effects of T. vaginalis harboring M. hominis on human vaginal epithelial cells and on MDCK cells are described. The results were analyzed by
R. G. Vancini; A. Pereira-Neves; R. Borojevic; M. Benchimol
Thirty young adult mice, of strain BALB/c, treated previously with progesterone, were inoculated intravaginally (10 mice) or directly into the uterus (10 mice) with Mycoplasma pulmonis and 10 mice remained uninoculated. Ten mice not treated with the hormone were also inoculated intrauterinely with M. pulmonis. The same numbers of mice treated with oestradiol were inoculated in the same ways with M. hominis. Vaginal swab specimens were obtained from all mice 7, 14 and 28 days after inoculation and samples of genital tract tissue were collected from pairs of mice at the same time intervals. Large numbers of M. pulmonis and M. hominis organisms were isolated from the vagina throughout the course of the experiments and they were cultured also from the cervix and uterine horns. Mycoplasma-like bodies were demonstrated by scanning electron microscopy in the cervix and in the uterus, but neither mycoplasmal species was found attached to vaginal epithelium. The results of silver-enhanced immunogold labelling in conjunction with scanning microscopy provided assurance that the mycoplasma-like bodies were, in fact, mycoplasmas. The importance of hormone treatment was indicated by the diminished susceptibility of untreated mice to M. pulmonis and the almost complete insusceptibility to M. hominis, shown by culture and scanning electron microscopy. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10
Furr, P. M.; Sarathchandra, P.; Hetherington, C. M.; Taylor-Robinson, D.
A polymerase chain reaction (PCR)-based assay was developed for the detection ofMycoplasmahominis. This assay generates a 152-bp PCR product which was part of an initial 471-bpM. hominisgenomic DNA fragment. The 471-bp DNA fragment was shown by hybridization analysis to be unique forM. hominis. The PCR assay can amplify as few as 18 molecules of target DNA. This diagnostic assay
Gary L. Gallia; Joseph M. Petroziello; Joseph M. Brogan; Ferne K. McCleskey; Vito G. DelVecchio
The molecular basis for the size and antigenic diversity of the variable adherence-associated (Vaa) antigen, a major surface protein and a putative adhesin (of Mycoplasmahominis, is described. Size-variant alleles of the single-copy vaa gene encode abundant surface lipoproteins containing one to four nearly identical, tandem repetitive units of 121 amino acids in the central region of the mature Vaa product. Gain or loss of central repeats in vaa genes gives rise to distinct size-variant Vaa antigens in clonal populations of this organism. The N-terminal and repeat regions of Vaa contain highly conserved sequences, while the C-terminal region, implicated as the adherence-mediating module, is highly variable and divergent among different strains of this pathogen. Sequence variation in this region may underlie the strain-dependent binding of some monoclonal antibodies to Vaa products. The Vaa antigen is expressed in vivo during chronic, active arthritis associated with M. hominis infection and is highly immunogenic in the human host. Size variation and C-terminal antigenic divergence of Vaa could affect the adherence of M. hominis and evasion of antibody-mediated immunity, thereby contributing to the organism's adaptive capability in the human host. Variation in vaa genes reveals a distinct pattern of mutations generating mycoplasma surface variation.
We used simulated blood cultures inoculated with clinical isolates of Mycoplasmahominis to determine whether liquid media of the BacT/ALERT (Organon Teknika, Durham, N.C.) will support growth of this fastidious organism and whether its presence can generate a positive signal with the instrument. Viability of clinical isolates of M. hominis was maintained for 7 days in BacT/ALERT media, and organisms were able to multiply when 1% gelatin was added to neutralize the mycoplasmastatic effects of the sodium polyanetholsulfonate anticoagulant. Without the addition of gelatin to BacT/ALERT bottles, the mycoplasmas declined in numbers or became completely nonviable. Mycoplasmal growth was further enhanced in BacT/ALERT PF both supplemented with gelatin, arginine, and DNA in comparison to broth with only gelatin added. No BacT/ALERT bottles containing M. hominis in simulated blood cultures were flagged positive by the instrument, despite growth of microorganisms of up to 107 CFU/ml after incubation for up to 7 days, suggesting that inadequate CO2 production or some other mechanism prevents the instrument from recognizing the presence of the organism and its metabolic products. The fastidious cultivation requirements and relatively slow growth of M. hominis warrant that dependence on automated systems and techniques designed to detect conventional bacteria will not be reliable for recovery of M. hominis and that specialized media and incubation conditions designed for optimum cultivation of mycoplasmas should be employed when this organism is suspected on clinical grounds.
BACKGROUND: Mycoplasmahominis is associated with pelvic inflammatory disease, bacterial vaginosis, post partum fever, sepsis and infections of the central nervous system often leading to serious conditions. Association with development of female infertility has also been suggested, but different publications present different results. We developed a sensitive and fast diagnostic real-time PCR to test clinical samples from women undergoing laparoscopic
We developed a multiplex polymerase chain reaction (M-PCR) assay to simultaneously detect Trichomonas vaginalis, Mycoplasmahominis, and Ureaplasma urealyticum. The test is extremely specific and has a sensitivity of 10 cells for T. vaginalis and U. urealyticum and of 1 cell for M. hominis. The technique was validated on vaginal swabs from 240 women presenting symptoms of vaginitis, and results
Mycoplasmahominis most frequently causes diseases of the genitourinary tract. Extragenital infections are uncommon, with almost all occurring in immunosuppressed persons or those predisposed due to trauma or surgery. We present the case of a previously well man who developed an M. hominis-associated parapharyngeal abscess following acute Epstein-Barr virus infection.
Mycoplasmahominis is an opportunistic human mycoplasma species that can be either commensal or pathogenic. Its detection by culture is considered to comprise the reference technique. Previously reported PCR techniques target the 16S rRNA or the gap gene, although sequence variations among clinical isolates may lead to variations in clinical sensitivity. The present study aimed to develop a specific TaqMan quantitative real-time PCR assay, targeting a gene conserved in all M. hominis isolates, and to compare it with quantitative culture. With the knowledge of the M. hominis PG21 genome sequence, the yidC gene, encoding a membrane protein translocase, was chosen as target. Its intraspecies heterogeneity was checked at the nucleotide level using 31 reference or clinical strains. The limit of detection, the analytical specificity and the reproducibility of the assay were assessed. Moreover, PCR and culture results were compared using 153 urogenital specimens. The limit of detection was seven copies/?L. The analytical specificity was 100%, with good inter- and intra-assay reproducibility. Among the 153 urogenital specimens, the yidC PCR and culture allowed detection of 55 and 45 M. hominis-positive samples, respectively. Comparison of the bacterial load among the 45 specimens found to be M. hominis-positive by both techniques revealed a statistically significant association between the quantitative results obtained. In conclusion, we developed a specific, sensitive and reproducible real-time PCR to detect all M. hominis clinical isolates. This PCR was shown to have higher sensitivity than culture, although both methods were correlated for quantification of M. hominis loads in urogenital specimens. PMID:20298269
Ureaplasma urealyticum (UU) and Mycoplasmahominis (MH) have been detected in the urine of women with systemic lupus erythematosus (SLE). We evaluated the presence of these mycoplasma in the endocervix of women presenting SLE. A total of 40 SLE patients (mean age 40.2 years), and 51 healthy women (mean age 30.9 years), were studied. Endocervical swabs were cultured in specific liquid media for MH or UU, detected by a quantitative color assay, and considered positive at >10(3) dilutions. Statistical analysis was performed using the two-tailed Fisher test. UU was detected in 52.5 % of patients and in 11.8% of controls (p= 0.000059). MH was detected in 20% of patients and 2% controls (p=0.003905). Both mycoplasmas were detected in 7.3% patients and 0% controls (p<0.000001). The results reported here corroborate the association of the mycoplasma infection and SLE. Thus, these agents may stimulate the production of autoreactive clones. PMID:11460209
Machado, A A; Zorzi, A R; Gléria, A E; Donadi, E A
Genital mycoplasmas play an important role in genitourinary tract disease. The purpose of this study was to isolate M. hominis and U. urealyticum from vaginal and throat swabs and urine from women sexually active or not. Samples were taken from women with (cases) or without (controls) genitourinary tract disease and were dipped inoculated into 1 ml of E broth with arginine or urea and ten-fold dilutions were done. Samples were incubated at 37 degrees C until phenol red indicator changed to color purple. Identification of species was done by polymerase chain reaction technique. M. hominis was identified with oligonucleotides that correspond to the nucleotide sequence of 16S rRNA gene and U. urealyticum was identified with oligonucleotides that correspond to the nucleotide sequence of the urease gene (Blanchard et al.). There was no statistical difference (X2 P > .05) between isolation percentages from vaginal swabs, while there was statistical difference between urine samples. These mycoplasmas were isolated in higher percentages from pubertal girls and were recovered until the fifth ten-fold dilution both from vaginal swabs and urine. For throat swabs they were only recovered from sexually active women. PMID:8986107
Gil-Juárez, C; Calderón, B A; Montero, J; Yáńez, A; Cedillo, L
Mycoplasmahominis and Ureaplasma urealyticum are associated with chorioamnionitis, preterm delivery and pelvic inflammatory disease. The aim of this study was to evaluate the possible risk factors of co-colonization by M. hominis in patients already colonized by U. urealyticum and compare demographic parameters, vaginal pH and microbiota of women colonized by U. urealyticum or M. hominis. A total of 452 patients positive for U. urealyticum or M. hominis were analysed, 421 (93.1%) of whom were positive for U. urealyticum and 31 (6.9%) for M. hominis. Patients positive for M. hominis compared to patients positive for U. urealyticum were more frequently colonized by Gardnerella vaginalis (71% vs 18.5%; p 0.0001), less frequently by lactobacilli (16.1% vs 61.5%; p 0.0001), and more frequently had a pH value higher than 4.5 (96.8% vs 57%; p 0.0001), all conditions associated to bacterial vaginosis (BV). Logistic regression analysis showed that only G. vaginalis colonization and pH higher than 4.5 were independently related to M. hominis colonization (respectively p 0.0001 and p 0.016). Thus, in women colonized by U. urealyticum, BV is an independent risk factor for M. hominis co-colonization. PMID:24008852
Mycoplasmahominis--one of the widely spread mycoplasmas (class Mollicutes), associated with the socially significant human diseases and contamination of cell cultures. The solution of the problem on controlling M. hominis infections is connected with determination of the molecular basis, responsible for mechanisms of bacterium survival under unfavorable conditions. As a result of proteomic approach (2-DIGE and MALDI TOF/TOF MS) for the first time, 53 M. hominis PG37 proteins were detected, different abundance of which occurred at cultivating the bacterium under stress (starvation and low temperature) conditions. According to the classification of proteins by functional category (clusters of orthologous groups of proteins--COG), 47 of the 53 proteins of the mycoplasma are involved in the fundamental cellular and biochemical processes--translation (12; 22.64%), transcription (2; 3.77%), posttranslational modification (7; 13.20%), cell cycle control (2; 3.77%), energy production and conversion (6; 11.32%), carbohydrate transport and metabolism (3; 5.66%), amino acid transport and metabolism (8; 15.09%), nucleotide transport and metabolism (6; 11.32%), inorganic ion transport and metabolism (1; 1.89%). The functions of six proteins (11.32%) have not been found; 24 proteins (45.28%) are the factors of bacterium virulence. M. hominis PG37 proteins, the expression modulation of which arises under the unfavorable environmental conditions, are the components of adaptation mechanisms of the mycoplasma to the stressors and potential targets for controlling infections caused by this bacterium. PMID:22393789
Chernov, V M; Chernova, O A; Baranova, N B; Gorshkov, O V; Medvedeva, E S; Sha?mardanova, G F
Background In Mycoplasmahominis, a facultative human pathogen of the human genital tract, OppA, the substrate-binding domain of the oligopeptide permease, is a multifunctional protein involved in nutrition uptake, cytoadhesion and hydrolysis of extracellular ATP. Results To map the function-related protein regions the ATPase activity and adhesive behavior of OppA mutants were analyzed. Mutations of the Walker BA motifs resulted in an inhibition of up to 8% of the OppA ATPase activity, whereas deletion of the N-terminal CS1 or the CS2 region, structural motifs that are conserved in bacterial OppA proteins, reduced ATPase activity to 60% and deletion of CS3, the third conserved region adjacent to the Walker B motif led to a reduction to 42% ATPase activity. Interestingly, adhesion of the OppA mutants to immobilized HeLa cells demonstrated that two distal regions are mainly involved in adherence of OppA: the CS1 region, deletion of which led to 35% of the cytoadhesion, and the Walker BA with the adjacent upstream region CS3, deletion of which led to 25% of the cytoadhesion. The influence of the ATPase activity on the adherence of M. hominis to HeLa cells was confirmed by the use of ATPase inhibitors which reduced mycoplasmal cytoadhesion to 50%. Conclusions These findings suggest that the OppA-mediated cytoadherence of Mycoplasmahominis depends on both, the topology of the neighbouring CS1 and ATPase domain regions and the functionality of the ecto-ATPase activity in addition.
Purpose: The prevalence of Ureaplasma urealyticum and Mycoplasmahominis m the endocervix at the time of oocyte collection in women undergoing in vitro fertilization (IVF; was examined using the polymerase chain reaction (PCR). Methods: All women were treated with tetracycline following sample collection. Results: U. urealyticum was identified in 56 (17.2%) of 326 women while M. hominis was present in
Steven S. Witkin; Isaac Kligman; James A. Grifo; Zev Rosenwaks
The uptake of fluoroquinolones was characterized for the fluoroquinolone-susceptible strain PG21 of My- coplasma hominis. Accumulation of fluoroquinolones appeared to occur by passive diffusion. Addition of arginine as the energizer significantly reduced the uptake of fluoroquinolones, suggesting the presence of an energy-dependent efflux process. Reserpine and orthovanadate, two multidrug pump inhibitors, increased significantly the ciprofloxacin (CIP) uptake. In contrast, such
S. Raherison; P. Gonzalez; H. Renaudin; A. Charron; C. Bebear
process. The affinity of (14C)erythromycin to ribosomes isolated from M. hominis was dramatically reduced relative to that to ribosomes isolated from M. pneumoniae. The nucleotide sequences of 23S rRNA of both ribosomal operons rrnA and rrnB and ribosomal proteins L4 and L22 of M. hominis were obtained. Compared to the sequence of M. pneumoniae, M. hominis harbored a G2057A transition
S. Pereyre; P. Gonzalez; B. de Barbeyrac; A. Darnige; H. Renaudin; A. Charron; S. Raherison; C. Bebear
This study was undertaken to determine the prevalence of Chlamydia trachomatis, mycoplasmas, and ureaplasmas in semen samples of infertile compared with fertile men and to evaluate the seminological variables of semen from infected and noninfected men. A total of 127 infertile and 188 fertile men seen in a maternity hospital clinic were recruited into the study over a period of 14 months. Specimens were obtained by masturbation and examined for the presence of Ureaplasma urealyticum, Mycoplasmahominis, Mycoplasma genitalium, and C trachomatis by polymerase chain reaction. Semen analysis was performed according to World Health Organization guidelines. U urealyticum, M hominis, M genitalium, and C trachomatis were demonstrated in the semen samples of 31 (24.4%) vs 49 (26.1%), 22 (17.1%) vs 61 (32.4%), 6 (4.7%) vs 6 (3.2%), and 5 (3.9%) vs 7 (3.7%), respectively, of infertile and control men. Mixed infections were detected in 14 (11%) of infertile and 29 (15.4%) of fertile men. The infertile men positive for M hominis had semen samples that showed statistically significant differences in the mean of sperm pH and leukocyte count between infected and uninfected men (P < .03 and P < .001, respectively). Similarly, there was statistically significant difference in the leukocyte counts of M genitalium and C trachomatis in infected compared with uninfected men. A similar trend was noted in infected fertile compared with uninfected men. The difference in prevalence of these urogenital pathogens among infertile compared with fertile men was not statistically significant. However, genital mycoplasmas and chlamydial infections appeared to influence semen quality negatively. PMID:22052774
Al-Sweih, Noura A; Al-Fadli, Amani H; Omu, Alexander E; Rotimi, Vincent O
Fluoroquinolone-resistant mutants ofMycoplasma hominiswere selected in vitro from the PG21 susceptible reference strain either by multistep selection on increasing concentrations of various fluoroquinolones or by one-step selection on agar medium with ofloxacin. The quinolone resistance-determining regions (QRDR) of the structural genes encoding the A and B subunits of DNA gyrase were amplified by PCR, and the nucleotide sequencesofeightmultistep-selectedresistantstrainswerecomparedtothoseofsusceptiblestrainPG21.Four high-level resistant
CECILE MARIE BEBEAR; JOSEPH MARIE BOVE ´; CHRISTIANE BEBEAR; ANDJOEL RENAUDIN
We conducted a prospective study on 100 couples consulting for infertility at the teaching Hospital of Tours, with the scope to determine if there is a benefit for systematic screening of Chlamydia trachomatis, Mycoplasmahominis and Ureaplasma urealyticum among genito-urinary specimen when exploring couples infertility. C. trachomatis was detected by PCR on sperm, endocervix and urine specimen. M. hominis and U. urealyticum were detected by culture on A7 agar medium and with minigaleries on sperm and endocervix specimen. Standard cultures were also performed on sperm, endocervix, vaginal and urine specimen. Only one specimen (sperm) was positive for C. trachomatis. Three percent of the specimen were positive for U. urealyticum (from which 2,5% of the sperm specimen). No specimen was positive for M. hominis. Our results show that screening of C. trachomatis, M. hominis and U. urealyticum is not systematically required for among check up of infertile couples, given the prevalence of chlamydiosis among the population studied. However, it would be interesting to perform it on a targeted population, according to anamnestic or clinical criteria. In addition, an important modification of vaginal flora was observed in 12% of cases, and 2 vaginosis were diagnosed; the putative consequences of this disequilibrium has to be further investigated. PMID:16298086
Rosemond, A; Lanotte, P; Watt, S; Sauget, A S; Guerif, F; Royčre, D; Goudeau, A; Mereghetti, L
AIMS: The aim of this study was to evaluate the antimicrobial effects of five natural substances against 50 clinical isolates of Mycoplasmahominis. METHODS AND RESULTS: The in vitro activity of selected natural compounds, cinnamon bark oil, anethole, carvacrol, eugenol and guaiazulene, was investigated against 50 M. hominis isolates cultivated from cervical swabs by the broth dilution method. All showed valuable antimicrobial activity against the tested isolates. Oil from the bark of Cinnamomum zeylanicum (MBC(90) = 500 µg/mL) however was found to be the most effective. Carvacrol (MBC(90) = 600 µg/mL) and eugenol (MBC(90) = 1000 µg/mL) also possessed strong antimycoplasmal activity. CONCLUSIONS: The results indicate that cinnamon bark oil, carvacrol and eugenol have strong antimycoplasmal activity and the potential for use as antimicrobial agents in the treatment of mycoplasmal infections. PMID:23128812
Fifteen strains of M. hominis isolated from patients with urogenital inflammations were analyzed. Variations in the quinolone resistance-determining regions (QRDR) have been found in fluoroquinolone-resistant M. hominis clinical isolates in comparison with the reference PG21 strain. In one isolate, parC had Asn substitute at position 91. PMID:11186458
Gushchin, A E; Ladygina, V G; Govorun, V M; Taraskina, A M; Savicheva, A M
This work reports the results of analyses of three complete mycoplasma genomes, a pathogenic (7448) and a nonpathogenic (J) strain of the swine pathogen Mycoplasma hyopneumoniae and a strain of the avian pathogen Mycoplasma synoviae; the genome sizes of the three strains were 920,079 bp, 897,405 bp, and 799,476 bp, respectively. These genomes were compared with other sequenced mycoplasma genomes
A. T. R. Vasconcelos; H. B. Ferreira; C. V. Bizarro; S. L. Bonatto; M. O. Carvalho; P. M. Pinto; D. F. Almeida; L. G. P. Almeida; R. Almeida; L. Alves-Filho; E. N. Assuncao; V. A. C. Azevedo; M. R. Bogo; M. M. Brigido; M. Brocchi; H. A. Burity; A. A. Camargo; S. S. Camargo; M. S. Carepo; D. M. Carraro; J. C. de Mattos Cascardo; L. A. Castro; G. Cavalcanti; G. Chemale; R. G. Collevatti; C. W. Cunha; B. Dallagiovanna; B. P. Dambros; Odir A. Dellagostin; C. Falcao; F. Fantinatti-Garboggini; M. S. S. Felipe; L. Fiorentin; G. R. Franco; N. S. A. Freitas; D. Frias; T. B. Grangeiro; E. C. Grisard; C. T. Guimaraes; M. Hungria; S. N. Jardim; M. A. Krieger; J. P. Laurino; L. F. A. Lima; M. I. Lopes; E. L. S. Loreto; H. M. F. Madeira; G. P. Manfio; A. Q. Maranhao; C. T. Martinkovics; S. R. B. Medeiros; M. A. M. Moreira; M. Neiva; C. E. Ramalho-Neto; M. F. Nicolas; S. C. Oliveira; R. F. C. Paixao; F. O. Pedrosa; S. D. J. Pena; M. Pereira; L. Pereira-Ferrari; I. Piffer; L. S. Pinto; D. P. Potrich; A. C. M. Salim; F. R. Santos; R. Schmitt; M. P. C. Schneider; A. Schrank; I. S. Schrank; A. F. Schuck; H. N. Seuanez; D. W. Silva; R. Silva; S. C. Silva; C. M. A. Soares; K. R. L. Souza; R. C. Souza; C. C. Staats; M. B. R. Steffens; S. M. R. Teixeira; T. P. Urmenyi; M. H. Vainstein; L. W. Zuccherato; A. J. G. Simpson; A. Zaha
Isolation of T strainmycoplasmas was found to be directly related to sexual activity in three groups of women. Metabolic inhibition titers followed the same pattern, the number of titers increased with sexual activity. The rate of mycoplasma isolations f...
A 30-year-old human immunodeficiency virus (HIV)-1 infected woman presented with vaginal discharge and associated vulval irritation. The vaginal swabs tested positive for Ureaplasma parvum and Mycoplasmahominis by both culture and polymerase chain reaction (PCR). The specimen also tested positive for Chlamydia trachomatis deoxyribonucleic acid (DNA) by cryptic plasmid and omp1 gene PCR assays. The patient was successfully treated with azithromycin based on the antibiotic susceptibility testing results of U. parvum and M. hominis by microbroth dilution. Since sexually transmitted infections enhance the transmission of HIV, HIV-positive patients should be screened routinely for these pathogens. PMID:23867681
The efficiency of the polymerase chain reaction (PCR) was compared with that of culture for detection ofUreaplasma urealyticum andMycoplasmahominis in 726 clinical specimens comprising 189 gynecological samples, 362 urological samples, and 175 samples from newborn infants. The sensitivity of PCR versus culture was 95% for both organisms, while the sensitivity of culture versus PCR was 91% forUreaplasma urealyticum and
M. Abele-Horn; C. Wolff; P. Dressel; A. Zimmermann; W. Vahlensieck; F. Pfaff; G. Ruckdeschel
BACKGROUND: In the facultative human pathogen Mycoplasmahominis, which belongs to the cell wall-less Mollicutes, the surface-localised substrate-binding domain OppA of the oligopeptide permease was characterised as the main ecto-ATPase. RESULTS: With the idea that extra-cellular ATP could only be provided by the infected host cells we analysed the ATP release of HeLa cells after incubation with different preparations of
This work reports the results of analyses of three complete mycoplasma genomes, a pathogenic (7448) and a nonpathogenic (J) strain of the swine pathogen Mycoplasma hyopneumoniae and a strain of the avian pathogen Mycoplasma synoviae; the genome sizes of the three strains were 920,079 bp, 897,405 bp, and 799,476 bp, respectively. These genomes were compared with other sequenced mycoplasma genomes reported in the literature to examine several aspects of mycoplasma evolution. Strain-specific regions, including integrative and conjugal elements, and genome rearrangements and alterations in adhesin sequences were observed in the M. hyopneumoniae strains, and all of these were potentially related to pathogenicity. Genomic comparisons revealed that reduction in genome size implied loss of redundant metabolic pathways, with maintenance of alternative routes in different species. Horizontal gene transfer was consistently observed between M. synoviae and Mycoplasma gallisepticum. Our analyses indicated a likely transfer event of hemagglutinin-coding DNA sequences from M. gallisepticum to M. synoviae.
Vasconcelos, Ana Tereza R.; Ferreira, Henrique B.; Bizarro, Cristiano V.; Bonatto, Sandro L.; Carvalho, Marcos O.; Pinto, Paulo M.; Almeida, Darcy F.; Almeida, Luiz G. P.; Almeida, Rosana; Alves-Filho, Leonardo; Assuncao, Enedina N.; Azevedo, Vasco A. C.; Bogo, Mauricio R.; Brigido, Marcelo M.; Brocchi, Marcelo; Burity, Helio A.; Camargo, Anamaria A.; Camargo, Sandro S.; Carepo, Marta S.; Carraro, Dirce M.; de Mattos Cascardo, Julio C.; Castro, Luiza A.; Cavalcanti, Gisele; Chemale, Gustavo; Collevatti, Rosane G.; Cunha, Cristina W.; Dallagiovanna, Bruno; Dambros, Bibiana P.; Dellagostin, Odir A.; Falcao, Clarissa; Fantinatti-Garboggini, Fabiana; Felipe, Maria S. S.; Fiorentin, Laurimar; Franco, Gloria R.; Freitas, Nara S. A.; Frias, Diego; Grangeiro, Thalles B.; Grisard, Edmundo C.; Guimaraes, Claudia T.; Hungria, Mariangela; Jardim, Silvia N.; Krieger, Marco A.; Laurino, Jomar P.; Lima, Lucymara F. A.; Lopes, Maryellen I.; Loreto, Elgion L. S.; Madeira, Humberto M. F.; Manfio, Gilson P.; Maranhao, Andrea Q.; Martinkovics, Christyanne T.; Medeiros, Silvia R. B.; Moreira, Miguel A. M.; Neiva, Marcia; Ramalho-Neto, Cicero E.; Nicolas, Marisa F.; Oliveira, Sergio C.; Paixao, Roger F. C.; Pedrosa, Fabio O.; Pena, Sergio D. J.; Pereira, Maristela; Pereira-Ferrari, Lilian; Piffer, Itamar; Pinto, Luciano S.; Potrich, Deise P.; Salim, Anna C. M.; Santos, Fabricio R.; Schmitt, Renata; Schneider, Maria P. C.; Schrank, Augusto; Schrank, Irene S.; Schuck, Adriana F.; Seuanez, Hector N.; Silva, Denise W.; Silva, Rosane; Silva, Sergio C.; Soares, Celia M. A.; Souza, Kelly R. L.; Souza, Rangel C.; Staats, Charley C.; Steffens, Maria B. R.; Teixeira, Santuza M. R.; Urmenyi, Turan P.; Vainstein, Marilene H.; Zuccherato, Luciana W.; Simpson, Andrew J. G.; Zaha, Arnaldo
T-strains of Mycoplasma, associated with 70-80 per cent of cases of nongonococcal urethritis in human males, were selectively inhibited by low concentrations of erythromycin in vitro. In an agar dilution system, T-strains of Mycoplasma were completely inh...
Eleven mycoplasmastrains were isolated from the semen of 24 stallions. Eight of these strains were identified as Mycoplasma equigenitalium. Three strains which hydrolized arginine could not be identified. The growth inhibition test with immune sera against M. arginini and M. equirhinis was negative. Antibiotic sensitivity test showed that all strains were sensitive to four antibiotic of tetracycline group (oxytetracyclin, minocycline, transcycline and vibramycin). Lincomycin and gentamycin appeared to be the most active against all the strains. Comparative analysis of routine semen examination did not reveal any difference between ejaculates infected with mycoplasma and free of these organisms. However, the levels of certain biochemical components of the semen plasma (glycerylphosphorylcholine, ergothioneine, fructose and of the semen plasma (glycerylphosphorylcholine, ergothioneine, fructose and total protein) from mycoplasma-positive ejaculates were significantly lower than in the semen plasma from mycoplasma free ejaculates. PMID:3833120
Zgórniak-Nowosielska, I; Kosiniak, K; Slagowska, A
Background Among the surface antigens of Mycoplasmahominis, the P120' protein was previously shown to elicit a subtle antibody response and appears to be relatively conserved. To get better insight into the evolution of this protein, we analysed the genetic variability of its surface exposed region in 27 M. hominis isolates recovered from the genital tract of Tunisian patients with infertility disorders. Methods All specimens were processed for culture and PCR amplification of the N-terminal surface exposed region of p120' gene. PCR products were sequenced to evaluate the genetic variability, to test for adaptive selection, and to infer the phylogenetic relationship of the M. hominis isolates. Results Sequence analysis showed a total of 25 single nucleotide polymorphisms distributed through 23 polymorphic sites, yielding 13 haplotypes. All but one mutation were confined within three distinct regions. Analysis of the amino acid-based phylogenetic tree showed a predominant group of 17 closely related isolates while the remaining appear to have significantly diverged. Conclusion By analysing a larger sample of M. hominis recovered from patients with urogenital infections, we show here that the P120' protein undergoes substantial level of genetic variability at its surface exposed region.
We report the cloning and characterization of the gyrA gene of the Mycoplasmahominis DNA gyrase, which was previously shown to be associated with quinolone resistance in this organism. The 2,733-bp gyrA gene encodes a protein of 911 amino acids with a calculated molecular mass of 102.5 kDa. As expected, M. hominis GyrA exhibits higher homology with the GyrA subunits of the gram-positive bacteria Clostridium acetobutylicum, Bacillus subtilis, Streptococcus pneumoniae, and Staphylococcus aureus than with its Escherichia coli counterpart. Knowing the entire sequence of the gyrA gene of M. hominis could be very useful for confirming the role of the GyrA subunit in fluoroquinolone resistance. Twenty-nine mutants of M. hominis were selected stepwise for resistance to trovafloxacin, a new potent fluoroquinolone, and their gyrA, gyrB, parC, and parE quinolone resistance-determining regions were characterized. Three rounds of selection yielded 3 first-step, 12 second-step, and 14 third-step mutants. The first-step mutants harbored a single substitution, Glu460?Lys (E. coli coordinates), in ParE. GyrA changes, Ser83?Leu, Glu87?Lys, and Ala119?Glu or Val, were found only in the second round of selection. At the third step, additional substitutions, at ParC Ser80, Ser81, and Glu84 and ParE Leu440, associated with high-level resistance to fluoroquinolones, appeared. Thus, high-level resistance to trovafloxacin required three steps and was associated with alterations in both fluoroquinolone targets. According to these genetic data, in M. hominis, as in Staphylococcus aureus and Streptococcus pneumoniae, topoisomerase IV seems to be the primary target of trovafloxacin.
Bebear, C. M.; Grau, O.; Charron, A.; Renaudin, H.; Gruson, D.; Bebear, C.
Infection by Mycoplasma bovis (M. bovis) can induce diseases, such as pneumonia and otitis media in young calves and mastitis and arthritis in older animals. Here, we report the finished and annotated genome sequence of M. bovis strain Hubei-1, a strain isolated in 2008 that caused calf pneumonia on a Chinese farm. The genome of M. bovis strain Hubei-1 contains a single circular chromosome of 953,114 bp with a 29.37% GC content. We identified 803 open reading frames (ORFs) that occupy 89.5% of the genome. While 34 ORFs were Hubei-1 specific, 662 ORFs had orthologs in the M. bovis type strain PG45 genome. Genome analysis validated lateral gene transfer between M. bovis and the Mycoplasma mycoides subspecies mycoides, while phylogenetic analysis found that the closest M. bovis neighbor is Mycoplasma agalactiae. Glycerol may be the main carbon and energy source of M. bovis, and most of the biosynthesis pathways were incomplete. We report that 47 lipoproteins, 12 extracellular proteins and 18 transmembrane proteins are phase-variable and may help M. bovis escape the immune response. Besides lipoproteins and phase-variable proteins, genomic analysis found two possible pathogenicity islands, which consist of four genes and 11 genes each, and several other virulence factors including hemolysin, lipoate protein ligase, dihydrolipoamide dehydrogenase, extracellular cysteine protease and 5?-nucleotidase.
The product nitroxoline was studied in vitro for its activity towards Ureaplasma urealyticum and Mycoplasmahominis. In view of the low MIC values obtained, it seems nitroxoline could be used in the treatment of urinary infections. It is bactericidal, and should not produce resistant strains. PMID:3543811
Increasing use of Mycoplasma gallisepticum (MG) live vaccines has led to a need for the differentiation of MG strains. The MG strains MK-7, MS-16, S6, FS-9 and R strains and the MG live vaccine strain F were compared by random amplification of polymorphic DNA (RAPD) in this study. Using RAPD, different patterns were found among the MG strains. In addition to this, we examined the differentiating potential of polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) primers targeted at the crmA, crmB, crmC, gapA, mgc2 and pvpA genes encoding cytadherence-related surface proteins. These proteins may take part in the pathogenesis of MG-induced disease. Differentiation of strain F is based on the identification of restriction enzyme sites in the PCR amplicons. Using HphI enzyme, crmC PCR amplicons produced different RFLP patterns. Digestion of amplicons of gapA-specific PCR with MboI enzyme also produced distinct patterns. Differences were observed among strains R and F by digestion of mgc2 PCR amplicons with HaelIl and VspI enzymes and digestion of pvpA PCR amplicons with AccI, PvulI and ScrFI endonucleases. This method can be used for the rapid differentiation of vaccine strain from wild strains. Differentiation of MG strains is a great advantage for diagnosticians or practitioners and it is useful for epidemiological studies. PMID:17278716
Biró, Judit; Erdei, Noémi; Székely, Ibolya; Stipkovits, L
We demonstrate here that the arbitrary primer polymerase-chain-reaction-based DNA fingerprinting method (also termed random amplified polymorphic DNA or RAPD) can be used to distinguish among strains of the avian pathogen Mycoplasma gallisepticum. Ten base oligonucleotide primers were used individually to prime DNA synthesis from genomic DNAs. Strain-specific arrays of DNA fragments were generated, which allowed us to identify and group isolates. Isolates of M. synoviae, M. gallinarum and M. iners yielded arrays of DNA fragments that differed markedly from those generated from the M. gallisepticum isolates using the same arbitrary primers. These results show that the RAPD fingerprinting method distinguishes genetically different strains of M. gallisepticum and indicates that it should be valuable for monitoring transmission of this pathogen. PMID:7870072
Geary, S J; Forsyth, M H; Aboul Saoud, S; Wang, G; Berg, D E; Berg, C M
Monoglucosyl diglyceride is synthesized from 1,2-diglyceride and uridine-5?-diphosphoglucose (UDP); diglucosyl diglyceride from monoglucosyl diglyceride, and uridine-5?-diphosphoglucose by membranes of Mycoplasma laidlawii strain B. All of these enzymatic activities reside in the membrane. Membranes solubilized by detergent action or succinylation and acetone powders of membranes were inactive. Requirements for Mg2+, UDP, and appropriate lipid acceptor were demonstrated for biosynthesis of both glycolipids. Glucose-1-phosphate plus uridine triphosphate could replace the UDP requirement. A medium of relatively high ionic strength and a critical concentration of sodium lauryl sulfate stimulated biosynthesis of the monoglucosyl diglyceride. The optimal pH for both reactions was 8.0. A specificity for 1,2-diglyceride from the homologous organism was found for optimal synthesis of the monoglucosyl diglyceride, and a specificity for monoglucosyl diglyceride was found in the case of diglucosyl diglyceride synthesis. Both reactions were specific for UDP.
Some Mycoplasma species are well-known avian pathogens and are of importance in poultry breeder flocks due to their pathogenic potential for embryos. Mycoplasmas are regularly detected in birds of prey, and a strain of Mycoplasma lipofaciens that was isolated from an egg of a Northern Goshawk (Accipiter gentilis) was examined for its pathogenicity in specific pathogen free chicken embryos since
Background Mycoplasmas-contamination of Orientia tsutsugamushi, one of the obligated intracellular bacteria, is a very serious problem in in vitro studies using cell cultures because mycoplasmas have significant influence on the results of scientific studies. Only a recommended decontamination method is to passage the contaminated O. tsutsugamushi strains through mice to eliminate only mycoplasmas under influence of their immunity. However, this method sometimes does not work especially for low virulent strains of O. tsutsugamushi which are difficult to propagate in mice. In this study, we tried to eliminate mycoplasmas contaminants from both high virulent and low virulent strains of the contaminated O. tsutsugamushi by repeating passage through cell cultures with antibiotics in vitro. Results We cultured a contaminated, high virulent strain of O. tsutsugamushi using a mouse lung fibroblasts cell line, L-929 cell in the culture medium containing lincomycin at various concentrations and repeated passages about every seven days. At the passage 5 only with 10 ?g/ml of lincomycin, we did not detect mycoplasmas by two PCR based methods whereas O. tsutsugamushi continued good growth. During following four passages without lincomycin, mycoplasmas did not recover. These results suggested that mycoplasmas were completely eliminated from the high virulent strain of O. tsutsugamushi. Furthermore, by the same procedures with 10 ?g/ml of lincomycin, we also eliminated mycoplasmas from a contaminated, low virulent strain of O. tsutsugamushi. Our additional assay showed that 50 ?g/ml of lyncomycin did not inhibit the growth of O. tsutsugamushi, although MICs of many mycoplasmas contaminants were less than 6 ?g/ml as shown previously. Conclusion Our results showed an alternative method to eliminate mycoplasmas from the contaminated O. tsutsugamushi strains in place of in vivo passage through mice. Especially this notable method works for the decontamination not only from the high virulent strain also from the low virulent strain of O. tsutsugamushi. For further elimination, lincomycin at the limit concentration, which does not inhibit the growth of O. tsutsugamushi, can possibly eliminate most mycoplasmas from contaminated O. tsutsugamushi strains.
BACKGROUND: While the genomic era is accumulating a tremendous amount of data, the question of how genomics can describe a bacterial species remains to be fully addressed. The recent sequencing of the genome of the Mycoplasma agalactiae type strain has challenged our general view on mycoplasmas by suggesting that these simple bacteria are able to exchange significant amount of genetic
Laurent X Nouvel; Pascal Sirand-Pugnet; Marc S Marenda; Eveline Sagné; Valérie Barbe; Sophie Mangenot; Chantal Schenowitz; Daniel Jacob; Aurélien Barré; Stéphane Claverol; Alain Blanchard; Christine Citti
Mycoplasma pneumoniae strain 309, a type 2a (subtype 2 variant) strain of this bacterium, has variations in the P1 protein, which is responsible for attachment of the bacterium to host cells. Here, we report the complete genome sequence of M. pneumoniae strain 309 isolated from a pneumonia patient in Japan.
Mycoplasma anatis, a member of the class Mollicutes, is the causative agent of a contagious infectious disease of domestic ducklings, wild birds, and eggs. Increasing reports show that coinfection of M. anatis with Escherichia coli results in substantial economic impacts on the duck farms in China. Here, we announce the first genome sequence of M. anatis.
The recently accomplished complete genomic sequence analysis of the type strain PG1 of Mycoplasma mycoides subsp. mycoides small-colony type revealed four large repeated segments of 24, 13, 12, and 8 kb that are flanked by insertion sequence (IS) elements. Genetic analysis of type strain PG1 and African, European, and Australian field and vaccine strains revealed that the 24-kb genetic locus is
\\u000a As is well known, medieval logicians gave a lot of attention to the analysis of different sophisms in which there are propositions\\u000a that include complex terms one of whose parts is a term in an oblique case. The propositions ‘cuiuslibet hominis asinus currit, ‘ab omni homine enuntiatum est verum \\/ ab utroque istorum enuntiatum est verum’, ‘omnem hominem videns currit’,
The prevalence of Mycoplasma fermentans, Mycoplasma pirum, Mycoplasma genitalium, Mycoplasma pneumoniae, Mycoplasmahominis, and Mycoplasma penetrans was investigated by using specific PCR assays with peripheral blood mononuclear cells from subjects infected or not infected with the human immunodeficiency virus (HIV). Only M. fermentans was detected in 5.8% of 154 HIV-seropositive and 11.1% of 90 HIV-seronegative subjects.
Mycoplasma gallisepticum (MG) is an important avian pathogen causing significant economic losses within the poultry industry. In an effort to develop tools to aid in MG research and diagnostics, we have compared sequences of the attenuated MG vaccine strain ts-11 to those of commonly used pathogenic challenge strains in search of a simple means of differentiation. Via gapA sequence alignments and comparisons, we have identified and designed primers facilitating strain differentiation. When applied to conventional polymerase chain reaction (PCR) assay at low annealing temperature, the primer sets allow for the differentiation of MG attenuated vaccine strains ts-11 as well as the attenuated MG vaccine strain 6/85 from the commonly utilized MG challenge strains R(low), R, and S6. Conventional PCR differentiation is based on the visualization of sole products with the attenuated MG strains ts-11 and 6/85 and the lack of the corresponding products from MG strains R(low), R, and S6. When applied to MG strain F, product visualization varies with the applied primer set. The differentiation of MG strains ts-11 and 6/85 from the pathogenic challenge strains was also accomplished via real-time analyses, however, the primer sets were not able to differentiate MG strains ts-11 and 6/85 from selected MG field isolates. PMID:18939641
Some Mycoplasma species are well-known avian pathogens and are of importance in poultry breeder flocks due to their pathogenic potential for embryos. Mycoplasmas are regularly detected in birds of prey, and a strain of Mycoplasma lipofaciens that was isolated from an egg of a Northern Goshawk (Accipiter gentilis) was examined for its pathogenicity in specific pathogen free chicken embryos since birds of prey eggs were not available for this purpose. The strain was found to be pathogenic, causing a high mortality as well as dwarfing, curled toes and infiltrations of heterophils in the liver, kidney, intestine and chorioallantoic membrane. PMID:17479376
Clinical findings in 36 immunosuppressed patients with lower respiratory tract infection or bacteremia with Actinobacillus hominis are described. Animal contact was only recorded for three patients; nine patients died despite appropriate antimicrobial treatment. Although infections with this microorganism seem to be rare, the fact that 37 of 46 strains characterized in this study have been found in Copenhagen indicates that under-reporting may occur. A. hominis is phenotypically relatively homogeneous but can be difficult to differentiate from other Actinobacillus species unless extensive biochemical testing is performed. Mannose-positive strains of A. hominis are especially difficult to differentiate from A. equuli. Attempts to identify A. hominis by automatic identification systems may lead to misidentifications. Ribotyping and DNA-DNA hybridization data show that A. hominis is a homogeneous species clearly separated from other species within the genus Actinobacillus.
DNA heterogeneity among strains and isolates of Mycoplasma gallisepticum (MG) was demonstrated with the arbitrarily primed polymerase chain reaction (AP-PCR) method. This method involves three cycles of low-stringency amplification followed by PCR at higher stringency. Reproducible DNA fragments of 25 different MG strains or isolates were generated with three arbitrarily chosen primers. The MG strains or isolates were distinguished according to the banding patterns of their amplified DNA on agarose gels, and the differences were characteristic for specific isolates. This method is rapid, simple, and reproducible, and it can also be used to determine the similarity between isolates of MG from various sources. PMID:8719206
Only limited information regarding the antimicrobial susceptibilities of Mycoplasma genitalium is available because of difficulties in isolating M. genitalium strains from clinical specimens. Antimicrobial susceptibilities of 15 clinical isolates, 7 ATCC strains, and an early passage of the M30 strain were examined by the broth dilution method. Azithromycin, clarithromycin, sitafloxacin, and moxifloxacin were the most active drugs against M. genitalium, and their MIC(90)s were 0.002, 0.008, 0.125, and 0.125 mg/liter, respectively. PMID:19738019
Only limited information regarding the antimicrobial susceptibilities of Mycoplasma genitalium is available because of difficulties in isolating M. genitalium strains from clinical specimens. Antimicrobial susceptibilities of 15 clinical isolates, 7 ATCC strains, and an early passage of the M30 strain were examined by the broth dilution method. Azithromycin, clarithromycin, sitafloxacin, and moxifloxacin were the most active drugs against M. genitalium, and their MIC90s were 0.002, 0.008, 0.125, and 0.125 mg/liter, respectively.
Twenty Mycoplasma arthritidis strains or isolates were compared by a combination of enzyme-linked immunosorbent assay by an antiserum adsorption technique, Western immunoblotting, and restriction analysis of chromosomal DNA. Antigenic markers that defined strains related to strains 158p10p9, PG6, and H606 were identified. In addition, restriction analysis allowed all 20 strains to be divided into six groups. Results of restriction analysis corresponded generally with antigenic similarities, although the former did not allow grouping with as fine a precision as the latter. However, intrastrain antigenic variability, which is common among many Mycoplasma species, including M. arthritidis, introduced a complicating factor into our attempts at antigenic analysis. While serologic and antigenic analyses remain useful, we recommend that they be used with caution and in combination with other techniques for identifying and characterizing new isolates and newly acquired strains. Combinations of these techniques have proven to be useful in our laboratory for quality control and for uncovering interesting relationships among strains subjected to animal passage and their less virulent antecedents and among strains originally classified as the same but obtained from different sources and maintained, sometimes for decades, in different laboratories.
Washburn, L R; Voelker, L L; Ehle, L J; Hirsch, S; Dutenhofer, C; Olson, K; Beck, B
The recently accomplished complete genomic sequence analysis of the type strain PG1 of Mycoplasma mycoides subsp. mycoides small-colony type revealed four large repeated segments of 24, 13, 12, and 8 kb that are flanked by insertion sequence (IS) elements. Genetic analysis of type strain PG1 and African, European, and Australian field and vaccine strains revealed that the 24-kb genetic locus is repeated only in PG1 and not in other M. mycoides subsp. mycoides SC strains. In contrast, the 13-kb genetic locus was found duplicated in some strains originating from Africa and Australia but not in strains that were isolated from the European outbreaks. The 12- and 8-kb genetic loci were found in two and three copies, respectively, in all 28 strains analyzed. The flanking IS elements are assumed to lead to these tandem duplications, thus contributing to genomic plasticity. This aspect must be considered when designing novel diagnostic approaches and recombinant vaccines. PMID:16919417
Mycoplasma gallisepticum (MG) is an economically important pathogen of poultry worldwide, causing chronic respiratory disease in chickens and turkeys. Differentiation of MG strains is critical, especially in countries where poultry flocks are vaccinated with live vaccines. In this study, oligonucleotide primers were designed based on a region preceding the trinucleotide repeat of a member of the vlhA gene family, and amplicons of 145-352 bp were generated from cultures of 10 different MG strains, including the ts-11, F and 6/85 vaccine strains. High-resolution melting (HRM) curve analysis of the resultant amplicons could differentiate all MG strains. Analysis of the nucleotide sequences of the amplicons from each strain revealed that each melting curve profile related to a unique DNA sequence. The HRM curve profiles (for ts-11) remained consistent after at least five passages under laboratory conditions. PCR-HRM curve analysis of 33 DNA extracts derived from respiratory swabs, or mycoplasma cultures grown from respiratory swabs, of ts-11-vaccinated commercial or specific pathogen-free chickens identified all these specimens, according to their sequences, as ts-11. The potential of the PCR-HRM curve analysis was also shown in the genotyping of 30 additional MG isolates from Europe, the USA and Israel. The results presented in this study indicate that PCR followed by HRM curve analysis provides a rapid and robust technique for genotyping of MG isolates/strains using both MG cultures and clinical swabs. PMID:20035007
Ghorashi, Seyed A; Noormohammadi, Amir H; Markham, Philip F
Several mycoplasma species have been shown to form biofilms that confer resistance to antimicrobials and which may affect the host immune system, thus making treatment and eradication of the pathogens difficult. The present study shows that the biofilms formed by two strains of the human pathogen Mycoplasma pneumoniae differ quantitatively and qualitatively. Compared with strain UAB PO1, strain M129 grows well but forms biofilms that are less robust, with towers that are less smooth at the margins. A polysaccharide containing N-acetylglucosamine is secreted by M129 into the culture medium but found in tight association with the cells of UAB PO1. The polysaccharide may have a role in biofilm formation, contributing to differences in virulence, chronicity and treatment outcome between strains of M. pneumoniae. The UAB PO1 genome was found to be that of a type 2 strain of M. pneumoniae, whereas M129 is type 1. Examination of other M. pneumoniae isolates suggests that the robustness of the biofilm correlates with the strain type. PMID:23412845
Simmons, Warren L; Daubenspeck, James M; Osborne, John D; Balish, Mitchell F; Waites, Ken B; Dybvig, Kevin
A Mycoplasma gallisepticum (MG) F-vaccine strain polymerase chain reaction (PCR) (MGF-PCR) was developed and standardized. The origin of the primers was a clone (p08-M6#17) that contained an MG F-strain-specific DNA fragment of 6.0 kilobase pairs designated fMGF-1. Both ends of fMGF-1 (BamHI and EcoRI) were sequenced, and regions adequate for the primers were chosen. Seven 25-base primers were synthesized, and two near the EcoRI end (MGF-P1 left [L] and right [R]) were selected for MGF-PCR, MGF-P1 L and R amplified a DNA product of 524 base pairs (bp) that was directed at F-strain-related MG only. None of 16 other species of avian mycoplasmas that were tested yielded MGF-PCR product. MGF-PCR was able to consistently detect F-strain samples containing 54 cells or more and inconsistently (at least one positive out of five replicates) in samples with fewer organisms. The MGF-PCR products were visualized either by gel electrophoresis or Southern blot hybridization with a probe containing an identical base sequence as the 524-bp product amplified by MGF-PCR. The MGF-PCR was 1000 to 10,000 times more sensitive than dot-blot assays using two MG F-strain-specific probes. PMID:8452497
The in vitro sensitivity of 20 wild strains of M. capricolum and that of the reference strain (California kid) against 14 antibiotics was investigated by means of a micromethod. The technique is based on the determination of the inhibitory minimum concentration (IMC) in a liquid medium as revealed by the inhibition of glucose metabolism. The following results were obtained: all strains were sensitive to five antibiotics (tylosin, oxytetracyclin, gentamycin, neomycin, nalidixic acid) with an IMC varying from 0.06 to 8 meq/ml. The variation in the IMC values from 8 to 32 meq/ml for spiramycin and erythromycin indicated that some of these strains were sensitive and other resistant to these two drugs. All strains were resistant to seven antibiotics (streptomycin, bacitracin, polymyxin, chloramphenicol, lincomycine, rifampicine and novobiocine), sometimes at a concentration exceeding 128 meg/ml. PMID:2132784
The pathomorphological findings and the expression and distribution of variable surface protein antigens (Vsp) of Mycoplasma (M.) bovis were characterised immunohistochemically in lungs of eight calves following inoculation with a Vsp A-expressing clonal variant of M. bovis type strain PG45. Within 48h post inoculation (p.i.) an innate immune response dominated by macrophages and neutrophils develops. The monoclonal antibodies (mAbs) 1A1
I. Buchenau; F. Poumarat; D. Le Grand; H. Linkner; R. Rosengarten; M. Hewicker-Trautwein
The aim of this study was to examine the incidence and antibiotic sensitivity of Ureaplasma urealyticum and Mycoplasmahominisstrains cultured from the genital discharges of sexually active individuals who attended our STD outpatient service. Samples were taken with universal swab (Biolab®, Budapest, Hungary) into the Urea-Myco DUO kit (Bio-Rad®, Budapest, Hungary) and incubated in ambient air for 48 h at 37 °C. The determination of antibiotic sensitivity was performed in U9 and arginin broth using the SIR Mycoplasma kit (Bio-Rad®, Budapest, Hungary) under the same conditions. Between 01.05.2008 and 31.12.2011, 373/4,466 (8.35 %) genito-urethral samples with U. urealyticum and 41/4,466 (0.91 %) genito-urethral samples with M. hominis infection were diagnosed in sexually active individuals in the National STD Center, Semmelweis University. U. urealyticum was isolated in 12.54 % in the cervix and 4.1 % in the male urethra, while M. hominis was isolated in 1.33 % in the cervix and 0.51 % in the male urethra. The affected age group was between 21 and 60 years old. U. urealyticum strains were sensitive to tetracycline (95.9 %), doxycycline (97.32 %), and azithromycin (85.79 %), and resistant to erythromycin (81.23 %), clindamycin (75.06 %), and ofloxacin (25.2 %). Cross-resistance occurred in 38.71 % of patients to erythromycin and clindamycin. M. hominisstrains were sensitive to clindamycin, ofloxacin, and doxycycline in more than 95 %, to tetracycline in 82.92 %, and no cross-resistance was detected among the antibiotics. Our study confirms that the continuously changing antibiotic resistance of ureaplasmas and mycoplasmas should be followed at least in a few centers in every country, so as to determine the best local therapy options for sexually transmitted infection (STI) patients. PMID:23686458
Pónyai, K; Mihalik, N; Ostorházi, E; Farkas, B; Párducz, L; Marschalkó, M; Kárpáti, S; Rozgonyi, F
In this study, six Chinese strains of Mycoplasma mycoides subsp. mycoides small colony type (MmmSC) isolated between 1953–1960 were analysed and their molecular characteristics compared to those of the African PG1 and Afade strains, the European C305 and 138\\/5 strains and the closely related caprine M. mycoides subsp.mycoides large colony type Y-goat strain. PCR amplification of long DNA fragments showed
Yuan Li; Jiuqing Xin; Yulong Gao; Jianhua Zhang; Robin A. J. Nicholas; Yiqing Lin
Mycoplasma sp., strain J, contains two glycolipids, cholesteryl beta-D-glucoside and 3,4,6-triacyl-beta-D-glucopyranose. The structure of the latter was proven by standard chemical procedures. Glycolipids comprise about 20% of the total lipids of the orga...
Mycoplasma gallisepticum causes respiratory disease and production losses in poultry. Vaccination of poultry with M. gallisepticum live vaccines is an approach to reduce susceptibility to infection and to prevent the economic losses. The development and evaluation of live vaccines usually requires the involvement of several vaccine and challenge strains in the same experimental setup. Our goal was to develop a tool to allow the differentiation between a set of known M. gallisepticum strains in a quantitative manner. We developed 5 real-time PCR assays that absolutely differentiated between one of the five commercial and laboratory vaccine strains: F, ts-11, 6/85, K5831, K5054, and the challenge strain R low when tested on in vitro cultures. The assay K5831 vs. R low was also tested on specimens from live birds that were vaccinated with K5831 and challenged with R low, and successfully differentiated between the vaccine and the challenge strains in a quantitative manner. This preliminary in vivo application of the method also shed light on possible protection mechanisms for the M. gallisepticum K5831 vaccine strain. PMID:18160233
Raviv, Ziv; Callison, Scott A; Ferguson-Noel, N; Kleven, Stanley H
Mycoplasmas are pathogens of different avian species, and they are able to be vertically transmitted. Even detected, Mycoplasma prevalence in raptor eggs is very low. In contrast to poultry, raptor eggs submitted for investigations are usually incubated. To investigate the influence of incubation length on the recovery of mycoplasmas from eggs, infertile specific-pathogen-free chicken eggs and embryos were infected with Mycoplasma lipofaciens (strain ML64), which had previously been isolated from an egg of a northern goshawk (Accipiter gentilis), in two different dosages. The eggs were investigated up to 12 days after infection (infertile eggs) or embryonic death. Mycoplasmas were recovered over the entire period after embryonic death by isolation. It was possible to re-isolate M. lipofaciens (strain ML64) from infertile eggs infected with 10(6) colony-forming units (CFUs) up to 12 days, but only up to 7 days if infected with 10(2) CFUs, which may be closer to the situation after natural infection. This study demonstrates that incubation of infertile eggs does have an influence on the recovery rate of mycoplasmas. This influence must be considered if interpreting results of Mycoplasma investigations in eggs of nonpoultry species. Additionally, it is recommended to use dead in shell embryos rather than infertile eggs for Mycoplasma detection. PMID:18939632
Four strains of Mycoplasma mycoides subsp. mycoides small colony type (MmmSC) isolated from recent outbreaks of contagious bovine pleuropneumonia (CBPP) in Africa have been investigated. One Botswanan strain, M375, displayed numerous and significant phenotypic differences from both contemporary field isolates and older field and vaccine strains (African, Australian, and European strains dating back to 1936). Differ- ences include altered morphology,
Strains of Mycoplasma mycoides subsp. mycoides small colony (SC) type, the agent of contagious bovine pleuropneumonia (CBPP), were analysed with respect to the polymorphism of distribution of a newly discovered insertion element, lS1296, on the chromosome. Analysis of 64 strains isolated from Europe, Africa and Australia, including four vaccine strains and the type strain PG1, revealed ten different IS patterns,
Xiaoxing Cheng; Jacques Nicolet; F. Poumarat; Jose Regalla; Franqois Thiaucourt; J. Frey
Through a survey of the phylogenetic distribution of sialidase among mycoplasmas, we detected activity secreted by the type strains of three of eleven species frequently or first isolated from dogs. The specific activity of washed cells of the type strains of Mycoplasma canis, Mycoplasma cynos, and Mycoplasma molare ranged from 5.2 ± 0.8 × 10-6 to 1.1 ± 0.3 × 10-5 enzymatic units per colony-forming unit (U/CFU). Cells of M. molare strain H542T had twice the specific activity (P < 0.05) of M. canis strain PG14T or M. cynos strain H831T. Significant differences in sialidase activity existed among nine clinical isolates of M. canis, ranging from not detectable to 2.1 ± 0.1 × 10-5 U/CFU. The type strains of other species previously isolated from dogs (Mycoplasma arginini, Mycoplasma bovigenitalium, Mycoplasma edwardii, Mycoplasma felis, Mycoplasma gatae, Mycoplasma maculosum, Mycoplasma opalescens, and Mycoplasma spumans) did not exhibit either secreted or cell-associated sialidase activity. Neither specific nor degenerate PCR primers complementary to the three known mycoplasmal sialidase alleles were able to amplify orthologs in M. canis, M. cynos, or M. molare, further evidence that the secreted sialidase of those species is distinct from the strictly cell-associated sialidases of Mycoplasma alligatoris, Mycoplasma synoviae, and Mycoplasma gallisepticum. This is the first report of a well-known bacterial virulence factor whose expression varies among strains of certain Mycoplasma species that infect dogs.
Although Mycoplasma gallisepticum (MG) is established in house finch (Carpodacus mexicanus) populations in at least 33 states, the potential risk of MG introduction to domestic poultry by infected finches currently is unknown. The objectives of this study were to determine if chickens could be infected with the finch strain of MG via direct, across-wire, and proximity (across-room) contact with naturally infected house finches and to determine if house finches could be infected through direct contact with experimentally infected chickens. Chickens were infected with the finch strain of MG through direct contact with naturally infected house finches, a determined by seroconversion (80%), polymerase chain reaction (PCR) (20%), and culture of MG (30%). Clinical disease was not observed in infected chickens. Isolates from chickens were identified as the original finch strain by arbitrary primed PCR. Transmission required an extended period of direct contact (10 wk) with infected finches, and no evidence of MG infection was detected in chickens exposed to infected finches across wire or across the room. Evidence of contact transmission of MG from infected chickens to house finches was limited to positive serum plate agglutination results, and infection could not be confirmed by PCR or culture. Results suggest that minimal biosecurity measures that restrict direct contact between chickens and house finches should significantly reduce the potential for MG transmission between these species. PMID:9645326
Stallknecht, D E; Luttrell, M P; Fischer, J R; Kleven, S H
The in vitro response of mouse thymocytes to various mycoplasmas was evaluated. Cultures of thymus cells from BALB mice were prepared in Earle minimal essential medium with 10 per cent fetal calf serum. After an initial drop in viability, cell populations stabilized at approximately 10-5 viable cells/ml for 3 to 5 days. Concentration of 10-6 to 10-8 colony-forming units of toxic isolates of Mycoplasma fermentans per ml killed over 50 per cent of these cells in a dose-dependent fashion. Four other mycoplasmas (M. pneumoniae, M. hominis, M. arthritidis and a nontoxic strain of M. fermentans) did not induce cytotoxicity of mouse thymocytes. Toxic isolates of M. fermentans multiplied in the presence of thymus cells as they were being inactivated. However, nonviable membrane preparations of these mycoplasma were also toxic, indicating that growth of the organisms is not a prerequisite for the toxic effect. The relevance of these findings for the isolation and identification of the membrane-associated toxic factor is discussed.
CDC coryneform group A-3 and A-4 bacteria were defined by Hollis and Weaver in 1981, but their taxonomic position is still unclear. By using biochemical and chemotaxonomical methods, four clinical strains belonging to CDC coryneform groups A-3 (n = 2) and A-4 (n = 2) were studied and could be assigned to the genus Cellulomonas, resulting in the first description of Cellulomonas strains isolated from clinical specimens. CDC coryneform group A-3 and A-4 strains were compared with the type strains of the seven species constituting the genus Cellulomonas at present as well as with the closely related species Oerskovia turbata, Oerskovia xanthineolytica, and Jonesia denitrificans, but their biochemical patterns were not compatible with the patterns of any of those species. Almost the entire sequences of the 16S rRNA genes of one representative strain of both CDC taxa were determined, and comparative sequence analysis confirmed the placement of the CDC coryneform group A-3 and A-4 strains studied in the Cellulomonas-Oerskovia subbranch of the actinomycetes. Both CDC taxa exhibited > 99% base pair homology within their 16S rDNAs. On the basis of phenotypic and molecular data, we formally propose a new species, Cellulomonas hominis sp. nov., for the CDC coryneform group A-3 bacteria examined. The type strain is DSM 9581. The precise taxonomic status of the CDC coryneform group A-4 strains studied remains to be established by quantitative DNA-DNA hybridizations.
Mycoplasma iowae is associated mainly with reduced hatchability in turkeys and is well known for the unusual ability of phenotypic variation in the Mycoplasma surface components as well as a relative resistance to heat, bile salts, and many antimicrobials. A subset of unique genes and a gene cluster responsible for these characteristics could be identified from the genome. Here, we report the first genome sequence of this species.
Objective: The involvement of the genital mycoplasmas Ureaplasma urealyticum and Mycoplasmahominis in complications of pregnancy has remained controversial especially because these microorganisms are frequent colonizers of the lower genital tract. Recovery of bacteria from the placenta appears to be the sole technique to represent a true infection and not vaginal contamination. Therefore, we investigated the presence of genital mycoplasmas, aerobic and anaerobic bacteria, and fungi in human placentas and evaluated their association with morbidity and mortality of pregnancy. Methods: We cultured placentas from 82 women with complicated pregnancies. One hundred placentas from women with uncomplicated pregnancies were evaluated as controls. When possible, placentas were examined histologically for presence of chorioamnionitis. Results: Microorganisms were recovered from 52% of the placentas of complicated pregnancies and U. urealyticum was the microorganism isolated most frequently from the placenta. A significant association between positive mycoplasma culture of the placenta and complication of pregnancy was found, and chorioamnionitis was positively related to isolation of mycoplasmas. Conclusions: These data suggest that genital mycoplasmas are able to infect the human placenta where they can cause chorioamnionitis. This infection of the placenta by genital mycoplasmas is related to preterm birth and fatal outcome of pregnancy.
Conjunctivitis in house finches (Carpodacus mexicanus), caused by Mycoplasma gallisepticum (MG), was first reported in 1994 and, since this time, has become endemic in house finch populations throughout eastern North America. Although the house finch is most commonly associated with MG-related conjunctivitis, MG has been reported from other wild bird species, and conjunctivitis (not confirmed as MG related) has been reported in over 30 species. To help define the host range of the house finch strain of MG and to better understand the effect of MG on other host species, we monitored a community of wild birds for exposure to MG and conducted experimental infections on nine avian species. For the field portion of our study, we conducted a 9-mo survey (August 2001 to April 2002) of wild avian species in a peri-urban environment on the campus of Auburn University. During this time 358 birds, representing 13 different families, were sampled. No clinical signs of mycoplasmosis were observed in any bird. Thirteen species from nine families had positive agglutination reactions for antibodies to MG, but all birds tested negative by polymerase chain reaction (PCR). Three mourning doves were PCR-positive for MG, but antibodies to MG were not detected. In the experimental infections, we exposed seven native avian species and two cage-bird species to MG (May 2000 to June 2002). After exposure, clinical disease was seen in all four species from the family Fringillidae and in eastern tufted titmice (Baeolophus bicolor). In addition, three other species were infected without clinical signs, suggesting that they may represent potential MG reservoirs. PMID:16107666
In this study, six Chinese strains of Mycoplasma mycoides subsp. mycoides small colony type (MmmSC) isolated between 1953-1960 were analysed and their molecular characteristics compared to those of the African PG1 and Afade strains, the European C305 and 138/5 strains and the closely related caprine M. mycoides subsp.mycoides large colony type Y-goat strain. PCR amplification of long DNA fragments showed that the six Chinese strains, the PG1 strain and the Y-goat strain, just like Afade, did not have the 8.84 kb deletion characteristic of the European strains C305 and 138/5. In comparison, the lppB gene sequence of the six MmmSC Chinese strains was found to be 99% homologous to that of PG1and Afade, but <93% homologous to the Y-goat sequence. The anti-rLppB antiserum reacted with PG1, Y-goat and the six Chinese strains at 67 kDa sites in Western blot, indicating that the lppB gene and its encoding protein exist in the Chinese strains. Multilocus sequence analysis (MLSA) of MmmSC strains from various regions confirmed that the Chinese strains were identical to the African and Australian cluster. This finding was further supported by the outcome of selective primer amplification. Based on these results, it is suggested that CBPP in China may have originated from Australia. PMID:17936044
Li, Yuan; Xin, Jiuqing; Gao, Yulong; Zhang, Jianhua; Nicholas, Robin A J; Lin, Yiqing
Mycoplasma sp. (strain F38) is the causative agent of contagious caprine pleuropneumonia, which is a goat disease of great global concern. Strain F38 belongs to the so-called "Mycoplasma mycoides cluster," and the members of this cluster have many biochemical and serological properties in common, which makes it difficult to differentiate between them by conventional methods. Their phylogenetic interrelationship are thus uncertain. The 16S rRNA gene of the rrnB operon from strain F38 was cloned and sequenced. The sequence was compared with the 16S rRNA sequences of related mycoplasmas, and phylogenetic trees were constructed by parsimony analysis. A three-way ambiguity among strain F38, Mycoplasma capricolum, and Mycoplasma sp. strain PG50 was observed in the trees. This observation is in agreement with a recent proposal to reclassify strain F38 and M. capricolum. A primer set was designed for in vitro amplification by PCR of a fragment of the 16S rRNA genes from the M. mycoides cluster. The amplimers of strain F38 could be distinguished easily from the corresponding amplimers from other members of the M. mycoides cluster by restriction enzyme analysis with PstI. This observation was utilized to design an identification system for strain F38. Part of the 16S rRNA gene of the rrnA operon from strain F38 was also cloned, and several sequence differences between the two rRNA operons were discovered, revealing microheterogeneity between the two 16S rRNA genes of this organism. Images
Bascunana, C R; Mattsson, J G; Bolske, G; Johansson, K E
A mycoplasma isolated from the cloaca of a tortoise was shown to be serologically distinct from 82 recognized Mycoplasma and Acholeplasma species. Three clones obtained from the isolated mycoplasma were examined in detail and proved indistinguishable from each other. One of these strains, 01008* (NCTC 11701), is designated the type strain of a new species, Mycoplasma testudinis. Microorganisms belonging to
Using published primers, detection of Mycoplasma synoviae and strain identification using the vlhA gene sequence was attempted. However, of 21 M. synoviae strains examined, three could not be amplified, so a new reverse primer was designed with a target in the conserved region of the vlhA gene. This allowed all 21 M. synoviae strains, a further nine strains and also material from 11 swab samples from M. synoviae-positive birds, to produce a PCR product, suggesting that the method could also be suitable for clinical specimens. The protocol was then tested on the type strains of M. synoviae and the other 22 recognised avian Mycoplasma species, with amplification of M. synoviae only. Further testing demonstrated that this PCR was equally or more sensitive than other PCR tests used to detect M. synoviae. Subsequent DNA sequence analysis of the PCR product based on percent similarity and evolutionary relationship appeared to be a useful tool for strain differentiation. PMID:19046834
Hammond, P P; Ramírez, A S; Morrow, C J; Bradbury, J M
Blastocystis hominis is a unicellular organism found commonly in the intestinal tract of humans and many other animals. Very little is known of the basic biology of the organism, and controversy surrounds its taxonomy and pathogenicity. There morphological forms (vacuolar, granular, and ameboid) have been recognized, but recent studies have revealed several additional forms (cyst, avacuolar, and multivacuolar). The biochemistry of the organism has not been studied to any extent, and organelles and structures of unknown function and composition are present in the cells. Several life cycles have been proposed but not experimentally validated. The form used for transmission has not been defined. Infections with the organism are worldwide and appear in both immunocompetent and immunodeficient individuals. Symptoms generally attributed to B. hominis infection are nonspecific, and the need for treatment is debated. If treatment appears warranted, metronidazole is suggested as the drug of choice, although failures of this drug in eradicating the organism have been reported. Infection is diagnosed by light microscopic examination of stained smears or wet mounts of fecal material. Most laboratories identify B. hominis by observing the vacuolar form, although morphological studies indicate that other forms, such as the cyst form and multivacuolar form, also should be sought for diagnosis.
Conjunctivitis in house finches (Carpodacus mexicanus), caused by Mycoplasma gal- lisepticum (MG), was first reported in 1994 and, since this time, has become endemic in house finch populations throughout eastern North America. Although the house finch is most commonly associated with MG-related conjunctivitis, MG has been reported from other wild bird species, and conjunctivitis (not confirmed as MG related) has
We present the complete genome sequence of Mycoplasma hyopneumoniae, an important member of the porcine respiratory disease complex. The genome is composed of 892,758 bp and has an average GC content of 28.6 mol%. There are 692 predicted protein coding sequences, the average protein size is 388 amino acids, and the mean coding density is 91%. Functions have been assigned
F. Chris Minion; Elliot J. Lefkowitz; Melissa L. Madsen; Barbara J. Cleary; Steven M. Swartzell; Gregory G. Mahairas
The polymerase chain reaction (PCR) technique was compared with culture for the detection of Ureaplasma urealyticum, Mycoplasmahominis, and Mycoplasma genitalium in clinical samples (vaginal secretions, throat and endotracheal secretions, and skin swabs) obtained from 47 high-risk pregnant women peripartum and eight newborn infants. Detection using PCR with homologous primers was highly specific, as a product with the expected length
N. Luki; P. Lebel; M. Boucher; B. Doray; J. Turgeon; R. Brousseau
Mycoplasma synoviae strain MS-H, developed by chemical mutagenesis of the Australian field strain 86079/7NS, is a live temperature-sensitive (ts (+)) vaccine used for control of M. synoviae infection in poultry worldwide. Genetic basis of temperature sensitivity and attenuation of MS-H has not been revealed thus far. Comparison of the complete genome sequence of MS-H, its parent strain 86079/7NS and two non-temperature sensitive (ts (-)) reisolates of MS-H revealed a mutation in a highly conserved domain of GTP binding protein Obg of MS-H, with reversion in ts (-) MS-H reisolates. Nucleotide change from G to A at position 369 of the obg gene resulted in an alteration of glycine to arginine at position 123 in Obg fold. Further analysis of the complete obg gene sequence in several MS-H reisolates revealed that a Gly123Arg substitution was associated with alteration in temperature sensitivity phenotype of MS-H. A second mutation, C to T at position 629, in obg gene was found in some of the MS-H reisolates and appeared to suppress the effects of the Gly123Arg substitution. In silico analysis of point mutations revealed that Gly123Arg has highly destabilizing effect on the MS-H Obg structure that can potentially abolish its biological functions in vivo especially at non-permissive temperature. Findings of this study implicate Obg alteration (Gly123Arg) as one of the possible causes of MS-H attenuation/temperature sensitivity and warrant further investigations into exploring the role of Obg-like proteins, an evolutionarily conserved protein from human to bacteria, in the biology of mycoplasmas. PMID:24069254
Shahid, Muhammad A; Markham, Philip F; Markham, John F; Marenda, Marc S; Noormohammadi, Amir H
Mycoplasma synoviae strain MS-H, developed by chemical mutagenesis of the Australian field strain 86079/7NS, is a live temperature-sensitive (ts+) vaccine used for control of M. synoviae infection in poultry worldwide. Genetic basis of temperature sensitivity and attenuation of MS-H has not been revealed thus far. Comparison of the complete genome sequence of MS-H, its parent strain 86079/7NS and two non-temperature sensitive (ts–) reisolates of MS-H revealed a mutation in a highly conserved domain of GTP binding protein Obg of MS-H, with reversion in ts– MS-H reisolates. Nucleotide change from G to A at position 369 of the obg gene resulted in an alteration of glycine to arginine at position 123 in Obg fold. Further analysis of the complete obg gene sequence in several MS-H reisolates revealed that a Gly123Arg substitution was associated with alteration in temperature sensitivity phenotype of MS-H. A second mutation, C to T at position 629, in obg gene was found in some of the MS-H reisolates and appeared to suppress the effects of the Gly123Arg substitution. In silico analysis of point mutations revealed that Gly123Arg has highly destabilizing effect on the MS-H Obg structure that can potentially abolish its biological functions in vivo especially at non-permissive temperature. Findings of this study implicate Obg alteration (Gly123Arg) as one of the possible causes of MS-H attenuation/temperature sensitivity and warrant further investigations into exploring the role of Obg-like proteins, an evolutionarily conserved protein from human to bacteria, in the biology of mycoplasmas.
Shahid, Muhammad A.; Markham, Philip F.; Markham, John F.; Marenda, Marc S.; Noormohammadi, Amir H.
This study examined the susceptibility to several antimicrobials of 28 isolates of Mycoplasma agalactiae obtained from goats in a region (southeastern Spain) where contagious agalactia is endemic. For each isolate, the minimum inhibitory concentration (MIC) against 12 antimicrobials of the quinolone, macrolide, aminoglycoside, and tetracycline families was determined. The antimicrobials with the lowest MIC were enrofloxacin, ciprofloxacin, tylosin, and doxycycline, all with MIC90 (concentration at which growth of 90% of the isolates is inhibited) <1µg/mL. Norfloxacin (a quinolone) showed a wide MIC range (0.1-12.8µg/mL), suggesting a resistance mechanism toward this antimicrobial that was not elicited by enrofloxacin or ciprofloxacin (the other quinolones tested). Erythromycin showed the highest MIC90 such that its use against Mycoplasma agalactiae is not recommended. Finally, Mycoplasma agalactiae isolates obtained from goat herds with clinical symptoms of contagious agalactia featured higher MIC90 and MIC50 (concentration at which growth of 50% of the isolates is inhibited) values for many of the antimicrobials compared with isolates from asymptomatic animals. The relationship between the extensive use of antimicrobials in herds with clinical contagious agalactia and variations in MIC requires further study. PMID:24035026
Paterna, A; Sánchez, A; Gómez-Martín, A; Corrales, J C; De la Fe, C; Contreras, A; Amores, J
An increased incidence of Mycoplasma pneumoniae infections was reported in 2011 in two cities in France, Bordeaux and Caen. Two complementary molecular typing methods, PCR-RFLP on adhesin P1 and multilocus variable number tandem repeat analysis (MLVA), were used to determine whether this phenomenon was clonal. In 2011, the percentage of M. pneumoniae-positive patients doubled in both cities compared with 2010. Macrolide resistance remained stable at 8.3% of patients. Eighteen MLVA types were identified among 94 M. pneumoniae-positive specimens, demonstrating that the phenomenon was multiclonal. Types P, J, U, X and E were the most frequent and 81.6% of the strains were adhesin P1 type 1. PMID:23279613
Five groups of light hybrid SPF chicks were inoculated at 1 day of age with different strains of Mycoplasma iowae via the right thoracic air sac and right foot pad. Control birds were inoculated with sterile mycoplasma broth at the same sites.The chicks were observed daily for clinical signs and at 3 and 6 weeks approximately half the birds in
The polymerase chain reaction (PCR) with primers complementary to the 16S rRNA genes was used to detect avian mycoplasmas. A primer pair designed for the detection of human and rodent mycoplasmal species was examined for its ability to detect the most important avian mycoplasmas. After testing the respective reference strains, we found that Mycoplasma iowae, Mycoplasma meleagridis and Mycoplasma synoviae
I. Kiss; Katalin Matiz; Éva Kaszanyitzky; Yleana Chávez; K.-E. Johansson
Mycoplasma gallisepticum is an important pathogen of chickens and turkeys that causes considerable economic losses to the poultry industry worldwide. The reemergence of M. gallisepticum outbreaks among poultry, the increased use of live M. gallisepticum vaccines, and the detection of M. gallisepticum in game and free-flying song birds has strengthened the need for molecular diagnostic and strain differentiation tests. Molecular techniques, including restriction fragment length polymorphism of genomic DNA (RFLP) and PCR-based random amplification of polymorphic DNA (RAPD), have already been utilized as powerful tools to detect intraspecies variation. However, certain intrinsic drawbacks constrain the application of these methods. The main goal of this study was to determine the feasibility of using an M. gallisepticum-specific gene encoding a phase-variable putative adhesin protein (PvpA) as the target for molecular typing. This was accomplished using a pvpA PCR-RFLP assay. Size variations among PCR products and nucleotide divergence of the C-terminus-encoding region of the pvpA gene were the basis for strain differentiation. This method can be used for rapid differentiation of vaccine strains from field isolates by amplification directly from clinical samples without the need for isolation by culture. Moreover, molecular epidemiology of M. gallisepticum outbreaks can be performed using RFLP and/or sequence analysis of the pvpA gene. PMID:11326008
Liu, T; García, M; Levisohn, S; Yogev, D; Kleven, S H
Live Mycoplasma gallisepticum (MG) vaccines have been USDA approved and licensed for use in commercial layer chickens since 1988; however, egg production and egg quality data exist only for the F strain of MG. Information pertinent to the effects of ts-11 MG on egg and eggshell quality parameters, as well as egg size distribution, is lacking. In this study, pullets were inoculated at 10 wk of age with ts-11 strain MG and placed in biological isolation units at 10 birds/unit. Hen-day egg production, eggshell strength, Haugh unit score, pimpling incidence, and blood/meat spot incidence were monitored and recorded in each trial through a 45-wk production cycle. Further, eggs from all treatments were collected daily, Monday-Thursday, and individually weighed. Results of this study indicate that no significant difference was observed between the treatments for the parameters measured or for egg size distribution. Therefore, these data should lessen producers' concerns pertaining to the impact of ts-11 strain MG on egg production, egg and eggshell quality parameters, and egg size distribution. PMID:11007009
Branton, S L; Lott, B D; May, J D; Maslin, W R; Pharr, G T; Bearson, S D; Collier, S D; Boykin, D L
Colonization of the avian respiratory tract with Mycoplasma gallisepticum results in a profound inflammatory response in the trachea, air sacs, conjunctiva, and lungs. A live attenuated M. gallisepticum vaccine strain, GT5, was previously shown to be protective in chickens upon challenge; however, the mechanisms by which this vaccine and others confer protection remain largely unknown. The current study evaluated several potential correlates of GT5 vaccine-mediated immune protection following challenge with the pathogenic M. gallisepticum strain Rlow. GT5-vaccinated chickens developed mild tracheal lesions, consisting of few and scattered, discrete, lymphofollicular aggregates in the lamina propria. In addition, low numbers of aggregated B, CD4+, and CD8+ cells were observed to infiltrate the trachea, in stark contrast to the large numbers infiltrating the tracheas of sham-vaccinated chickens challenged with Rlow. Lymphofollicular aggregates were rarely observed prior to day 12 postchallenge in sham-vaccinated chickens. Instead, they contained an increasingly more cellular inflammatory response characterized by expansion of the lamina propria by lymphoplasmacytic and histiocytic infiltrates. This was due in part to expansion of interfollicular zones by large numbers of infiltrating CD4+ and CD8+ cells and a sizeable population of immunoglobulin A (IgA)- and IgG-secreting plasma cells. GT5-vaccinated chickens also had higher serum IgG concentrations, and significantly higher numbers of M. gallisepticum-specific IgG- and IgA-secreting plasma/B cells within the trachea, than did sham-vaccinated chickens. These responses were observed as early as day 4 postchallenge, indicating the importance of antibody-mediated clearance of mycoplasma in GT5-vaccinated chickens.
Javed, Mohammed A.; Frasca, Salvatore; Rood, Debra; Cecchini, Katharine; Gladd, Martha; Geary, Steven J.; Silbart, Lawrence K.
To determine the Mycoplasma gallisepticum (MG) rapid serum plate agglutination (RSPA) test response of broiler breeders after ts-11 strain vaccination, 55 Cobb pullets derived from a nonvaccinated, MG-negative, commercial, broiler breeder grandparent flock were monitored from 8 to 20 wk of age (over a 12-wk trial period). To evaluate the effect of lateral spread of the ts-11 vaccine strain on RSPA test results from commingled and adjacently penned birds, treatment groups included (A) birds vaccinated with ts-11strain MG at 8 wk of age, (B) commingled nonvaccinates in the same pen as the vaccinated birds, (C) nonvaccinates in a second pen separated from the first pen by a distance of 2 m, and (D) birds vaccinated with ts-11 strain MG at 8 wk of age and kept in a separate room. Rapid serum plate agglutination tests were performed once a week for 6 wk and then every 2 wk for 6 more wk, postvaccination. A polymerase chain reaction (PCR) assay specific fbr ts-11 strain MG was used to confirm vaccination, and a second PCR specific for non-ts-11 strain MG was used to confirm the absence of field infection. Seroconversion was first detected by the RSPA test 2 wk postvaccination and attained maximum positive rates of 58% at 12 wk postvaccination in treatment A and 60% at 8 wk postvaccination in treatment D. Seroconversion rates in nonvaccinated, commingled pullets was 10% at 5 wk and 30% at 12 wk after the vaccination of pen mates. The ts-11-specific PCR detected the vaccine strain in 80%-100% of the vaccinated birds 2 wk after vaccination. One of 15 nonvaccinated birds penned 2 m from vaccinated birds yielded ts-11 by PCR assay 12 wk after vaccination, which indicates that the spread of ts-11 over short distances may be possible in situations in which there is a common caretaker. PCR on tracheal swabs taken 12 wk postvaccination detected ts-l1 in 50% and 60% of the vaccinated birds in treatments A and D, respectively; in 30% of the commingled nonvaccinates; and in 6.6% of the separately penned nonvaccinates. In contrast, choanal swabs collected from vaccinated birds at 12 wk were 21% and 40% PCR positive for ts-11 strain MG, while those from nonvaccinates were negative. All samples were PCR negative for field strain MG. The pattern of seroconversion as measured by RSPA test in small groups of broiler breeders was different from that previously reported for leghorns. Lateral spread of the ts-11 strain to commingled pen mates occurred rapidly, causing RSPA seroconversion patterns that mimicked those of the vaccinated pen mates. PMID:15839426
Collett, S R; Thomson, D K; York, D; Bisschop, S P R
Two trials were conducted concurrently to determine and compare, blood pH, blood gases, hematocrit and hemoglobin in F-strainMycoplasma gallisepticum (FMG) inoculated layers and FMG contact- infected broilers. At the termination of the study, FMG-inoculated layers had the highest partial pressure of O and the lowest partial pressure of CO as compared with the other treatment groups. Blood pH values
H. A. Olanrewaju; J. L. Purswell; S. D. Collier; S. L. Branton
Mycoplasma alligatoris causes lethal invasive disease of alligators and caimans. A homolog of the nagH gene, encoding a hyaluronidase secreted by Clostridium perfringens, and a C. perfringens hyaluronidase nagI or nagK pseudogene were discovered in the M. alligatoris genome. The nagH gene was detected by PCR in the closest relative of M. alligatoris, Mycoplasma crocodyli, but not in 40 other species representing the Mycoplasmahominis, Mycoplasma pneumoniae, and Spiroplasma phylogenetic clusters. The hyaluronidase activity in the cellular fraction of M. alligatoris and M. crocodyli SP4 broth cultures was equivalent to 10?16 U of Streptomyces hyalurolyticus hyaluronidase CFU?1. Negligible activity was present in the cell-free supernatant fraction. No chondroitinase activity was detected. There is also a novel homolog of the nanI gene, which encodes a sialidase secreted by C. perfringens, in the M. alligatoris genome. The signature YRIP and SXDXGXTW motifs and catalytic residues of the clostridial sialidase are conserved in the mycoplasmal gene, but the leader sequence necessary for its secretion by C. perfringens is absent. The gene was not detected by PCR in any other mycoplasma. Potent cell-associated sialidase activity was present in M. alligatoris colonies on agar but not in the cell-free supernatants of broth cultures or in M. crocodyli. The presence of hyaluronidase and sialidase in M. alligatoris is consistent with the rapid invasiveness and necrotizing effects of this organism, and the lack of sialidase in M. crocodyli is consistent with its comparatively attenuated virulence. This genetic and biochemical evidence suggests that the spreading factors hyaluronidase and sialidase, a combination unprecedented in mycoplasmas, are the basis of the virulence of M. alligatoris.
Background Mycoplasma hyopneumoniae is the primary cause of enzootic pneumonia in pigs. Although vaccination is an important control tool, the results observed under field conditions are variable. This may be due to antigenic differences between the strains circulating in pig herds and the vaccine strain. This study compared the protective efficacy of four bacterins against challenge infection with a highly virulent field strain of M. hyopneumoniae. Seventy eight, one-week old piglets were randomly assigned to five treatment groups (A, B, C, D, E), 14 piglets each, and a negative control group (F) consisting of 8 piglets. All pigs were injected at 1 (D7) and 4 weeks of age (D28), with 2 ml of either a placebo or a bacterin based on selected M. hyopneumoniae strains, namely A (F7.2C), B (F20.1L), C (B2V1W20 1A-F), D (J strain), E (placebo; positive control), F (placebo; negative control). At D56, all pigs except those of group F were challenged intratracheally with 7 ml culture medium containing 107 CCU/ml of M. hyopneumoniae strain F7.2C. All pigs were euthanized and necropsied at D84. The severity of coughing and pneumonia lesions were the main parameters. Immunofluorescence (IF) testing, nested PCR testing of bronchoalveolar lavage (BAL) fluid and serology for M. hyopneumoniae were also performed. Results The different bacterins only slightly improved clinical symptoms (average 0.38 in vaccinated groups vs. 0.45 in group E) and histopathological lung lesions (average 3.20 in vaccinated groups vs. 3.45 in group E), but did not improve macroscopic lung lesions (score 4.30 vs. 4.03 in group E). None of the vaccines was significantly and/or consistently better or worse than the other ones. All bacterins evoked a serological response in the vaccinated animals. All pigs, except those from group F, were positive with nPCR in BAL fluid at D84. Conclusion The bacterins did not induce a clear overall protection against challenge infection, and there were no significant differences in protective efficacy between bacterins containing homologous and heterologous M. hyopneumoniae strains. Further research is necessary to better characterize the antigens involved in protection and to elucidate the protective immunity responses following M. hyopneumoniae vaccination and/or infection.
The effects of 6/85-strainMycoplasma gallisepticum (6/85MG) inoculation alone or in conjunction with F-strain M. gallisepticum (FMG) overlays and their timing on the blood characteristics of commercial egg-laying hens were investigated. Control birds received sham inoculations at 10 wk of age. Birds in a second treatment group were inoculated with 6/85MG at 10 wk of age, those in a third treatment group were inoculated with 6/85MG at 10 wk followed by an overlay inoculation of FMG at 22 wk, and those in a fourth treatment group were inoculated with 6/85MG at 10 wk followed by an overlay inoculation of FMG at 45 wk. Parameters investigated at 24, 32, 43, and 47 wk were hematocrit, plasma total protein, and serum calcium, triglycerides, and cholesterol. No significant treatment effects were noted for hematocrit, serum triglycerides, or serum cholesterol. However, at wk 32, plasma protein was greater in birds that received 6/85MG at 10 wk or 6/85MG at 10 wk and FMG at 22 wk in comparison to controls. Also, at wk 47, serum calcium concentration was greater in birds that received 6/85MG at 10 wk and FMG at 45 wk compared with controls and those that received 6/85MG at 10 wk and FMG at 22 wk. These results suggest that the prelay inoculation of pullets with 6/85MG may subsequently elevate plasma protein, and in conjunction with an FMG overlay at 45 wk, may increase serum calcium concentrations in laying hens. PMID:18809862
Peebles, E D; Viscione, K A; Branton, S L; Vance, A M; Gerard, P D; Whitmarsh, S K
Bacteriological methods applicable to the characterization and differentiation of Mycoplasma species were investigated. After appropriate modification and standardization, these methods were tested on 53 Mycoplasmastrains comprising more than 22 species....
B. B. Aluotto R. G. Wittler C. O. Williams J. Faber
A series of meatal swabs, taken from 17 men over a period of 17 months during their tour at an Antarctic base was examined for mycoplasmas. The number of organisms isolated never exceeded 104 and not every specimen from each man yielded mycoplasmas. Nevertheless, Mycoplasmahominis was isolated from 71% and T-mycoplasmas from 59% of the men at some time during their stay. M. hominis persisted in the presence of serum IHA antibody titres of 1/64. Three subjects yielded only M. hominis and one only T-mycoplasmas. Six men had already spent a year at the base when the study began and mycoplasmas were still being isolated from some of them at the end of a 31 month period of isolation. The persistence of mycoplasmas in the male genital tract can therefore be independent of sexual contact. Two modes of persistence are suggested; either a few men act as carriers and reinfect the others by contaminating their environment, or as seems more likely, most men have chronic infections.
Holmes, M. J.; Furr, Patricia M.; Taylor-Robinson, D.
Initial efforts by the poultry industry at controlling and containing Mycoplasma gallisepticum (MG) included testing and slaughter of reactor flocks. Ultimately, using the aforementioned measures coupled with heat treatment of hatching eggs together with biosecurity and biosurveillance procedures, ...
The present study describes the use of microarray technology for rapid identification and differentiation of Mycoplasma mycoides subsp. mycoides from other mycoplasmas that may be pathogenic to ruminants, including those of the Mycoplasma mycoides cluster, genetically and antigenically strictly correlated with Mycoplasma mycoides subsp. mycoides. A microarray containing genetic sequences of 55 different bacterial species from Acholeplasma, Mycoplasma, Spiroplasma and Ureaplasma genera was constructed. Sequences to genes of interest were collected in FASTA format from NCBI. The collected sequences were processed with OligoPicker software. Oligonucleotides were then checked for their selectivity with BLAST searches in GenBank. The microarray was tested with ATCC/NCTC strains of Mycoplasma spp. of veterinary importance in ruminants including Mycoplasma belonging to the mycoides cluster as well as Mycoplasma mycoides subsp. mycoides and Mycoplasma mycoides subsp. capri field strains. The results showed that but one ATCC/NCTC reference strains hybridized with their species-specific sequences showed a profile/signature different and distinct from each other. The heat-map of the hybridization results for the nine genes interrogated for Mycoplasma mycoides subsp. mycoides demonstrated that the reference strainMycoplasma mycoides subsp mycoides PG1 was positive for all of the gene sequences spotted on the microarray. CBPP field, vaccine and reference strains were all typed to be M. mycoides subsp. mycoides, and seven of the nine strains gave positive hybridization results for all of the nine genes. Two Italian strains were negative for some of the genes. Comparison with non-Mycoplasma mycoides subsp. mycoides reference strains showed some positive signals or considerable homology to Mycoplasma mycoides subsp. mycoides genes. As expected, some correlations were observed between the strictly genetically and antigenically correlated Mycoplasma mycoides subsp. mycoides and Mycoplasma mycoides subsp. capri strains. Specifically, we observed that some Italian Mycoplasma mycoides subsp. mycoides strains were positive for two out of the three Mycoplasma mycoides subsp. capri genes, differently from what has been observed for other European or African Mycoplasma mycoides subsp. mycoides strains. This study highlighted the use of microarray technology as a simple and effective method for a single-step identification and differentiation of Mycoplasma mycoides subsp. mycoides from other mycoplasmas that may be pathogenic to ruminants, including those of the Mycoplasma mycoides cluster, genetically and antigenically strictly correlated with Mycoplasma mycoides subsp. mycoides. The opportunity to discriminate several mycoplasmas in a single analysis enhances diagnostic rapidity and may represent a useful tool to screen occasionally mycoplasmas affecting animal farming in territories where diagnostic laboratory support is limited. The heat-map of the hybridization results of the comparative genomic hybridizations DNA-designed chip clearly indicates that the microarray performs well for the identification of the tested Mycoplasma mycoides subsp. mycoides reference and field strains, discriminating them from other mycoplasmas. PMID:22271459
Tonelli, A; Sacchini, F; Krasteva, I; Zilli, K; Scacchia, M; Beaurepaire, C; Nantel, A; Pini, A
Duplicate vaginal swabs were collected from 100 women, and comparisons were made between an in-house broth-agar culture system and a commercially available kit, the Mycoplasma IST kit (bioMerieux), for the detection of Mycoplasmahominis and Ureaplasma urealyticum. There was good agreement between the two systems for detection of the genital mycoplasmas in terms of sensitivity, with values of >92% being
ALISON CLEGG; MEGAN PASSEY; MITION YOANNES; ANDAUDREY MICHAEL
Mycoplasma gallisepticum (MG), a reproductive/respiratory pathogen in poultry, has been implicated in suboptimum egg production and decreased hatchability. Commercial layer hens raised in a controlled environment were inoculated with the S6 strain of MG at 20 wk of age. The S6 inoculation had no effect on bird weight, egg production, digestive tract weight and length, or histopathologic lesion scores, although significant differences were noted in the lengths and weights of various portions of the reproductive tract. This study shows that S6MG inoculation does not detrimentally affect layer hen performance when in the absence of environmental stressors customary to a caged layer facility. PMID:12713163
Parker, T A; Branton, S L; Jones, M S; Peebles, E D; Gerard, P D; Willeford, K O; Pharr, G T; Maslin, W R
The polymerase chain reaction (PCR) technique was compared with culture for the detection ofUreaplasma urealyticum, Mycoplasmahominis, andMycoplasma genitalium in clinical samples (vaginal secretions, throat and endotracheal secretions, and skin swabs) obtained from 47 high-risk pregnant women peripartum and eight newborn infants. Detection using PCR with homologous primers was highly specific, as a product with the expected length was consistently
N. Luki; P. Lebel; M. Boucher; B. Doray; J. Turgeon; R. Brousseau
Mycoplasma synoviae is a major pathogen of chickens and turkeys, causing economic losses to the poultry industry worldwide. In this study, we validated and applied polymerase chain reaction (PCR) and DNA sequence analysis on the N-terminal end of the hemagglutinin encoding gene vlhA as an alternative for the detection and initial typing of field strains of M. synoviae in commercial poultry. PCR primers were tested against isolates of M. synoviae from various sources along with other avian mycoplasma and other bacterial species. The vlhA gene-targeted PCR assay was highly specific in the identification of M. synoviae, with a detection limit of 4.7 x 10(2) color changing units/ml. DNA sequence analysis of amplified products was also conducted to validate the potential for typing M. synoviae strains using the N-terminal region of the vlhA gene. To evaluate the test, we applied the PCR assay to tracheal swabs collected from chickens challenged with M. synoviae strain K1968 and compared the results to the serologic detection. The PCR assay was also evaluated directly on tracheal samples collected from commercial layers. Overall, this vlhA gene-targeted PCR is a useful tool for detection and initial typing of M. synoviae and can be applied in the preliminary identification of M. synoviae isolates directly from clinical samples. PMID:15529983
Hong, Yang; García, Maricarmen; Leiting, Victoria; Bencina, Dulan; Dufour-Zavala, Louise; Zavala, Guillermo; Kleven, Stanley H
The macrolide class of antibiotics, including tylosin and tilmicosin, is widely used in the veterinary field for prophylaxis and treatment of mycoplasmosis. In vitro susceptibility testing of 50 strains of M. gallisepticum isolated in Israel during the period 1997-2010 revealed that acquired resistance to tylosin as well as to tilmicosin was present in 50% of them. Moreover, 72% (13/18) of the strains isolated from clinical samples since 2006 showed acquired resistance to enrofloxacin, tylosin and tilmicosin. Molecular typing of the field isolates, performed by gene-target sequencing (GTS), detected 13 molecular types (I-XIII). Type II was the predominant type prior to 2006 whereas type X, first detected in 2008, is currently prevalent. All ten type X strains were resistant to both fluoroquinolones and macrolides, suggesting selective pressure leading to clonal dissemination of resistance. However, this was not a unique event since resistant strains with other GTS molecular types were also found. Concurrently, the molecular basis for macrolide resistance in M. gallisepticum was identified. Our results revealed a clear-cut correlation between single point mutations A2058G or A2059G in domain V of the gene encoding 23S rRNA (rrnA, MGA_01) and acquired macrolide resistance in M. gallisepticum. Indeed, all isolates with MIC ? 0.63 ?g/mL to tylosin and with MIC ? 1.25 ?g/mL to tilmicosin possess one of these mutations, suggesting an essential role in decreased susceptibility of M. gallisepticum to 16-membered macrolides.
'Candidatus Mycoplasma haemobos', sometimes causative of bovine infectious anemia at various extents, has been demonstrated throughout the world. Here, we show two distinct types of 'Ca. M. haemobos' are distributed among cattle in Japan, by examining the primary and secondary structures of the 16S-23S rRNA intergenic spacer region that has been shown to be a stable genetic marker for mycoplasma species. Our results may explain differences in severity of anemic condition as well as provide a genetic marker for an epidemiological study of bovine hemoplasma infections. PMID:23064449
Background In Mycoplasma synoviae, type strain WVU 1853, a single member of the haemaglutinin vlhA gene family has been previously shown to be expressed. Variants of vlhA are expressed from the same unique vlhA promoter by recruiting pseudogene sequences via site-specific recombination events, thus generating antigenic variability. Using a bacterial stock of M. synoviae WVU 1853 that had been colony purified thrice and maintained in our laboratory at low passage level, we previously identified a vlhA gene-related partial coding sequence, referred to as MS2/28.1. The E. coli-expressed product of this partial coding sequence was found to be immunodominant, suggesting that it might be expressed. Results Reverse transcription-PCR amplification (RT-PCR), using a sense primer located at the 5'-end region of the expected vlhA transcript and a reverse primer located at the 3' end of MS2/28.1 coding sequence, yielded a consistent amplification product showing that MS2/28.1 was indeed transcribed. Nucleotide sequence analysis of the RT-PCR product identified an 1815-nucleotide full-length open reading frame (ORF), immediately preceded by a nucleotide sequence identical to that previously reported for expressed vlhA genes. PCR amplifications using genomic DNA isolated from single colonies further confirmed that the full-length ORF of MS2/28.1 was located downstream of the unique vlhA promoter sequence. The deduced 604-amino acid (aa) sequence showed a perfect sequence identity to the previously reported vlhA expressed genes along the first 224 residues, then highly diverged with only 37.6% aa identity. Despite the fact that this M. synoviae clone expressed a highly divergent and considerably shorter C-terminal haemagglutinin product, it was found to be expressed at the surface of the bacterium and was able to haemagglutinate chicken erythrocytes. Importantly, the E. coli-expressed C-terminal highly divergent 60 residues of MS2/28.1 proved haemagglutination competent. Conclusions In contrast to the previously characterized vlhA expressedvariants, MS2/28.1 displayed a highly divergent sequence, while still able to haemagglutinate erythrocytes. Overall, the data provide an indication as to which extent the M. synoviae vlhA gene could vary its antigenic repertoire.
In two consecutive trials of the current study, the effect of the age of application of S6-strainMycoplasma gallisepticum (S6MG) inoculation on the blood characteristics of commercial layers housed and maintained under controlled conditions was determined. The ages of inoculation compared were tho...
The effects of F-strainMycoplasma gallisepticum (FMG) inoculation and 1.5 % supplemental dietary poultry fat (PF) on the digestive and reproductive organ characteristics of commercial layers at 58 wk of age were investigated. Sham and FMG inoculations were administered at 12 (before lay) and 22 (e...
The effects of 2 supplemental levels of dietary poultry fat (PF) and the combination of PF, phytase (PHY) and 25-hydroxycholecalciferol [25(OH)D] on the blood characteristics of commercial layers inoculated with F-strainMycoplasma gallisepticum (FMG) were investigated in 2 trials. Sham and FMG ino...
In 3 trials, the effects of dietary supple mentation with phytase (PHY) and 25-hydroxycholecalciferol (25-D3) on the digestive and reproductive organ characteristics of commercial layers that were inoculated pre-lay (12 wk of age) or at the onset of lay (22 wk of age) with F-strainMycoplasma gallis...
The polymerase chain reaction (PCR) was evaluated to detect mycoplasma contamination of avian live virus vaccines. The specificity of the primers showed that 34 strains belonging to nine species of avian mycoplasma DNA could be detected. The sensitivity of PCR to detect mycoplasma DNA was 100·2colony forming units (cfu) ofMycoplasma synoviaeand 100·7cfu ofMycoplasma gallisepticum. WhenM. synoviaeandM. gallisepticumwere spiked into several
Increasing use of Mycoplasma gallisepticum (MG) live vaccines has led to a need for a rapid test for differentiation of MG field strains from the live vaccine strains ts-11 and 6/85. We examined the differentiating potential of diagnostic polymerase chain reaction (PCR) primers targeted to the gene mgc2, encoding a cytadherence-related surface protein uniquely present in MG. The mgc2-PCR diagnostic primers are specific for MG in tests of all avian mycoplasmas or bacteria present in the chicken trachea and are sensitive enough to readily detect MG in tracheal swabs from field outbreaks. Differentiation of vaccine strain ts-11 was based on identification of restriction enzyme sites in the 300-base-pair (bp) mgc2-PCR amplicon present in ts-11 and missing in MG isolates from field outbreaks in Israel. Restriction sites for the enzymes HaeII and SfaN1 were identified in the amplified region in strain ts-11 and were not found in 28 field isolates of MG, comprising a representative cross section of all the MG isolates from the period 1997-2003. In practice, differential diagnosis of MG is achieved within 1 day of submission of tracheal swab samples by mgc2-PCR amplification and restriction of the amplicon with HaeII, giving a 270-bp fragment for ts-11 or no restriction for other MG strains tested. Application of the mgc2-PCR-restriction fragment length polymorphism (mgc2-PCR-RFLP) assay enabled differential diagnosis of both components of a mixture of ts-11 and non-ts-11 DNA, detecting the field strain in the presence of a large excess of ts-11. The test was successfully applied in vivo for monitoring vaccinates in a ts-11 vaccine trial. In principle, the test may also be used to identify the 6/85 vaccine strain, which yields a 237-bp product, readily differentiated from the approximately 300-bp PCR product of all other strains tested. Further testing of field isolates will be necessary to determine the applicability of this test in the United States and other countries. PMID:16094829
Although they lack a cell wall, mycoplasmas do possess a glycocalyx. The interactions between the glycocalyx, mycoplasmal surface proteins and host complement were explored using the murine pathogen Mycoplasma pulmonis as a model. It was previously shown that the length of the tandem repeat region of the surface lipoprotein Vsa is associated with susceptibility to complement-mediated killing. Cells producing a long Vsa containing about 40 repeats are resistant to complement, whereas strains that produce a short Vsa of five or fewer repeats are susceptible. We show here that the length of the Vsa protein modulates the affinity of the M. pulmonis EPS-I polysaccharide for the mycoplasma cell surface, with more EPS-I being associated with mycoplasmas producing a short Vsa protein. An examination of mutants that lack EPS-I revealed that planktonic mycoplasmas were highly susceptible to complement killing even when the Vsa protein was long, demonstrating that both EPS-I and Vsa length contribute to resistance. In contrast, the mycoplasmas were resistant to complement even in the absence of EPS-I when the cells were encased in a biofilm. PMID:22504437
Bolland, Jeffrey R; Simmons, Warren L; Daubenspeck, James M; Dybvig, Kevin
Summary We have developed a simple method for rapid detection of mycoplasma contamination in cell cultures using SYBR Green-based\\u000a real-time polymerase chain reaction (PCR). To detect eight common contaminant mollicutes, including Mycoplasma (M. arginini, M. fermentans, M. orale, M. hyorhinis, M. hominis, M. salivarium, M. pirum) and Acholeplasma laidlawii, four primers were prepared based on the 23S rRNA regions. Using these
Background Among Coagulase-Negative Staphylococci (CoNS), Staphylococcus hominis represents the third most common organism recoverable from the blood of immunocompromised patients. The aim of this study was to characterize biofilm formation, antibiotic resistance, define the SCCmec (Staphylococcal Chromosomal Cassette mec) type, and genetic relatedness of clinical S. hominis isolates. Methodology S. hominis blood isolates (n?=?21) were screened for biofilm formation using crystal violet staining. Methicillin resistance was evaluated using the cefoxitin disk test and the mecA gene was detected by PCR. Antibiotic resistance was determined by the broth microdilution method. Genetic relatedness was determined by pulsed-field gel electrophoresis (PFGE) and SCCmec typed by multiplex PCR using two different methodologies described for Staphylococcus aureus. Results Of the S. hominis isolates screened, 47.6% (10/21) were categorized as strong biofilm producers and 23.8% (5/21) as weak producers. Furthermore, 81% (17/21) of the isolates were methicillin resistant and mecA gene carriers. Resistance to ampicillin, erythromycin, and trimethoprim was observed in >70% of isolates screened. Each isolate showed a different PFGE macrorestriction pattern with similarity ranging between 0–95%. Among mecA-positive isolates, 14 (82%) harbored a non-typeable SCCmec type: eight isolates were not positive for any ccr complex; four contained the mec complex A ccrAB1 and ccrC, one isolate contained mec complex A, ccrAB4 and ccrC, and one isolate contained the mec complex A, ccrAB1, ccrAB4, and ccrC. Two isolates harbored the association: mec complex A and ccrAB1. Only one strain was typeable as SCCmec III. Conclusions The S. hominis isolates analyzed were variable biofilm producers had a high prevalence of methicillin resistance and resistance to other antibiotics, and high genetic diversity. The results of this study strongly suggested that S. hominis isolates harbor new SCCmec structural elements and might be reservoirs of ccrC1 in addition to ccrAB1 and mec complex A.
The precarious status of desert (Gopherus agassizii) and gopher (G. polyphemus) tortoises has resulted in conservation efforts that now include health assessment as an important component of management decision-making. Mycoplasmal upper respiratory tract disease (URTD) is one of very few diseases in chelonians for which comprehensive and rigorously validated diagnostic tests exist. In this study, serum samples obtained from eight Gopherus tortoises documented at necropsy to (i) be enzyme-linked immunosorbent assay (ELISA) seropositive using the PS6 antigen, (ii) be infected with Mycoplasma agassizii as indicated by direct isolation of the pathogen from the respiratory surfaces, and (iii) have histological lesions of mycoplasmal URTD were used to evaluate four distinct clinical isolates of M. agassizii as antigens for ELISA and Western blot analyses. Each animal sample reacted in the Western blot with its homologous M. agassizii strain, but recognition of heterologous M. agassizii strains was variable. Further, individual animals varied significantly with respect to the specific proteins recognized by the humoral immune response. An additional 114 Gopherus serum samples were evaluated using ELISA antigens prepared from the four distinct M. agassizii strains; A405 values were significantly correlated (r2 goodness of fit range, 0.708 to 0.771; P < 0.0001) for all antigens tested. The results confirm that strain variation is responsible for the observed differences between Western blot binding patterns. Thus, reliance on a single M. agassizii strain as an antigen in Western blot assays may provide false-negative results. This could have adverse consequences for the well-being of these environmentally sensitive hosts if false-negative animals were relocated to sites consisting of true-negative populations.
Wendland, Lori D.; Klein, Paul A.; Jacobson, Elliott R.; Brown, Mary B.
Certain Mycoplasma spp. are pathogens of poultry, but little is known of the role of mycoplasmas in disease of birds of prey. Species-specific polymerase chain reactions (PCRs) for the detection of the poultry pathogens Mycoplasma gallisepticum, Mycoplasma imitans, Mycoplasma iowae, Mycoplasma meleagridis and Mycoplasma synoviae were therefore evaluated for use in birds of prey. The specificities of the PCR methods
Summary: The history, replication, genetics, characteristics (both biological and physical), and factors involved in the pathogenesis of Mycoplasma genitalium are presented. The latter factors include adhesion, the influence of hormones, motility, possible toxin production, and immunological responses. The preferred site of colonization, together with current detection procedures, mainly by PCR technology, is discussed. The relationships between M. genitalium and various diseases are highlighted. These diseases include acute and chronic nongonococcal urethritis, balanoposthitis, chronic prostatitis, and acute epididymitis in men and urethritis, bacterial vaginosis, vaginitis, cervicitis, pelvic inflammatory disease, and reproductive disease in women. A causative relationship, or otherwise strong association, between several of these diseases and M. genitalium is apparent, and the extent of this, on a subjective basis, is presented; also provided is a comparison between M. genitalium and two other genital tract-orientated mollicutes, namely, Mycoplasmahominis, the first mycoplasma of human origin to be discovered, and Ureaplasma species. Also discussed is the relationship between M. genitalium and infertility and also arthritis in both men and women, as is infection in homosexual and immunodeficient patients. Decreased immunity, as in HIV infections, may enhance mycoplasmal detection and increase disease severity. Finally, aspects of the antimicrobial susceptibility and resistance of M. genitalium, together with the treatment and possible prevention of mycoplasmal disease, are discussed.
Against 198 viridans group streptococci, 25 Streptococcus bovis strains, and 5 Cardiobacterium hominisstrains, MICs of DX-619, a des-F(6)-quinolone, were between 0.004 and 0.25 ?g/ml. These MICs were lower than those of other quinolones (?0.008 to >32 ?g/ml). ?-Lactam MICs were between ?0.008 and 16 ?g/ml. Azithromycin resistance was found in most species, while most were telithromycin susceptible. Glycopeptides and linezolid were active against viridans group strains but inactive against C. hominis.
Kosowska-Shick, Klaudia; Smith, Kathy; Bogdanovich, Tatiana; Ednie, Lois M.; Jones, Ronald N.; Appelbaum, Peter C.
Strains 1517(T) and 61D(T) were characterized by phenotypic and molecular taxonomic methods. These Gram-positive lactic acid bacteria were homo-fermentative, facultatively anaerobic short rods. They were phylogenetically related to the genus Lactobacillus according to 16S rRNA gene sequence analysis, with 99 % similarity between strain 1517(T) and the type strain of Lactobacillus gigeriorum, and 98.6, 98.5 and 98.4 % between strain 61D(T) and Lactobacillus gasseri, Lactobacillus taiwanensis and Lactobacillus johnsonii, respectively. Multilocus sequence analysis and metabolic analysis of both strains showed variation between the two strains and their close relatives, with variation in the position of the pheS and rpoA genes. The DNA-DNA relatedness of 43.5 % between strain 1517(T) and L. gigeriorum, and 38.6, 29.9 and 39.7 % between strain 61D(T) and L. johnsonii, L. taiwanensis and L. gasseri, respectively, confirmed their status as novel species. Based on phenotypic and genotypic characteristics, two novel species of Lactobacillus are proposed: Lactobacillus pasteurii sp. nov., with 1517(T) ( = CRBIP 24.76(T) = DSM 23907(T)) as the type strain, and Lactobacillus hominis sp. nov., with 61D(T) (=CRBIP 24.179(T) = DSM 23910(T)) as the type strain. PMID:22328611
Glycopeptides and linezolid are the most widely used antibiotics to treat infections by methicillin-resistant Staphylococcus spp. We report the presence of various isolates of methicillin-resistant S. hominis subsp. hominis with resistance to linezolid and reduced susceptibility to glycopeptides. We studied ten blood culture isolates of S. hominis subsp. hominis from nine patients admitted to our hospital. Etest was used to study susceptibility to antibiotics commonly prescribed against staphylococci. Domain V region of the 23S rRNA gene was amplified and sequenced to detect possible mutations that confer resistance to linezolid. Pulsed-field gel electrophoresis (PFGE) was used for the clonality study of isolates. All isolates were resistant to oxacillin, gentamicin, levofloxacin, cotrimoxazole, and linezolid, and susceptible to tigecycline and daptomycin. Nine of the isolates were resistant to erythromycin and clindamycin, and showed heterogeneous resistance to glycopeptides. C2190T, G2603T, and G2474T mutations were detected in domain V of the 23S rRNA gene. PFGE showed the presence of two different clones. This report alerts to the possible appearance of clinical strains of methicillin-resistant staphylococci with intermediate resistance to glycopeptides, resistance to linezolid, and multiple resistance to other second-line antibiotics. PMID:19876662
Sorlozano, A; Gutierrez, J; Martinez, T; Yuste, M E; Perez-Lopez, J A; Vindel, A; Guillen, J; Boquete, T
Mycoplasma mycoides subsp. mycoidesSC (MmymySC)is the etiological agent of contagious bovine pleuropneumonia (CBPP), a highly contagious respiratory disease in cattle. The genome of Mmymy SC type strain PG1(T) has been sequenced to map all the genes and to facilitate further studies regarding the cell function of the organism and CBPP. The genome is characterized by a single circular chromosome of 1211703 bp with the lowest G+C content (24 mole%)and the highest density of insertion sequences (13% of the genome size)of all sequenced bacterial genomes. The genome contains 985 putative genes, of which 72 are part of insertion sequences and encode transposases. Anomalies in the GC-skew pattern and the presence of large repetitive sequences indicate a high genomic plasticity. A variety of potential virulence factors was identified, including genes encoding putative variable surface proteins and enzymes and transport proteins responsible for the production of hydrogen peroxide and the capsule, which is believed to have toxic effects on the animal. PMID:14762060
Mycoplasma (M.) bovis was identified and reported in Austria as agent of infection and disease in cattle only once, namely in 2005 associated with a case of mastitis in a smallholding, but in 2007 it unexpectedly emerged as the cause of a devastating disease outbreak in a dairy herd of 100 individuals and spill over infection to pigs, both kept on the same mountain pasture. In 2008, M. bovis remained endemic at a low level in this region followed by the re-emergence of the agent in 2009 and a dramatic spread of the disease to further Alpine areas and their foothills in 2010 and 2011. From these outbreaks, a total of 94 M. bovis isolates including 7 porcine isolates were selected for genotyping. Two molecular tools, randomly amplified polymorphic DNA (RAPD) analysis and multi-locus variable number of tandem-repeat analysis (MLVA) were chosen to identify strain types involved in these outbreaks and to trace routes of infection and dynamics of dissemination. With both typing methods, the majority of Alpine isolates (96.8%) recovered over time from different areas and hosts was clustered into one group exhibiting a unique and indistinguishable profile which significantly differed from those of geographically unrelated strains including the type strain PG45 and 3 Alpine isolates which suddenly appeared and disappeared in 2009. Stability of the unique profile strongly indicated that a single M. bovis strain initiated the outbreak in 2007, crossed the host species barrier by infecting pigs, re-emerged in 2009 and became widespread in the Austrian Alps in 2010 and 2011. The remarkable dissemination and persistence of a single and unique M. bovis strain may reflect peculiarities of dairy management practices in the Alps based on Alpine transhumance and cooperative use of mountain pastures. As the source of the outbreak strain remains unknown, the findings of this study underscore the importance of continuous surveillance by monitoring further spread and persistence of M. bovis infections for effective control to minimize losses in Alpine dairy farming. PMID:23490560
Neuraminidases are virulence factors in many pathogenic microorganisms. They are present also in some Mycoplasma species that cause disease in birds, dogs and alligators. Thirty-seven Mycoplasma species have been examined previously for neuraminidase (sialidase) activity, whereas many of the species causing disease in man, ruminants, pigs, rodents and other animals have not. In this study neuraminidase enzymatic activity (NEAC) was examined in 45 previously untested Mycoplasma species, including those causing diseases in man, farm animals and laboratory animals. The only species in which NEAC was found was Mycoplasma neurolyticum, specifically, its type strain (Type A(T)) which is capable of inducing neurologic signs in inoculated young mice and rats. The NEAC of washed cells was relatively weak, but it differed even more than 10-fold among cells of cultures derived from individual colonies of M. neurolyticum. A weak NEAC was also detected in the supernatant of the M. neurolyticum broth culture. Canine Mycoplasma spp. with high sialidase activity reported previously, Mycoplasma canis, Mycoplasma cynos and Mycoplasma molare had 100-fold more NEAC than M. neurolyticum, but apparent differences in NEAC levels existed among strains of M. canis and of M. cynos. Zymograms using neuraminidase-specific chromogenic substrate were used to show proteins having NEAC. In M. canis (a field isolate Larissa and the type strain PG14(T)), M. cynos (isolate 896) and M. molare (type strain H542(T)) proteins with NEAC had molecular masses of ?130kDa, 105kDa and 110kDa, respectively. Identification of these neuraminidases could provide the basis for their molecular characterization. PMID:21937171
Ber?i?, Rebeka Lucijana; Cizelj, Ivanka; Ben?ina, Mateja; Narat, Mojca; Bradbury, Janet M; Dov?, Peter; Ben?ina, Dušan
Experimental inoculation with the F-strain of Mycoplasma gallisepticum (FMG) between 8 and 18 wk of age is known to affect reproductive performance in commercial layers. Therefore, two trials were conducted to determine if changes in digestive and reproductive organ characteristics also occur in commercial laying hens infected with FMG at 12 wk of age. In Trial 1, liver weight, liver lipid and moisture contents, ovary weight, ovarian follicular hierarchy, and the weights, lengths, and histologies of the infundibulum, magnum, isthmus, uterus, and vagina were determined. In Trial 2, fatty liver hemorrhagic syndrome (FLHS) incidence and the weights, lengths, and histologies of the duodenum, jejunum, and ileum were determined in addition to the parameters examined in Trial 1. In both trials, the average number of mature (diameter > or = 12 mm) ovarian follicles was lower in FMG-inoculated hens in comparison to controls. Also, magnum/oviduct (cm/cm) length was reduced in treated birds. In Trial 2, isthmus/BW and isthmus/oviduct (g/ g) weight were decreased at 46 wk of age, and vagina/ BW and vagina/oviduct (g/g) weight were decreased at both 20 and 36 wk of age due to FMG treatment. In Trial 2, FMG treatment resulted in a 50% increase in the number of FLHS birds. Furthermore, treatment caused a decrease at 20 wk of age and an increase at 44 wk of age in liver moisture content. However, the intestinal characteristics examined were not affected by FMG inoculation. Altered liver, ovarian, and reproductive organ characteristics were associated with FMG infection in commercial layers. More specifically, FMG inoculation at 12 wk resulted in a higher incidence of FLHS, ovarian follicular regression, and decreased isthmal and vaginal proportions of the reproductive tract. These data clearly demonstrate that alterations in performance and egg characteristics of layers inoculated with FMG at 12 wk of age are related to mutual functional disturbances in the liver, ovary, and oviduct without concomitant intestinal changes. PMID:12512582
Burnham, M R; Peebles, E D; Branton, S L; Jones, M S; Gerard, P D; Maslin, W R
Samples of cervico-vaginal mucus from 633 animals from 110 herds were cultured and yielded the following mycoplasmas: T-strain--88: Mycoplasma bovigenitalium--79, Mycoplasma spp. (Leach Group 7)--7, Acholeplasma laidlawii--4, Mycoplasma bovirhinis--2 and one not typable. Uterine exudates and endometrial scrapings from 80 infertile cows in two herds were examined. Four animals were positive, M. bovigenitalium was isolated three times, A. laidlawii and Mycoplasma arginini once each. Sixty-five normal uterine contents from pregnant cows were examined, one yielded M. bovigenigalium and the same organism was recovered from the fetal kidney. T-strainmycoplasma, M. bovigenitalium and other Mycoplasma spp. appear to be a part of the normal flora of the cervico-vaginal region of clinically normal one and two year old bred heifers in Alberta and Saskatchewan. Although M. arginini was not recovered from the cervico-vaginal region, a single recovery was made from the uterus of an infertile cow.
The history of B. hominis is unique. Few infectious agents have provoked the many misconceptions that plague this enigmatic parasitic ameba. Conflicting descriptions of its nature and pathogenesis have continued throughout the 20th century. As seen by the greatly expanded number of reports in recent years, B. hominis is now a major subject of study, particularly for evidence of disease causation. Physicians are treating patients with intestinal disease caused by B. hominis. Many mild cases resolve in about 3 days without treatment, but others are acute and chronic disease is common. As with E. histolytica, the carrier state is often seen without symptoms. Treatment is usually with metronidazole, but emetine (for refractory infections), trimethoprim-sulfamethoxazole, and pentamidine are also effective. In fecal samples, this complex protozoan appears in a variety of cell forms which makes microscopic diagnosis difficult. As yet, no specific fluorescent-antibody test is available for diagnosis. A culture method to demonstrate the more easily recognized CB form is available, but probably not feasible for most diagnostic laboratories. The common cell forms are the CB form, the granular (mitochondria) form, and the ameba form. The unexpected size range of these forms in clinical material, from yeast size (ca. 7 microns) to giant cells of 20 to 40 microns, makes diagnosis difficult Pseudopodia may be demonstrated by the ameba form in heated microscope stage culture chambers. The anaerobic B. hominis has no cyst form. Its mitochondria are uniquely anaerobic and have no cytochrome protein or oxidative mitochondrial enzymes. Because of its many cell forms and anaerobic mitochondria, B. hominis is an organism of great interest for morphologic and biochemical study. Reproduction is asexual, usually by binary fission. Shizogony occurs in cultured cells. The CB appears to be an organelle whose specific purpose is for reproduction by shizogony. From 2 to 30 progeny are derived from schizogony. The ameba form reproduces by plasmotomy; it has no CB. The pathology of B. hominis infections has been studied in gnotobiotic guinea pigs in which inflammation of the intestinal mucosa and invasion of the superficial layers were seen. Only limited studies of human pathology are available. Those who have studied mucosal histopathology report inflammation and cellular changes that resolve after treatment. More study in this area is strongly indicated (32, 44, 57, 62, 67, 75). Ultrastructural details of B. hominis major forms, except for the schizont, are complete. The organism has no cell wall. The concentric CB takes up as much as 95% of the cell. The major organelles, which include multiple nuclei, Golgi apparatus, mitochondria, endoplasmic reticulum, fat, and other inclusions, are confined in two or four opposed pods in a thin band of peripheral cytoplasm between the spherical entire plasma membrane and the CB membrane. The pods buldge the CB membrane inward. There is evidence of a bacteroid endosymbiont. Education about B. hominis is needed. Entry of recent findings into new textbooks is imperative for its understanding among medical practitioners. Laboratory workers need to be aware of it for many reasons. The College of American Pathologists includes B. hominis in its proficiency testing samples and requires that it be reported from clinical samples. Images
Two trials were conducted to determine the effects of a prelay ts-11-strainMycoplasma gallisepticum (ts-11MG) vaccination alone or in conjunction with F-strain M. gallisepticum (FMG) inoculation overlays at 2 different age periods during lay on the digestive and reproductive organ characteristics of commercial egg-laying hens. In each trial, the following 4 treatments were utilized: sham vaccination at 10 wk of age, ts-11MG vaccination at 10 wk of age, ts-11MG at 10 wk of age overlaid by FMG inoculation at 22 wk of age, and ts-11MG at 10 wk of age overlaid by FMG at 45 wk of age. Necropsies were performed at the end of both trials (58 wk of age), using 2 birds from each of 4 replicate units per treatment, to observe treatment effects on the following parameters: liver weight, liver lipid and moisture concentrations, incidence of fatty liver hemorrhagic syndrome, ovary weight, number of mature ovarian follicles, and the total and segmental weights, lengths, and histologies of the oviduct and small intestine. Treatments affected only vaginal length as a percentage of total oviduct length. Vaginas were relatively longer in hens that had only been vaccinated with ts-11MG at 10 wk in comparison to all the other treatment groups, including controls. Except for relative vaginal length, the digestive and reproductive organs of layers were not influenced by the ts-11MG and FMG treatment regimens imposed in this study. These results confirm that when coupled with FMG inoculations during lay, prelay ts-11MG vaccinations may be a practical substitute for prelay FMG inoculations for providing continual protection against field-strain M. gallisepticum infections in layers. PMID:19359686
Vance, A M; Branton, S L; Collier, S D; Gerard, P D; Peebles, E D
Quantitative methods of estimation of similarity between gene orders have been used to compare the genomes of 14 strains of\\u000a mycoplasmas and 2 strains of phytoplasmas, i.e., all genomes of bacteria of the class Mollicutes sequenced to date. Reconstructions\\u000a of the mycoplasma phylogeny based on comparisons of (a) gene orders in a chromosome and (b) nucleotide or amino acid sequences
Two trials were conducted to determine the effects of a prelay 6/85-strainMycoplasma gallisepticum (6/85MG) vaccination alone or in conjunction with time-specific F-strain M. gallisepticum (FMG) inoculation overlays on the gross reproductive and digestive organ characteristics of commercial egg-laying hens. In each trial, the following 4 treatments were applied: 1) sham vaccination at 10 wk of age; 2) vaccination of 6/85MG at 10 wk; 3) 6/85MG at 10 wk overlaid by FMG inoculation at 22 wk; and 4) 6/85MG at 10 wk overlaid by FMG at 45 wk. Two birds per isolation pen (experimental replicate unit) were necropsied at the end of both trials to observe the effects of treatment on liver weight, liver lipid and moisture concentrations, incidence of fatty liver hemorrhagic syndrome, ovary weight, mature ovarian follicle numbers, and the total and segmental weights, lengths, and histologies of the oviduct and small intestine. The applied treatments affected only liver moisture. Liver moisture content was greater in birds vaccinated with 6/85MG at 10 wk alone or in conjunction with FMG at 45 wk in comparison with sham vaccinated controls and birds that received a 6/85MG vaccination at 10 wk overlaid by an FMG inoculation at 22 wk. Prelay 6/85MG vaccinations may be a suitable substitute for prelay FMG inoculations, and FMG overlays during lay on prelay 6/85MG vaccinations may also provide continual protection against field-strain MG infections without eliciting any subsequent suppressive effects on performance, as noted in an earlier study. PMID:19211526
Viscione, K A; Branton, S L; Vance, A M; Gerard, P D; Womack, S K; Peebles, E D
Background The Mycoplasma mycoides cluster consists of five species or subspecies that are ruminant pathogens. One subspecies, Mycoplasma mycoides subspecies mycoides Small Colony (MmmSC), is the causative agent of contagious bovine pleuropneumonia. Its very close relative, Mycoplasma mycoides subsp. capri (Mmc), is a more ubiquitous pathogen in small ruminants causing mastitis, arthritis, keratitis, pneumonia and septicaemia\\u000a and is also found as
Francois Thiaucourt; Lucia Manso-Silvan; Woubit Salah; Valérie Barbe; Benoit Vacherie; Daniel Jacob; Marc Breton; Virginie Dupuy; Anne Marie Lomenech; Alain Blanchard; Pascal Sirand-Pugnet
After the first outbreak of fatal Mycoplasma ovis infection (eperythrozoonosis) in a sheep flock in Hungary (1997), a second wave of the disease was noted in 2006, with different seasonal pattern and affected age group, as well as increased mortality (5.5%). The aim of the present study was to molecularly characterize the causative agent and to reveal underlying factors of
Sándor Hornok; Marina L. Meli; András Erd?s; István Hajtós; Hans Lutz; Regina Hofmann-Lehmann
The effects of a prelay 6/85-strainMycoplasma gallisepticum (6/85 MG) inoculation alone or in conjunction with subsequent F-strain M. gallisepticum (FMG) inoculations during lay on the internal egg characteristics of commercial egg-laying hens were investigated. In the first 2 treatment groups, birds were sham inoculated or were inoculated with 6/85 MG at 10 wk of age. In a third treatment group, birds were inoculated with 6/85 MG at 10 wk in conjunction with a subsequent inoculation of FMG at 22 wk, and in a fourth treatment group, birds were inoculated with 6/85 MG at 10 wk in conjunction with a subsequent inoculation of FMG at 45 wk. Percentage yolk weight, albumen weight, yolk moisture, and yolk lipid were determined at 24, 32, 43, 47, and 58 wk of hen age. The data from wk 24, 32, and 43 were analyzed separately from those at wk 47 and 58. Furthermore, yolk fatty acid profiles were determined at wk 58. The applied treatments affected yolk moisture and fatty acid profiles. Across wk 24, 32, and 43, yolk moisture content was higher in birds inoculated with 6/85 MG at 10 wk and FMG at 22 wk when compared with control birds and those inoculated with 6/85 MG at 10 wk alone. In addition, at wk 58, yolk palmitic, oleic, and linolenic acid concentrations were affected differently by treatment. A 22-wk FMG inoculation in birds previously inoculated with 6/85 MG at 10 wk may increase yolk moisture content, and alterations in yolk palmitic, oleic, and linolenic acid levels with treatment may be manifested by disturbances in fatty acid synthesis. PMID:18493000
Viscione, K A; Branton, S L; Gerard, P D; Whitmarsh, S K; Peebles, E D
The influences of F-strainMycoplasma gallisepticum (FMG) vaccine inoculation during the pullet period on the subsequent productive and reproductive performance of parent broiler chicken breeders on a multi-age farm were evaluated. Three thousand breeders were randomly divided into 2 treatment groups that were either vaccinated with FMG (FMG-vaccinated group) or not vaccinated with FMG (FMG-free group). Body weight and egg production were determined through approximately 50 wk of age. Egg weight and feed conversion was determined at 26, 32, 35, 38, and 43 wk of age. Egg quality parameters, including eggshell strength, egg-specific gravity, egg shape index, blood-meat spots, Haugh unit score, eggshell thickness, yolk:albumen ratio, percentage yolk, albumen and eggshell weights, and percentage fertility, hatchability, and second-quality chicks were determined at 26, 32, and 43 wk of age. Air sacs were examined and lesions were scored at 20, 32, and 50 wk of age. The number of mature ovarian follicles, histologies of ovary, and lengths, and histologies of the infundibulum, magnum, isthmus, uterus, and vagina were determined. In the present study, an increase in egg production of broiler breeder hens in the FMG-vaccinated group during peak of lay was compared with the FMG-free group. Feed conversion of hens in the FMG-vaccinated group was significantly less at 32, 35, 38, and 43 wk of age. Eggs from hens in the FMG-vaccinated group had a significantly higher Haugh units score at 26 wk of age and had a significantly higher eggshell thickness and lower incidence of blood-meat spots at 32 wk. Hatching eggs from hens in the FMG-vaccinated group had a significantly higher hatchability. The mean lesion score of air-sac lesion of birds in the FMG-vaccinated group was significantly less than FMG-vaccinated group. Uteruses of hens in the FMG-vaccinated group had a significantly longer length compared with the FMG-free group at 32 wk of age. The results indicate that inoculation of commercial parent broiler chicken breeders with the FMG vaccine before laying may prevent infection by field M. gallisepticum, and facilitate productive and reproductive performance. PMID:23687149
Background Mycoplasma bovis is associated with pneumonia in calves characterized by the development of chronic caseonecrotic lesions with the agent persisting within the lesion. The purposes of this study were to characterize the morphology of lung lesions, examine the presence of M. bovis variable surface protein (Vsp) antigens and study the local immune responses in calves after infection with M. bovis strain 1067. Methods Lung tissue samples from eight calves euthanased three weeks after experimental infection with M. bovis were examined by bacteriology and pathology. Lung lesions were evaluated by immunohistochemical (IHC) staining for wide spectrum cytokeratin and for M. bovis Vsp antigens and pMB67 antigen. IHC identification and quantitative evaluation of CD4+ and CD8+ T lymphocytes and immunoglobulin (IgG1, IgG2, IgM, IgA)-containing plasma cells was performed. Additionally, expression of major histocompatibility complex class II (MHC class II) was studied by IHC. Results Suppurative pneumonic lesions were found in all calves. In two calves with caseonecrotic pneumonia, necrotic foci were surrounded by epithelial cells resembling bronchial or bronchiolar epithelium. In all calves, M. bovis Vsp antigens were constantly present in the cytoplasm of macrophages and were also present extracellularly at the periphery of necrotic foci. There was a considerable increase in numbers of IgG1- and IgG2-positive plasma cells among which IgG1-containing plasma cells clearly predominated. Statistical evaluation of the numbers of CD4+ and CD8+ T cells, however, did not reveal statistically significant differences between inoculated and control calves. In M. bovis infected calves, hyperplasia of bronchus-associated lymphoid tissue (BALT) was characterized by strong MHC class II expression of lymphoid cells, but only few of the macrophages demarcating the caseonecrotic foci were positive for MHC class II. Conclusions The results from this study show that infection of calves with M. bovis results in various lung lesions including caseonecrotic pneumonia originating from bronchioli and bronchi. There is long-term persistence of M. bovis as demonstrated by bacteriology and immunohistochemistry for M. bovis antigens, i.e. Vsp antigens and pMB67. The persistence of the pathogen and its ability to evade the specific immune response may in part result from local downregulation of antigen presenting mechanisms and an ineffective humoral immune response with prevalence of IgG1 antibodies that, compared to IgG2 antibodies, are poor opsonins.
Contagious agalactia is a mycoplasmal infection caused by Mycoplasma agalactiae, Mycoplasma mycoides subsp. mycoides LC, M. mycoides subsp. capri, Mycoplasma capricolum subsp. capricolum and Mycoplasma putrefaciens. Identification of the causative organisms is usually performed by isolation and classical biochemical and serological tests, though this is a lengthy and cumbersome process for mycoplasmas. Specific PCR assays have been developed for the identification of Mycoplasma agalactiae and M. putrefaciens. For members of the M. mycoides cluster existing PCR tests are based on the amplification of highly conserved genes coding for ribosomal proteins, hence a possibility of cross-reactions. The gene glk, coding for a glucokinase, that is found in this cluster is very distantly related to any other bacterial glucokinase described so far. It was therefore chosen as target to design a new PCR test. The validation was performed independently in three laboratories in France and India using over 100 mycoplasmastrains of various geographical origins. All strains belonging to the M. mycoides cluster were detected by amplification of the expected PCR product (428 bp) while no amplification was obtained from M. agalactiae strains. Our results demonstrate the universality of this PCR in spite of the great heterogeneity found within this cluster. This new tool may be of great help for the implementation of control measures directed towards contagious agalactia. PMID:17606362
Woubit, S; Manso-Silván, L; Lorenzon, S; Gaurivaud, P; Poumarat, F; Pellet, M-P; Singh, V P; Thiaucourt, F
SUMMARY Eleven strains of the avian pathogen Mycoplasma synoviae were evaluated for the presence of sialidase activity by using the fluorogenic substrate 2?-(4-methylumbelliferyl)-?-D-N-acetylneuraminic acid and the sialidase inhibitor 2-deoxy-2,3-didehydro-N-acetylneuraminic acid. The kinetics of in vitro growth in modified Frey’s medium were also assessed for each strain. Five strains had been isolated from clinically symptomatic chickens, and strains WVU1853T and K3344 have been demonstrated to be capable of reproducing disease in specific pathogen-free chickens. All strains exhibited sialidase activity, although the amount varied 65 fold (P < 0.0001) from 1.3 x 10?7 to 2.0 x 10?9 activity units/colony-forming unit among strains. Strains originally isolated from clinically symptomatic birds had more (P < 0.05) sialidase activity than strains from asymptomatic birds. Strain WVU1853T exhibited the most sialidase activity (P < 0.0001) and grew to the highest culture density (P < 0.0001) among strains, but across strains the rank correlation of growth rate with sialidase activity was not significant. Negligible activity was detected in conditioned culture supernatant fluid. This is the first report of sialidase activity in pathogenic strains of M. synoviae, which suggests a potential enzymatic basis for virulence of the organism.
The effects of 6/85-strainMycoplasma gallisepticum (6/85MG) inoculation alone or in conjunction with a F-strain M. gallisepticum (FMG) overlay and its timing on the performance of commercial egg-laying hens were investigated. Control birds received sham inoculations at 10 wk of age. A second treated group of birds was inoculated with 6/85MG at 10 wk of age, a third treatment group of birds was inoculated with 6/85MG at 10 wk and received an overlay of FMG at 22 wk, and a fourth treatment group was inoculated with 6/85MG at 10 wk followed by a 45-wk overlay inoculation of FMG. Parameters investigated between 20 and 55 wk of age included BW, mortality, weekly and total egg production, egg weight, relative eggshell conductance, eggshell weight per unit of surface area, percentage shell weight, yolk/albumen ratio, and egg shape index. Hen age effects were reported for BW, egg weight, yolk/albumen ratio, and egg production. No treatment effects or hen age x treatment interactions were noted for those parameters except for yolk/albumen ratio, and no significant effects of any kind were noted for the remaining parameters examined. At wk 47, a significant treatment effect for yolk/albumen ratio was noted. The yolk/albumen ratio in the group of birds that received 6/85MG at 10 wk followed by an overlay of FMG at 22 wk was significantly lower than the sham control birds and those that were inoculated with 6/85MG at 10 wk followed by a 45-wk overlay inoculation of FMG. Prelay 6/85MG inoculations may be a suitable substitute for prelay FMG inoculations, and FMG overlays during lay on prelay 6/85MG inoculations may provide continual protection without eliciting any subsequent suppressive effects on performance. PMID:18281589
Viscione, K A; Branton, S L; Vance, A M; Gerard, P D; Whitmarsh, S K; Peebles, E D
An integrative conjugative element, ICEA, was characterized in Mycoplasma agalactiae strain 5632, in which it occurs as multiple chromosomal copies and as a free circular form. The distribution of ICEA sequences in M. agalactiae strains and their occurrence in Mycoplasma bovis suggest the spreading of the element within or between species.
Automated solid-phase DNA sequencing was used for determination of partial 16S ribosomal DNA sequences of mycoplasmas. The sequence information was used to establish phylogenetic relationships of 11 different mycoplasmas whose 16S rRNA sequences had not been determined earlier. A biotinylated fragment corresponding to positions 344 to 939 in the Escherichia coli sequence was generated by PCR. The PCR product was immobilized onto streptavidin-coated paramagnetic beads, and direct sequencing was performed in both directions. One previously unclassified avian mycoplasma was found to belong to the Mycoplasma lipophilum cluster of the hominis group. Microheterogeneities were discovered in the rRNA operons of Mycoplasma mycoides subsp. mycoides (SC type), confirming the existence of two different rRNA operons. The 16S rRNA sequence of M. mycoides subsp. capri was identical to that of M. mycoides subsp. mycoides (type SC), except that no microheterogeneities were revealed. Furthermore, automated solid-phase DNA sequencing was used to identify a mycoplasmal contamination of a cell culture as Mycoplasma hyorhinis, which proved to be very difficult by conventional methods. The results suggest that the direct solid-phase DNA sequencing procedure is a powerful tool for identification of mycoplasmas and is also useful in taxonomic studies. Images
Based on biological and molecular data, the species of Cryptosporidium infecting the intestine of humans is considered a new species for which the name Cryptosporidium hominis is proposed. The structure and infectivity of the oocysts of C. hominis from the feces of humans are described. Oocysts are ...
Within the genus Mycoplasma are species whose cells have terminal organelles, polarized structures associated with cytadherence and gliding motility. Mycoplasma penetrans, found mostly in HIV-infected patients, and Mycoplasma iowae, an economically significant poultry pathogen, are members of the Mycoplasma muris phylogenetic cluster. Both species have terminal organelles that interact with host cells, yet the structures in these species, or any in the M. muris cluster, remain uncharacterized. Time-lapse microcinematography of two strains of M. penetrans, GTU-54-6A1 and HF-2, and two serovars of M. iowae, K and N, show that the terminal organelles of both species play a role in gliding motility, with differences in speed within and between the two species. The strains and serovars also differed in their hemadsorption abilities that positively correlated with differences in motility speeds. No morphological differences were observed between M. penetrans and M. iowae by scanning electron microscopy (SEM). SEM and light microscopy of M. penetrans and M. iowae showed the presence of membranous filaments connecting pairs of dividing cells. Breaking of this filament during cell division was observed for M. penetrans by microcinematography, and this suggests a role for motility during division. The Triton X-100-insoluble fractions of M. penetrans and M. iowae consisted of similar structures that were unique compared to those identified in other mycoplasma species. Like other polarized mycoplasmas, M. penetrans and M. iowae have terminal organelles with cytadherence and gliding functions. The difference in function and morphology of the terminal organelles suggests that mycoplasmas have evolved terminal organelles independently of one another.
An immunodominant protein, P40, of Mycoplasma agalactiae was analyzed genetically and functionally. The gene encoding P40 was cloned from type strain PG2, sequenced, submitted to point mutagenesis in order to convert mycoplasma-specific TGATrp codon to the universal TGGTrp codon, and subsequently expressed in Escherichia coli. Nucleotide sequence-derived amino acid sequence comparisons revealed a similarity of P40 to the adhesin P50
Benedicte Fleury; Dominique Bergonier; Xavier Berthelot; Ernst Peterhans; Joachim Frey; Edy M. Vilei
The vsa genes of Mycoplasma pulmonis encode the V-1 lipoproteins. Most V-1 proteins contain repetitive domains and are thought to be involved in mycoplasma-host cell interactions. Previously, we have reported the isolation and characterization of six vsa genes comprising a 10-kb region of the genome of M. pulmonis strain KD735-15. In the current study, vsa-specific probes were used to clone
XUEJUN SHEN; JULIANN GUMULAK; HUILAN YU; C. TODD FRENCH; NIANXIANG ZOU; KEVIN DYBVIG
Somerson, Norman L. (National Institute of Allergy and Infectious Diseases, Bethesda, Md.), and M. K. Cook. Suppression of Rous sarcoma virus growth in tissue cultures by Mycoplasma orale. J. Bacteriol. 90:534–540. 1965.—An agent which produced cell destruction in human diploid and chick-embryo fibroblasts was isolated from WI-26 strain of human diploid fibroblasts and shown to be a mycoplasma. The multiplication of Rous sarcoma virus (RSV) and Rous associated virus (RAV) was inhibited in WI-26, WI-38, and chick-embryo fibroblasts infected with this mycoplasma. The mycoplasma isolate, designated strain 941, reacted strongly in the complement-fixation test with antiserum to Mycoplasma orale CH19299, an isolate obtained from the human oral cavity. The cytopathic effect of mycoplasmastrain 941 could be eliminated by growing the mycoplasma on an artificial agar medium before inoculation into chick-embryo fibroblasts. Serial passage in chick-embryo fibroblasts restored the cytopathogenicity of the agar-grown mycoplasma. However, growth of RSV and RAV was inhibited by both the tissue culture-grown and the agar-grown 941 strain, and also by the CH19299 strain which did not produce any cytopathic effect. Images
The effects of dietary supplementation with phytase and 25-hydroxycholecalciferol on the performance characteristics of commercial layers that were inoculated prelay (12 wk of age) or at the onset of lay (22 wk of age) with F-strainMycoplasma gallisepticum were assessed. Experimental layer diets, which included a basal control diet or the same diet supplemented with 0.025% phytase and 25-hydroxycholecalciferol, were fed from 20 through 58 wk of age. Weekly and total egg production were determined from 22 through 58 wk, and egg weight and various internal egg and eggshell quality characteristics were examined at 34, 50, and 58 wk of age. F-strain M. gallisepticum inoculation decreased egg production at the beginning of lay (wk 22 and 23) but increased post-peak lay at wk 45. However, there were no treatment effects of any kind on total egg production, egg weight, or any of the internal egg and eggshell characteristics examined during lay. In conclusion, dietary supplementation with phytase and 25-hydroxycholecalciferol did not affect layer performance or interact with the effects of F-strain M. gallisepticum inoculation; however, F-strain M. gallisepticum inoculation resulted in a shift in egg production from wk 22 to 45 without having an overall effect on total egg production. PMID:18281591
Peebles, E D; Branton, S L; Burnham, M R; Whitmarsh, S K; Gerard, P D
The polymerase chain reaction (PCR) with primers complementary to the 16S rRNA genes was used to detect avian mycoplasmas. A primer pair designed for the detection of human and rodent mycoplasmal species was examined for its ability to detect the most important avian mycoplasmas. After testing the respective reference strains, we found that Mycoplasma iowae, Mycoplasma meleagridis and Mycoplasma synoviae could be detected by PCR with this primer pair, and distinction could be made among them by restriction fragment length polymorphism (RFLP) assay with two restriction enzymes (BamHI and RsaI). For the detection of Mycoplasma gallisepticum by PCR, we needed species-specific primers. The results of the PCR- and RFLP, based identification procedures of 17 different field isolates agreed with those obtained by conventional methods. PMID:9451458
Kiss, I; Matiz, K; Kaszanyitzky, E; Chávez, Y; Johansson, K E
The polymerase chain reaction (PCR) was evaluated to detect mycoplasma contamination of avian live virus vaccines. The specificity of the primers showed that 34 strains belonging to nine species of avian mycoplasma DNA could be detected. The sensitivity of PCR to detect mycoplasma DNA was 10(0.2) colony forming units (cfu) of Mycoplasma synoviae and 10(0.7) cfu of Mycoplasma gallisepticum. When M. synoviae and M. gallisepticum were spiked into several avian live virus vaccines, PCR gave a positive reaction except for the avian pox and the avian encephalomyelitis vaccines which were prepared from organ homogenates. Short-term incubation of avian encephalomyelitis vaccines improved the sensitivity of PCR to detect both M. synoviae and M. gallisepticum. Therefore, PCR, combined with the short-term incubation, were shown to be most effective in detecting mycoplasma contamination in all of avian live virus vaccines. PMID:9467032
Kojima, A; Takahashi, T; Kijima, M; Ogikubo, Y; Nishimura, M; Nishimura, S; Harasawa, R; Tamura, Y
Recently we have shown that a low (Rlow) and a high laboratory passage (Rhigh) of the poultry pathogen Mycoplasma gallisepticum prototype strain R differ markedly in their capability to invade non-phagocytic eukaryotic cells. In the present study the infection traits of these two mycoplasma passages were compared in an in vivo setting. After aerosol inoculation of chickens, M. gallisepticum was
BACKGROUND: Phytoplasmas are insect-transmitted, uncultivable bacterial plant pathogens that cause diseases in hundreds of economically important plants. They represent a monophyletic group within the class Mollicutes (trivial name mycoplasmas) and are characterized by a small genome with a low GC content, and the lack of a firm cell wall. All mycoplasmas, including strains of 'Candidatus (Ca.) Phytoplasma asteris' and 'Ca.
Michael Kube; Bernd Schneider; Heiner Kuhl; Thomas Dandekar; Katja Heitmann; Alexander M Migdoll; Richard Reinhardt; Erich Seemüller
Many lipoproteins are expressed on the surfaces of mycoplasmas, and some have been implicated as playing roles in pathogenesis. Family 2 lipoproteins of Mycoplasma pneumoniae have a conserved “mycoplasma lipoprotein X” central domain and a “mycoplasma lipoprotein 10” C-terminal domain and are differentially expressed in response to environmental conditions. Homologues of family 2 lipoproteins are Mycoplasma specific and include the lipoprotein of Mycoplasma gallisepticum, encoded by the MGA0674 gene. Comparative transcriptomic analysis of the M. gallisepticum live attenuated vaccine strain F and the virulent strain Rlow, reported in this study, indicated that MGA0674 is one of several differentially expressed genes. The MGA0674-encoded lipoprotein is a proteolytically processed, immunogenic, TX-114 detergent-phase protein which appears to have antigenic divergence between field strains Rlow and S6. We examined the virulence of an Rlow ?MGA0674 mutant (P1H9) in vivo and observed reduced recovery and attenuated virulence in the tracheas of experimentally infected chickens. The virulence of two additional Rlow ?MGA0674 mutants, 2162 and 2204, was assessed in a second in vivo virulence experiment. These mutants exhibited partial to complete attenuation in vivo, but recovery was observed more frequently. Since only Mycoplasma species harbor homologues of MGA0674, the gene product has been renamed “Mycoplasma-specific lipoprotein A” (MslA). Collectively, these data indicate that MslA is an immunogenic lipoprotein exhibiting reduced expression in an attenuated strain and plays a role in M. gallisepticum virulence.
Szczepanek, S. M.; Frasca, S.; Schumacher, V. L.; Liao, X.; Padula, M.; Djordjevic, S. P.; Geary, S. J.
... is not required to be tested for mycoplasma ... and points to consider require mycoplasma testing for viral ... we agreed it is prudent to test the working ... More results from www.fda.gov/biologicsbloodvaccines/vaccines/approvedproducts
Localization of the lipid of Blastocystis hominis was histochemically demonstrated. B. hominis was fixed with glutaraldehyde, treated with detergents, and stained with Sudan black B or Nile blue. The central vacuole of B. hominis stained with Sudan black B showed many black droplets with great variation in staining intensity. In Nile blue staining, the central vacuole showed many pinkish granules or homogeneous blue reactions. These results indicated the presence of neutral lipids and/or acidic lipids in the central vacuole. At the ultrastructural level, the central vacuole and some cytoplasmic vesicles showed enhanced electron density after post-fixation with imidazole-buffered osmium tetroxide solution. Great variation in the density and distribution of the electron-dense materials was observed in the central vacuole. Since the electron-dense vesicles were frequently observed in the cytoplasm, B. hominis may accumulate the lipid in the central vacuole, the function of which is not well known. PMID:7544390
Yoshikawa H. & Hayakawa A. 1996. Freeze-fracture cytochemistry of membrane cholesterol in Blastocystis hominis. International Journal for Parasitology 26: 1111–1114. Membrane cholesterol in Blastocystis hominis was detected by freeze-fracture methods using a polyene antibiotic, filipin. Since the intramembrane particles (IMP) were distributed heterogeneously on both plasma and vacuole membranes, many IMF-free areas were observed. Even in filipin-treated cells, filipin-cholesterol complexes
The in vitro activities of valnemulin (Econor®) and two other antimicrobial agents were determined against recent field strains of Mycoplasma hyopneumoniae and Mycoplasma hyosynoviae using a broth microdilution method. Valnemulin showed exceptional activity against M hyopneumoniae (MIC90 0·0005 ?g ml?1) and M hyosynoviae (MIC range 0·0001 ?g ml?1 to 0·00025 ?g ml?1) field strains. Tiamulin was 100-fold less active (MIC90
The Mycoplasma mycoides cluster consists of six pathogenic mycoplasmas causing disease in ruminants, which share many genotypic and phenotypic traits. The M. mycoides cluster comprises five recognized taxa: Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC), M. mycoides subsp. mycoides Large Colony (MmmLC), M. mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and M. capricolum subsp. capripneumoniae (Mccp). The group of strains known as Mycoplasma sp. bovine group 7 of Leach (MBG7) has remained unassigned, due to conflicting data obtained by different classification methods. In the present paper, all available data, including recent phylogenetic analyses, have been reviewed, resulting in a proposal for an emended taxonomy of this cluster: (i) the MBG7 strains, although related phylogenetically to M. capricolum, hold sufficient characteristic traits to be assigned as a separate species, i.e. Mycoplasma leachii sp. nov. (type strain, PG50(T) = N29(T) = NCTC 10133(T) = DSM 21131(T)); (ii) MmmLC and Mmc, which can only be distinguished by serological methods and are related more distantly to MmmSC, should be combined into a single subspecies, i.e. Mycoplasma mycoides subsp. capri, leaving M. mycoides subsp. mycoides (MmmSC) as the exclusive designation for the agent of contagious bovine pleuropneumonia. A taxonomic description of M. leachii sp. nov. and emended descriptions of M. mycoides subsp. mycoides and M. mycoides subsp. capri are presented. As a result of these emendments, the M. mycoides cluster will hereafter be composed of five taxa comprising three subclusters, which correspond to the M. mycoides subspecies, the M. capricolum subspecies and the novel species M. leachii. PMID:19502315
Manso-Silván, L; Vilei, E M; Sachse, K; Djordjevic, S P; Thiaucourt, F; Frey, J
Mycoplasmas evolved by a drastic reduction in genome size, but their genomes contain numerous repeated sequences with important roles in their evolution. We have established a bioinformatic strategy to detect the major recombination hot-spots in the genomes of Mycoplasma pneumoniae, Mycoplasma genitalium, Ureaplasma urealyticum and Mycoplasma pulmonis. This allowed the identification of large numbers of potentially variable regions, as well as a comparison of the relative recombination potentials of different genomic regions. Different trends are perceptible among mycoplasmas, probably due to different functional and structural constraints. The largest potential for illegitimate recombination in M.pulmonis is found at the vsa locus and its comparison in two different strains reveals numerous changes since divergence. On the other hand, the main M.pneumoniae and M.genitalium adhesins rely on large distant repeats and, hence, homologous recombination for variation. However, the relation between the existence of repeats and antigenic variation is not necessarily straightforward, since repeats of P1 adhesin were found to be anti-correlated with epitopes recognized by patient antibodies. These different strategies have important consequences for the structures of genomes, since large distant repeats correlate well with the major chromosomal rearrangements. Probably to avoid such events, mycoplasmas strongly avoid inverse repeats, in comparison to co-oriented repeats.
In 3 trials, the effects of dietary supplementation with phytase (PHY) and 25-hydroxycholecalciferol on BW and the blood characteristics of commercial layers that were inoculated prelay (12 wk of age) or at the onset of lay (22 wk of age) with F-strainMycoplasma gallisepticum were assessed at 34, 50, and 58 wk of age. Experimental layer diets, which included either a basal control diet or the same diet supplemented with 0.025% PHY and 25-hydroxycholecalciferol, were fed from 20 through 58 wk of age. The supplemented diet decreased blood hematocrit values across bird age, inoculation type (sham vs. F-strain M. gallisepticum), and age of inoculation (prelay vs. onset of lay). Phytase- and 25-hydroxycholecalciferol-supplemented diets reduced bird BW in sham-inoculated control birds across bird age and age of inoculation. This effect was not observed in F-strain M. gallisepticum-inoculated birds. Furthermore, across diet (control vs. supplemented) and inoculation type, total plasma protein concentration at 34 wk of age was higher in birds that were inoculated at the onset of lay compared with those inoculated prelay. Diet, inoculation type, and inoculation age had no effect on mortality, reproductive organ histopathological lesion scores, or serum cholesterol and Ca concentrations. In conclusion, throughout lay, the supplementation of commercial layer diets with PHY may lower hematocrit, and inoculation with F-strain M. gallisepticum prelay or at the onset of lay may ameliorate the depressing effects of dietary PHY and 25-hydroxycholecalciferol supplementation on hen BW. PMID:17369552
Peebles, E D; Branton, S L; Burnham, M R; Whitmarsh, S K; Gerard, P D
Recently, it was demonstrated using in vitro assays that the avian pathogen Mycoplasma gallisepticum is able to invade nonphagocytic cells. It was also shown that this mycoplasma can survive and multiply intracellularly for at least 48 h and that this cell invasion capacity contributes to the systemic spread of M. gallisepticum from the respiratory tract to the inner organs. Using the gentamicin invasion assay and a differential immunofluorescence technique combined with confocal laser scanning microscopy, we were able to demonstrate in in vitro experiments that M. gallisepticum is also capable of invading sheep and chicken erythrocytes. The frequencies of invasion of three well-defined M. gallisepticum strains were examined over a period of 24 h, and a significant increase in invasiveness occurred after 8 h of infection. In addition, blood samples derived from chickens experimentally infected via the aerosol route with the virulent strain M. gallisepticum Rlow were analyzed. Surprisingly, M. gallisepticum Rlow was detected in the bloodstream of infected chickens by nested PCR, as well as by differential immunofluorescence and interference contrast microscopy that showed that mycoplasmas were not only on the surface but also inside chicken erythrocytes. This finding provides novel insight into the pathomechanism of M. gallisepticum and may have implications for the development of preventive strategies.
Vogl, Gunther; Plaickner, Astrid; Szathmary, Susan; Stipkovits, Laszlo; Rosengarten, Renate; Szostak, Michael P.
Homologous recombination between repeated DNA elements in the genomes of Mycoplasma species has been hypothesized to be a crucial causal factor in sequence variation of antigenic proteins at the bacterial surface. To investigate this notion, studies were initiated to identify and characterize the proteins that form part of the homologous DNA recombination machinery in Mycoplasma pneumoniae as well as Mycoplasma genitalium. Among the most likely participants of this machinery are homologs of the Holliday junction migration motor protein RuvB. In both M. pneumoniae and M. genitalium, genes have been identified that have the capacity to encode RuvB homologs (MPN536 and MG359, respectively). Here, the characteristics of the MPN536- and MG359-encoded proteins (the RuvB proteins from M. pneumoniae strain FH [RuvB(FH)] and M. genitalium [RuvB(Mge)], respectively) are described. Both RuvB(FH) and RuvB(Mge) were found to have ATPase activity and to bind DNA. In addition, both proteins displayed divalent cation- and ATP-dependent DNA helicase activity on partially double-stranded DNA substrates. The helicase activity of RuvB(Mge), however, was significantly lower than that of RuvB(FH). Interestingly, we found RuvB(FH) to be expressed exclusively by subtype 2 strains of M. pneumoniae. In strains belonging to the other major subtype (subtype 1), a version of the protein is expressed (the RuvB protein from M. pneumoniae strain M129 [RuvB(M129)]) that differs from RuvB(FH) in a single amino acid residue (at position 140). In contrast to RuvB(FH), RuvB(M129) displayed only marginal levels of DNA-unwinding activity. These results demonstrate that M. pneumoniae strains (as well as closely related Mycoplasma spp.) can differ significantly in the function of components of their DNA recombination and repair machinery. PMID:21949077
Estevăo, Silvia; Sluijter, Marcel; Hartwig, Nico G; van Rossum, Annemarie M C; Vink, Cornelis
On the basis of the aligned 16S rRNA sequences of Mollicutes, a pair of primers was chosen for the detection of Mycoplasma gallisepticum. When used in the polymerase chain reaction (PCR), the primers detected a specific amplification of all Mg strains tested, yielding an expected 330 bp product. Amplification was not detected when other Mollicutes or E. coli were used as PCR templates. SPF chickens were experimentally inoculated with two strains of M. gallisepticum or Mycoplasma iowae. Tracheal swabs were collected 8, 15, 20 and 28 days after inoculation, and cultured for mycoplasma or tested by PCR. PCR products were detected by hybridization with a digoxigenin-labeled probe and by chemiluminescence. The results showed that culture was positive for 49/73 swabs while PCR detected 70/72 positive samples. Thus, PCR can provide the basis of a sensitive, specific, rapid and non-radio-active method for detecting M. gallisepticum. PMID:18671058
Kempf, I; Blanchard, A; Gesbert, F; Guittet, M; Bennejean, G
Domermuth, C. H. (Statens Seruminstitut, Copenhagen, Denmark), M. Nielsen, E. A. Freundt, and A. Birch-Andersen. Gross morphology and ultrastructure of Mycoplasma gallisepticum. J. Bacteriol. 88:1428-1432. 1964.-The ultrastructure and gross morphology of Mycoplasma gallisepticum strains JA and W were studied with an electron microscope. Intact specimens were grown on Parlodion membranes, fixed with formaldehyde, and studied in situ. Sectioned specimens were grown on agar and were prepared for study by conventional sectioning techniques. Grossly, single and multiple (as many as four) protrusions were frequently observed extending from the surface of cells of the W strain. Single short protrusions extended from many of the cells of strain JA. In sectioned material, rows of what appeared to be developing elementary bodies (as many as four) were observed in cells of strain W, whereas, single, apparently developing elementary bodies were observed in strain JA. In both strains, these bodies were located within the protruding areas of the cell wall. The inclusion-containing portions of the cell appeared to be the protrusions which extended from the surface of the intact cell. PMID:14234802
DOMERMUTH, C H; NIELSEN, M; FREUNDT, E A; BIRCH-ANDERSEN, A
Certain Mycoplasma spp. are pathogens of poultry, but little is known of the role of mycoplasmas in disease of birds of prey. Species-specific polymerase chain reactions (PCRs) for the detection of the poultry pathogens Mycoplasma gallisepticum, Mycoplasma imitans, Mycoplasma iowae, Mycoplasma meleagridis and Mycoplasma synoviae were therefore evaluated for use in birds of prey. The specificities of the PCR methods were established using avian and other mycoplasmas and also selected walled bacteria. The sensitivities of the different PCR assays varied between 100 fg and 10 pg DNA. Fifty-three tracheal swabs from healthy captive and free-ranging birds of prey were then investigated using these PCRs, and in no case was an amplicon obtained for M. gallisepticum/M. imitans, M. iowae or M. synoviae. Species-specific primers for M. meleagridis amplified a product from eight birds of prey but restriction enzyme analysis as well as sequencing of PCR products demonstrated these results to be false positives. Alignment studies of the sequenced products with the 16S rRNA gene sequence of various Mycoplasma species in GenBank demonstrated an identity of 91% to M. meleagridis but of 98% to Mycoplasma buteonis or Mycoplasma gallopavonis. Isolation and attempted identification of these mycoplasmas suggested it may be a previously unrecognized species. PMID:18661307
The in vitro activities of 5-chloro-8-hydroxyquinoline (CHQ) against single strains of 12 different species of mycoplasma and the impacts of repeated exposure of these strains to CHQ on their susceptibility to this agent have been studied. On initial exposure, the minimal inhibitory concentrations for these strains ranged from 0.24 to 1.92 micrograms of CHQ per ml of test medium; activities remained unchanged during 10 serial transfers in CHQ-containing medium.
The in vitro activities of 5-chloro-8-hydroxyquinoline (CHQ) against single strains of 12 different species of mycoplasma and the impacts of repeated exposure of these strains to CHQ on their susceptibility to this agent have been studied. On initial exposure, the minimal inhibitory concentrations for these strains ranged from 0.24 to 1.92 micrograms of CHQ per ml of test medium; activities remained unchanged during 10 serial transfers in CHQ-containing medium. PMID:263891
Pentatrichomonas hominis is considered to be a commensal protozoan of the vertebrate digestive tract. On the basis of light microscopic examination of feces, some investigators presumptively identified P. hominis as a causative agent of feline diarrhea. However, molecular identification of P. hominis infection in the cat has not been reported. Another trichomonad, Tritrichomonas foetus, is recognized as an intestinal pathogen
Jody L. Gookin; Stephen H. Stauffer; Michael G. Levy
Hominy feed is a low value ($83.7/metric ton) coproduct of the corn dry milling process that accounts for nearly 35% of the starting corn quantity. The average composition of hominy feed on a dry basis is 56.9% starch, 25.2% neutral detergent fiber, 11.1% protein, and 5.3% fat. Starch in hominy fe...
Experimental inoculation of commercial laying hens, maintained under controlled conditions, with the S6-strain of Mycoplasma gallisepticum (S6MG) at 10 wk of age has previously been shown to affect the lengths and weights of various portions of the reproductive tract without affecting subsequent performance. Two trials were conducted to compare the effects of S6MG inoculation at 10 wk of age (prior to lay), 22 wk of age (onset of lay), and 45 wk of age (during lay) on performance characteristics in commercial layers housed and maintained under controlled conditions, as in previous studies. In each trial, BW, mortality, egg production, egg weight, eggshell weight per unit of surface area, percentage eggshell weight, percentage albumen weight, percentage yolk weight, and yolk weight per albumen weight ratio were examined at various ages throughout an entire laying cycle. Across wk 47 and 58 (age periods after the last 45 wk inoculation), eggshell weight per unit of surface area and percentage eggshell weight were significantly reduced in birds that had received an S6MG inoculation at 45 wk of age when compared with birds that had not received an S6MG inoculation or had been inoculated with S6MG at either 10 or 22 wk of age. Alterations in eggshell quality in response to S6MG may become evident only in older birds that are experiencing declines in production when housed under controlled conditions. PMID:16463961
Basenko, E Y; Peebles, E D; Branton, S L; Whitmarsh, S K; Gerard, P D
Experimental inoculation of commercial laying hens with the S6-strain of Mycoplasma gallisepticum (S6MG) at 20 wk of age, while being maintained under ideal conditions, has previously been shown to affect the lengths and weights of various portions of the reproductive tract. Two trials were conducted in the current study to compare the effects of S6MG inoculation prior to lay at 10 wk of age, during onset of lay at 22 wk of age, and during lay at 45 wk of age on the digestive and reproductive organs of commercial layers similarly housed and maintained under ideal conditions. In each trial, liver weight, liver moisture and lipid concentration, incidence of fatty liver hemorrhagic syndrome, ovary weight, ovarian mature follicle numbers, weights and lengths of the oviduct and oviductal regions, and weights and lengths of the small intestine and small intestinal regions were examined at 60 wk of hen age. At 60 wk, liver lipid concentration was depressed, and isthmus weight, as a percentage of total oviduct weight, was increased in birds that had been inoculated with S6MG at 45 wk. Alterations in liver lipid content and weight of the isthmal portion of the oviduct may occur in response to S6MG inoculation during the later stages of production in layers housed under ideal conditions. PMID:16673758
Peebles, E D; Basenko, E Y; Branton, S L; Whitmarsh, S K; Gerard, P D
In 3 trials, the effects of dietary supplementation with phytase (PHY) and 25-hydroxycholecalciferol (25-D3) on the digestive and reproductive organ characteristics of commercial layers that were inoculated prelay (12 wk of age) or at the onset of lay (22 wk of age) with F-strainMycoplasma gallisepticum (FMG) were assessed at 58 wk of age. Experimental layer diets that included a basal control diet or a control diet supplemented with 0.025% PHY and 25-D3 were fed from 20 through 58 wk of age. As a percentage of total oviduct weight, magnum weight was lower in birds that were inoculated (sham or FMG) at lay onset compared with those that were inoculated prelay, and in FMG-inoculated birds, relative duodenum length was greater in those inoculated at 12 compared with 22 wk. Also, as percentages of organ weight or length, infundibulum length and isthmus weight were increased, whereas duodenum length was decreased by dietary supplementation with PHY and 25-D3. The overall timing (12 vs. 22 wk) of inoculation can affect the reproductive organ characteristics of layers, whereas, more specifically, the timing of an FMG inoculation may affect their digestive organ structure. Furthermore, independent of inoculation timing and type, the reproductive organ and digestive systems of laying hens may be influenced by dietary supplementation with PHY and 25-D3. PMID:17626828
Peebles, E D; Branton, S L; Burnham, M R; Whitmarsh, S K; Gerard, P D
The protein and antigen profiles of 60 isolates, strains and the type strain PG1 of Mycoplasma mycoides subsp. mycoides SC were compared by sodium dodecyl sulphate polyacrylamide gel electrophoresis and immunoblot analysis. Analysis using contagious bovine pleuropneumonia antisera and hyperimmune rabbit sera against several representative strains revealed some differences in protein profiles and variability in antigens among strains from different
Rosário Gonçalves; José Regalla; Jacques Nicolet; Joachim Frey; Robin Nicholas; John Bashiruddin; Paola de Santis; Aires Penha Gonçalves
A urease color test fluid medium (U-9) for the detection and identification of T (T-strain) mycoplasmas in clinical material is described which is sensitive and specific for this group of mycoplasmas. The medium was prepared from commercially available co...
On the basis of the aligned 16S rRNA sequences of Mollicutes, a pair of primers was chosen for the detection of Mycoplasma gallisepticum. When used in the polymerase chain reaction (PCR), the primers detected a specific amplification of all Mg strains tested, yielding an expected 330 bp product. Amplification was not detected when other Mollicutes or E. coli were used
Isabelle Kempf; A. Blanchard; Fabienne Gesbert; Michele Guittet; G. Bennejean
Newcastle disease is characterized by respiratory manifestations in association with nervous and/or digestive symptoms. Its prevention is done by vaccination with live attenuated (lentogenic strains) and/or killed vaccines. The lentogenic strains can lead to strong post-vaccination reaction, principally due to the presence of other pathogenic agents. Among them, Mycoplasma synoviae is worldwide important, mainly in Brazil. The dissemination of this agent in poultry flocks has been achieved due to difficulties in diagnosis and disease reproduction, virulence variations among different M.synoviae strains, and attribution of typical M.synoviae disease manifestation to other disease agents. This experimental study in SPF chicks (Gallus gallus), previously infected by M.synoviae and thereafter vaccinated against Newcastle disease, was done with the objective of evaluating M.synoviae pathogenicity through assessment of post-vaccinal respiratory reactions and serologic responses to Newcastle disease virus vaccine in the absence of environmental factors. A total of 86 three days old chicks were used, being 57 infected by eye and nostril drop, with chicken activated M. synoviae strain WVU 1853. Seven days later, 21 mycoplasma infected birds plus 29 not mycoplasma infected ones were vaccinated against Newcastle disease. As results, the not infected and vaccinated birds yielded, significantly, higher and longer lasting serologic responses to Newcastle disease vaccine virus than those infected and vaccinated. Similarly, the infected and vaccinated birds yielded lower serologic reactions to M.synoviae than those only mycoplasma infected. No post-vaccinal respiratory reaction was observed in the vaccinated birds.
de Cassia Figueira Silva, Rita; do Nascimento, Elmiro Rosendo; de Almeida Pereira, Virginia Leo; Barreto, Maria Lucia; do Nascimento, Maria da Graca Fichel
Mycoplasma gallinarum colonizes poultry as well as mammals, but is considered to have a commensal relationship with its hosts. Though unable to cause poultry disease by its self, reports have been published suggesting a synergism during mixed infections between M. gallinarum and respiratory viruses or their vaccine strains that can precipitate airsacculitis. Currently, little research is being done on M.
Mycoplasmastrain 163K was isolated from the gills of a tench (Tinca tinca L.) with red disease. The cells are elongated, ovoid or flask shaped, consisting of a thicker body and a more slender part ending in a hemispherical terminal structure, that is apparently stabilized by a cytoskeleton. They are able to attach to inert surfaces and living cells. The
H. KIRCHHOFF; P. BEYENE; M. FISCHER; J. FLOSSDORF; J. HEITMANN; B. KHATTAB; D. LOPATTA; R. ROSENGARTEN; G. SEIDEL; C. YOUSEF
Eighty-one strans of Mycoplasma arginini were isolated from swabs and tissues cultured from a series of 555 cats. The majority of these strains (84%) were recovered from the oropharyngeal regions. M. arginini was experimentally inoculated into young kittens. No clinical disease was produced. However, there was rapid colonization in the inoculated sites.
The in vitro emergence of resistance to enrofloxacin, erythromycin, tylosin, tiamulin, and oxytetracycline in three avian Mycoplasma species, Mycoplasmagallisepticum, Mycoplasma synoviae and Mycoplasma iowae was studied. Mutants were selected stepwise and their MICs were determined after 10 passages in subinhibitory concentrations of antibiotic. High-level resistance to erythromycin and tylosin developed within 2–6 passages in the three Mycoplasma species. Resistance to
A. V Gautier-Bouchardon; A. K Reinhardt; M Kobisch; I Kempf
Objective This study was performed to evaluate the relationship among the Nugent score for the diagnosis of bacterial vaginosis (BV), the results of vaginal fluid culture for genital mycoplasma, and the subsequent occurrence of preterm birth. Methods The Nugent score and culture for genital mycoplasmas were performed in vaginal fluid obtained from 977 pregnant women (gestational age 13–30 weeks). Vaginal samples were obtained with sterile cotton swabs. The relationship among the Nugent score, vaginal fluid culture results and the occurrence of spontaneous preterm birth was examined. Results (1) Of the 977 women, 14% (137) had a Nugent score of ? 8; (2) The prevalence of a positive vaginal culture for genital mycoplasmas was 30% (288); Ureaplasma urealyticum was isolated in 252 (88%), Mycoplasmahominis in 9 (3%), and both in 27 (9%) women; (3) Cases with a Nugent score of ? 8 had a higher rate of a positive vaginal culture for genital mycoplasmas than those with the lower Nugent score (55% vs. 25%; p<0.001); (4) Women with a Nugent score of ? 8 had a significantly higher rate of spontaneous preterm birth <37 (10% vs. 4%), <34 (5% vs. 2%), and <32 (4% vs. 1%) weeks of gestation than those with the lower Nugent score (At each gestational age, p<0.05); (5) In contrast, a positive vaginal culture for genital mycoplasmas was not associated with an increased risk for spontaneous preterm birth; (6) Among patients with a positive culture and a Nugent score of ? 8, the frequency of spontaneous preterm delivery (<37 weeks) was 10% (7/72); (7) There was no difference in the incidence of spontaneous preterm delivery according to the results of vaginal culture in patients with a Nugent score of ? 8, as well as in those with a lower Nugent score. Conclusion A high Nugent score (? 8) for the detection of BV but not a positive vaginal culture for genital mycoplasmas is a risk factor for spontaneous preterm birth.
SUMMARY Mycoplasmas are the smallest known free-living form of life, and differ from bacteria in a number of characteristics. They are widely distributed in the animal kingdom and may give rise to both acute and latent infections as well as being present as normal flora. The three principal rodent pathogens so far described are Mycoplasma pulmonis, Mycoplasma arthritidis and Mycoplasma
A beige-pigmented bacterium (strain CCUG 53761A(T)) was isolated from human blood from an 85-year-old man in Göteborg, Sweden. Comparative analysis of 16S rRNA gene sequences showed that this bacterium displayed <95 % similarity to all described species of the genera of the family Alcaligenaceae. It grouped within the radiation of the genus Alcaligenes, but showed only 93.0-94.8 % similarity to type strains of members of this genus (Alcaligenes faecalis subsp. parafaecalis, 94.8 %; Alcaligenes faecalis subsp. faecalis, 94.2 %; Alcaligenes faecalis subsp. phenolicus, 93.4 %). This discrimination was supported by chemotaxonomic differences. The polyamine pattern consisted of the predominant compound putrescine, moderate amounts of spermidine and minor to trace amounts of spermine and cadaverine; 2-hydroxyputrescine was not detectable. The quinone system was ubiquinone Q-8 with minor amounts of Q-7. The polar lipid profile was composed of the major lipids diphosphatidylglycerol and phosphatidylethanolamine and moderate amounts of phosphatidylglycerol and an unknown phospholipid; minor lipids were also detected. The fatty acid profile, with large amounts of C(16 : 0) and C(17 : 0) cyclo and the absence of C(12 : 0) 2-OH as hydroxylated fatty acid, also differed significantly from those reported for Alcaligenes species. On the basis of these data, it is proposed that strain CCUG 53761A(T) represents a novel genus and species, for which the name Paenalcaligenes hominis gen. nov., sp. nov. is proposed. The type strain of Paenalcaligenes hominis is CCUG 53761A(T) =CCM 7698(T). PMID:19684310
Furuncular myiasis caused by Dermatobia hominis is endemic throughout Central and South America. We report a case of furuncular myiasis in a traveler returned from Costa Rica. The case is unique because the primary care physician obtained magnetic resonance images. The images, however, do not show any characteristic features that assist in diagnosis.
Bhandari, Ramanath; Janos, David P.; Sinnis, Photini
This is the first in vitro study on the activity of 20 kinds of crude extracts of traditional Chinese medicine (TCM) on the intestinal parasite, Blastocystis hominis using the criteria of living cell count (LCC) and living cell rate (LCR). LCC and LCR were applied as observation indicators, the former as a fixed-quantity and the latter as a fixed-quality method.
L. Q. Yang; Mulkit Singh; E. H. Yap; G. C. Ng; H. X. Xu; K. Y. Sim
Mycoplasma salivarium infections outside the oral cavity are rare. We describe a 49-year-old man with laryngeal cancer and right pleural space infection with M. salivarium. To our knowledge, this is the first report of empyema due to Mycoplasma salivarium.
Recently, the genomes of two Mycoplasma fermentans strains, namely M64 and JER, have been completely sequenced. Gross comparison indicated that the genome of M64 is significantly bigger than the other strain and the difference is mainly contributed by the repetitive sequences including seven families of simple and complex transposable elements ranging from 973 to 23,778 bps. Analysis of these repeats resulted in the identification of a new distinct family of Integrative Conjugal Elements of M. fermentans, designated as ICEF-III. Using the concept of “reaction connectivity”, the metabolic capabilities in M. fermentans manifested by the complete and partial connected biomodules were revealed. A comparison of the reported M. pulmonis, M. arthritidis, M. genitalium, B. subtilis, and E. coli essential genes and the genes predicted from the M64 genome indicated that more than 73% of the Mycoplasmas essential genes are preserved in M. fermentans. Further examination of the highly and partly connected reactions by a novel combinatorial phylogenetic tree, metabolic network, and essential gene analysis indicated that some of the pathways (e.g. purine and pyrimidine metabolisms) with partial connected reactions may be important for the conversions of intermediate metabolites. Taken together, in light of systems and network analyses, the diversity among the Mycoplasma species was manifested on the variations of their limited metabolic abilities during evolution.
The de novo synthesized polar lipids of Mycoplasma species are rather simple, comprising primarily of the acidic glycerophospholipids PG and CL. In addition, when grown in a medium containing serum, significant amounts of PC and SPM are incorporated into the mycoplasma cell membrane although these lipids are very uncommon in wall-covered bacteria. The exogenous lipids are either incorporated unchanged or the PC incorporated is modified by a deacylation-acylation enzymatic cycle to form disaturated PC. Although their small genome, in some Mycoplasma species, other genes involved in lipid biosynthesis were detected, resulting in the synthesis of a variety of glycolipis, phosphoglycolipids and ether lipids. We suggest that analyses and comparisons of mycoplasma polar lipids may serve as a novel and useful tool for classification. Nonetheless, to evaluate the importance of polar lipids in mycoplasma, further systematic and extensive studies on more Mycoplasma species are needed. While studies are needed to elucidate the role of lipids in the mechanisms governing the interaction of mycoplasmas with host eukaryotic cells, the finding that a terminal phosphocholine containing glycolipids of M. fermentans serves both as a major immune determinants and as a trigger of the inflammatory responses, and the findings that the fusogenicity of M. fermentans with host cells is markedly stimulated by lyso-ether lipids, are important steps toward understanding the molecular mechanisms of M. fermentans pathogenicity.
Dermatobia hominis (human Bot fly) causes furuncular myiasis (larval infection) in Central and South America. This report describes a case in a member of the UK Armed Forces who had recently taken part in an overseas training exercise in Belize. The importance of clinical history (including travel history) is highlighted. We also describe the outcomes of conservative treatment and surgical intervention for separate lesions in the same patient. PMID:24079201
HeLa cells sensitive to the mitochondrial protein synthesis inhibitors erythromycin (ERY) and chloramphenicol (CAP) and HeLa variants resistant to the effects of these drugs were purposefully infected with drug-sensitive and -resistnat mycoplasmastrains. Mycoplasma hyorhinis and the ERY-resistant strain of Mycoplasma orale, MO-ERY/sup r/, did not influence the growth of HeLa and ERY-resistant ERY2301 cells in the presence or absence of ERY. M. hyorhinis also did not affect the growth of HeLa and CAP-resistant Cap-2 cells in the presence or absence of CAP. However, both HeLa and Cap-2 cells infected with the CAP-resistant strain of M. hyorhinis, MH-CAP/sup r/, were more sensitive to the cytotoxix effect of CAP. This maybe due to the glucose dependence of the cells, which was compromised by the increased utilization of glucose by MH-CAP/sup r/ in these infected cell cultures. In vitro protein synthesis by isolated mitochondria was significantly altered by mycoplasma infection of the various cell lines. A substantial number of mycoplasmas copurified with the mitochondria, resulting in up to a sevenfold increase in the incorporation of (/sup 3/H)leucine into the trichloroacetic acid-insoluble material. More importantly, the apparent drug sensitivity or resistance of mitchondrial preparations from mycoplasma-infected cells reflected the drug sensitivity or resistance of the contaminating mycoplasmas. These results illustrate the hazards in interpreting mitochondrial protein synthesis data derived from mycoplasma-infected cell lines, particularly putative mitochondrially encoded mutants resistant to inhibitors of mitochondrial protein synthesis.
Discard milk from sick or antibiotic-treated cows is often used as an economical alternative to milk re- placer at dairy farms. This practice poses a health risk to calves if the discard milk is from cows with mycoplasma mastitis. Mycoplasma bovis, Mycoplasma californicum, and Mycoplasma canadense are among the agents known to cause contagious mastitis in cattle and occasionally pneumonia,
J. A. Butler; S. A. Sickles; C. J. Johanns; R. F. Rosenbusch
Very little is understood of the structure of mycoplasma promoters, and this limits interpretation of genomic sequence data in these species. In this study the transcriptional start points of 22 genes of Mycoplasma pneumoniae were identified and the regions 5' to the start point compared. Although a strong consensus -10 region could be seen, there was only a weak consensus in the -35 region. A high proportion of transcripts had heterogeneous 5'-ends and characterisation of the sequence of the 5'-ends of two transcripts established that the heterogeneity was derived from initiation of transcription at reduced levels between 1 and 4 bases 5' to the major starting point. In addition to this apparently unique feature, a high proportion of transcripts lacked a 5' untranslated leader region that could contain a ribosomal binding site. Such leaderless transcripts are seen rarely in other bacterial species. Although the promoter regions for a number of members of lipoprotein multigene families were examined, no obvious explanation for regulation of expression was apparent. Using the data from this study an improved matrix for prediction of M.pneumoniae promoters was derived. Application of this matrix to the sequences immediately 3' and 5' to each predicted start codon in the genome suggested that most M. pneumoniae transcriptional start points were likely to occur between 5 and 30 bases 5' to the start codon. PMID:11071937
An inactivated Mycoplasma pneumoniae vaccine was evaluated in a double-blind study for safety, antigenicity and efficacy in marine recruits. The vaccine appeared to be safe and resulted in a significant reduction in the number of vaccinees hospitalized wi...
Commercial laying hens maintained under controlled conditions were experimentally inoculated with the S6 strain of Mycoplasma gallisepticum (S6MG) at 45 wk of age. This resulted in depressed liver lipid concentration, and inoculations at 20 and 45 wk affected the size of various portions of the reproductive tract. In 2 consecutive trials of the current study, the effect of age of application of S6MG inoculation on the egg yolk characteristics of commercial layers similarly housed and maintained under controlled conditions was determined. The ages of inoculation compared were prior to lay at 10 wk of age, during onset of lay at 22 wk of age, and during postpeak lay at 45 wk of age. In each trial, yolk moisture and total lipid content were determined at 24, 32, 43, 47, and 58 wk of age. Yolk cholesterol concentration and yolk fatty acid profiles at wk 47 and 58 were also examined. Data from wk 24, 32, and 43 (effects of S6MG inoculations at 10 and 22 wk) and data from wk 47 and 58 (effects of S6MG inoculations at 10, 22, and 45 wk) were analyzed separately. The data of both trials were pooled then analyzed together. Across wk 47 and 58, percentage yolk lipid was significantly lower in eggs laid by birds inoculated at 10 wk compared with those inoculated at 45 wk. Sham-inoculated control and 22-wk inoculated groups had intermediate percentage yolk lipids. Compared with sham-control and 10-wk S6MG inoculation groups across wk 47 and 58, yolk myristic, oleic, and linolenic acid concentrations were reduced, whereas yolk stearic and arachidonic acid levels were increased by either 22- or 45-wk S6MG inoculations. In comparison with all other treatment groups at wk 47, yolk linoleic acid concentration was reduced by S6MG inoculation at 45 wk. Variable postpeak alterations in yolk total lipid and fatty acid content occur in response to the timing of S6MG inoculation in layers housed under controlled conditions. PMID:16903485
Peebles, E D; Basenko, E Y; Branton, S L; Whitmarsh, S K; Maurice, D V; Gerard, P D
In 2 consecutive trials of the current study, the effect of the age of application of S6-strainMycoplasma gallisepticum (S6MG) inoculation on the blood characteristics of commercial layers housed and maintained under controlled conditions was determined. The ages of inoculation compared were those before lay at 10 wk of age, during onset of lay at 22 wk of age, and during postpeak lay at 45 wk of age. In each trial, hematocrit, plasma protein, and serum cholesterol, triglycerides, and Ca were determined at 20, 24, 32, 43, 47, and 58 wk of age. The data from both trials were pooled then analyzed together, whereas, data from wk 20 (effect of 10-wk S6MG inoculation); data from wk 24, 32, and 43 (effects of 10- and 22-wk S6MG inoculations); and data from wk 47 and 58 (effects of 10-, 22-, and 45-wk S6MG inoculations) were analyzed separately. At wk 20, hematocrit was higher in birds inoculated with S6MG at 10 wk compared with sham-inoculated birds, and across wk 24, 32, and 43, serum Ca was higher in birds inoculated with S6MG at 10 or 22 wk compared with those that were sham-inoculated. Serum Ca level across wk 47 and 58 was higher in birds inoculated with S6MG at 10 wk compared with sham-inoculated controls and birds inoculated with S6MG at 22 wk, with 45-wk S6MG-inoculated birds being intermediate. The response of serum cholesterol level at 47 wk to S6MG inoculation at either 10, 22, or 45 wk compared with controls was nearly opposite to that of the response observed at 58 wk. However, serum triglycerides were depressed only at wk 47 due to the 45-wk S6MG inoculation compared with all other treatments. Variable post-peak alterations in serum Ca and lipids occur in response to the timing of S6MG inoculation in layers housed under controlled conditions. PMID:17032838
Peebles, E D; Basenko, E Y; Branton, S L; Whitmarsh, S K; Gerard, P D
Effects of 2 supplemental concentrations of dietary poultry fat (PF) and the combination of PF, phytase (PHY), and 25-hydroxycholecalciferol [25(OH)D] on the gross digestive and reproductive organ characteristics of commercial layers inoculated with F-strainMycoplasma gallisepticum (FMG) were investigated in 2 trials. Sham and FMG inoculations were administered at 12 wk (before lay) or 22 wk (onset of lay), dietary treatments [basal control diet (BCD); BCD with 0.75% supplemental PF (LPFD); BCD with 1.50% supplemental PF (HPFD); HPFD additionally supplemented with 0.013% PHY and 0.025% 25(OH)D] were initiated at 20 wk of age, and organ characteristics were determined at 58 wk of age. In proportion to small intestine weight, jejuna were heavier in birds inoculated at 22 wk rather than at 12 wk of age. In hens inoculated at 22 wk of age, percentage of infundibulum weight was increased by FMG. The proportional length of infundibula in birds fed HPFD with PHY and 25(OH)D was longer than in those fed LPFD. In birds inoculated with FMG at 22 wk of age, BW was greater in those fed HPFD with or without added PHY and 25(OH)D in comparison with those fed LPFD, whereas LPFD increased percentage of oviduct and magnum weights when compared with the HPFD and BCD groups, respectively. Percentage of duodenum weight in birds that were fed HPFD with PHY and 25(OH)D was greater compared with those fed LPFD in the wk 22 sham and wk 12 FMG inoculation groups, but was also greater than in those fed BCD in the wk 12 FMG inoculation group. Conversely, percentage of duodenum weight was greater in birds fed LPFD compared with those fed HPFD after a wk 22 FMG inoculation. However, despite the effects of the supplemental combination of 1.50% PF, PHY, and 25(OH)D on the oviduct and small intestine structures, it did not result in a subsequent influence on layer performance, as indicated in a previous companion report. PMID:21406365
Peebles, E D; Park, S W; Branton, S L; Gerard, P D; Womack, S K
Effects of F-strainMycoplasma gallisepticum (FMG) inoculation and 1.5% supplemental dietary poultry fat (PF) on the digestive and reproductive organ characteristics of commercial layers at 58 wk of age were investigated. Sham and FMG inoculations were administered at 12 (before lay) and 22 (early in lay) wk of age, and dietary treatments (basal control diets and basal control diets with PF) were initiated at 20 wk of age. Supplemental PF increased BW and decreased isthmal length relative to total oviduct length in hens. Various oviduct segments were also affected by the type and age of inoculation, and these effects were further influenced by the use of PF. In comparison to their time-specific sham-inoculated controls, infundibulum weight relative to BW was increased when birds were inoculated with FMG at 22 wk, whereas isthmus weight relative to total oviduct weight was increased by FMG inoculation at 12 wk of age. However, PF affected infundibulum length relative to total oviduct length only in sham-inoculated birds, and PF increased magnum weight relative to total oviduct weight only in birds inoculated at 22 wk of age (sham or FMG). Furthermore, PF decreased isthmus weight relative to total oviduct weight only in birds that were sham-inoculated (12 or 22 wk). In conclusion, the inoculation of FMG at 12 or 22 wk may increase the relative contributions of the isthmus and infundibulum, respectively, to the total mass of the oviduct. In addition, PF may decrease the relative length of the isthmus and increase the relative weight of the magnum in the oviducts of birds that have been inoculated at 22 wk of age (sham or FMG). Previous studies have shown 1.5% supplemental dietary PF to influence feed consumption throughout lay and performance early in lay in hens that were inoculated with FMG at 12 wk of age. However, the current results suggest that these influences are associated with gross changes in the oviduct but not the digestive tract of layers. PMID:20075276
Park, S W; Burnham, M R; Branton, S L; Gerard, P D; Womack, S K; Peebles, E D
Mycoplasma pneumoniae is a common cause of community-acquired pneumonia, but this organism can also affect almost every organ system besides the lung. Neurological manifestations (meningitis, encephalitis, transverse myelitis, Guillain-Barrč syndrome) are the most frequent extrapulmonary complications of Mycoplasma pneumoniae. We report a case of a immunocompetent child affected by encephalitis caused by Mycoplasma pneumoniae. PMID:18710062
The use of animal and plant derived raw materials in mammalian cell culture processes may provide a possible route of entry for adventitious contaminants such as mycoplasma. Mycoplasma contaminations of cell culture represent a serious challenge to the production of biotechnology derived therapeutics. The slow growing nature of mycoplasma can disguise their infection of cultures since cells may continue to
The growth of three pathogenic goat mycoplasmas, strains Y, KH1 and Mycoplasma mycoides var. capri (PG3), was studied. They formed classical colonies on agar containing 1/500 thallium acetate. They were inactivated during storage at 2 to 4 C and by freezing and thawing but not by shaking. Only KH1 was killed by sonic treatment. Ultraviolet inactivation curves showed that their colony-forming units were single binucleate cells. Details of their growth curves are given. Filtration through 0.45- or 0.3-?m membrane filters removed up to 97% of the cells. Less than 0.003% passed 0.22-?m membranes. In electron micrographs, the cells were seen replicating by budding and most were 0.6 to 0.9 ?m in diameter; but cells between 0.1 and 0.2 ?m reproduced. They usually multiplied by producing one bud, a form of binary fission. However, two buds were produced by some synchronized cells, indicating that both nuclei had divided simultaneously to form progeny, an alternate method of multiplication. Images
Cryptosporidium hominis and Cryptosporidium parvum are associated with massive disease outbreaks world- wide. Because these two species have different transmission cycles, identification of these parasites to the species level in clinical samples may provide laboratory data of crucial importance in epidemiologic investi- gations. To date, the most reliable way to differentiate C. hominis and C. parvum is based on DNA
Kakali Bandyopadhyay; Kathryn L. Kellar; Iaci Moura; Maria Cristina Casaqui Carollo; Thaddeus K. Graczyk; Susan Slemenda; Stephanie P. Johnston; Alexandre J. da Silva
A prospective study was carried out to investigate the epidemiology and clinical significance of Blastocystis hominis in the following groups of the population of the city of Salamanca (Spain): in children attending 11 day car centres and 7 primary schools, two fecal samples were obtained from each child, and in 1231 patients attending the Clinical Hospital. A B. hominis incidence
A. M. Martín-Sánchez; A. Canut-Blasco; J. Rodriguez-Hernandez; I. Montes-Martínez; J. A. García-Rodríguez
The DNA recombination and repair machinery of Mycoplasma pneumoniae is composed of a limited set of approximately 11 proteins. Two of these proteins were predicted to be encoded by neighboring open reading frames (ORFs) MPN340 and MPN341. Both ORFs were found to have sequence similarity with genes that encode proteins belonging to the DNA helicase superfamily 1 (SF1). Interestingly, while a homolog of the MPN341 ORF is present in the genome of Mycoplasma genitalium (ORF MG244), MPN340 is an M. pneumoniae-specific ORF that is not found in other mycoplasmas. Moreover, the length of MPN340 (1590 base pairs [bp]) is considerably shorter than that of MPN341 (2148 bp). Examination of the MPN340-encoded amino acid sequence indicated that it may lack a so-called 2B subdomain, which is found in most SF1 DNA helicases. Also, the MPN340-encoded amino acid sequence was found to differ between subtype 1 strain M129 and subtype 2 strain FH at three amino acid positions. Both protein variants, which were termed PcrA(s) M129 and PcrA(s) FH, respectively, as well as the MPN341- and MG244-encoded proteins (PcrA Mpn and PcrA Mge , respectively), were purified, and tested for their ability to interact with DNA. While PcrA Mpn and PcrA Mge were found to bind preferentially to single-stranded DNA, both PcrA(s) M129 and PcrA(s) FH did not demonstrate significant DNA binding. However, all four proteins were found to have divalent cation- and ATP-dependent DNA helicase activity. The proteins displayed highest activity on partially double-stranded DNA substrates carrying 3' single-stranded extensions. PMID:23894687
Estevăo, Silvia; van der Heul, Helga U; Sluijter, Marcel; Hoogenboezem, Theo; Hartwig, Nico G; van Rossum, Annemarie M C; Vink, Cornelis
Powelson, Dorothy M. (Stanford Research Institute, Menlo Park, Calif.). Metabolism of animal cells infected with mycoplasma. J. Bacteriol. 82:288–297. 1961.—The effect of pleuropneumonia-like organisms (PPLO) upon the metabolism of tissue cultures was tested by comparing the assimilation and accumulation of the amino acids in the medium during growth and maintenance of monolayers of mouse fibroblasts (L strain) and human bone marrow cells (Mox). This preliminary study indicates that PPLO do alter the amino acid metabolism of animal cells. The observed changes in metabolic patterns shown by the infected fibroblast cultures did not mirror the metabolic patterns of the PPLO in the medium alone. Different strains of animal cells showed different responses to one PPLO strain, and different strains of PPLO caused different responses in one strain of animal cells. The PPLO did not grow in the tissue culture medium (no. 199 plus 2% horse serum and 20 to 40 units of penicillin/ml) nor in spent culture fluids and rapidly died off at 37 C but survived for months at 4 C. The altered metabolism of the infected tissue cultures appeared to reflect a true host-parasite interaction.
The lipids of the sterol nonrequiring Mycoplasmastrain S743 were found to include both ester glycerophosphatides (phosphatidylglycerol, acylphosphatidylglycerol, and diphosphatidylglycerol) and ceramide glycerophosphate compounds containing N-hydroxyacyl groups. The major phosphosphingolipid was tentatively identified as a hydroxyceramidephosphorylglycerol containing an O-acyl group. These compounds became labeled during growth in the presence of 32P-orthophosphate, 14C-glycerol, or 14C-palmitate. The lipid fraction also contained free long-chain base. 14C-palmitate was converted to labeled sphinganine. The long-chain base composition of the lipids was modified by growing the organisms in media containing different fatty acids, which were converted to bases containing two more C atoms per molecule. Ninety per cent of the long-chain base from cells grown in medium supplemented with elaidate consisted of monounsaturated C20 base. Images
The biosynthesis of cholesteryl glucoside by Mycoplasma gallinarum strain J proceeds by the transfer of glucose from uridine-5?-diphosphoglucose to membrane-bound sterol. Galactose also can be coupled to cholesterol via uridine-5?-diphosphogalactose. The reaction is specific for the uridine-5?-diphospho sugars. Enzymatic activity is associated with the membrane. Treatment of the membrane to remove endogenous sterol inactivates the enzyme. Only sterol which has been bound to the membrane participates in the reaction. The optimum pH is about 8.0, and Mg2+ is required. The reaction is unaffected by nucleotide triphosphate, uridine-5?-monophosphate, and uridine-5?-diphosphate. Reduction of pH to the optimum for ?-glucosidase in the membrane results in loss of synthesized glucoside. The enzyme is saturated at 0.5 mm uridine-5?-diphosphoglucose. The apparent Km of 2.05 × 10?7 indicates a high affinity of the enzyme for the nucleotide sugar.
...2013-04-01 false Mycoplasma detection media and components. 864.2360 Section...Products Â§ 864.2360 Mycoplasma detection media and components. (a) Identification. Mycoplasma detection media and components are used to detect...
Polyomavirus hominis 1 (BK virus, BKV) is an important pathogen in the field of transplantation medicine. BKV reactivation among renal-transplant recipients could cause BK associated nephropathy, which has unfavorable prognosis and is a cause for graft rejection. It is not clear why only few transplanted patients develop BK associated nephropathy while most exhibit asymptomatic viruria. One of the possible reasons lies in the mutations of the VP1 gene, encoding the main structural protein, bearing important determinants for the recognition of specific cellular receptors. The change of amino acid sequence could result in altered pathogenicity of BKV. The amplified sequences of BK in this research were from urines of patients with various clinical conditions along with healthy individuals. Nevertheless the sequence analysis which was undertaken did not show correlation between the viral genotype and the clinical condition. It was demonstrated that the most common BKV genotype in Bulgaria is genotype I and that the strains common in Bulgaria (genotypes I and IV) have typical European origin. Most of the sequenced BKV DNA samples (8/10) were correlated with the highest degree of similarity (81%) to the subcluster Ib. A specific place among the samples is taken by Pr-9, amplified from the urine of a pregnant woman that has a different evolutionary origins and might establish the beginning of a new distinct BKV strain. PMID:20029813
Cell of Mycoplasma pneumoniae FH gliding on a glass surface in liquid medium were examined by microscopic observation and quantitatively by microcinematography (30 frames per min). Comparisons were made only within the individual experiments. The cells moved in an irregular pattern with numerous narrow bends and circles. They never changed their leading end. The average speed (without pauses) was relatively constant between o.2 and 0.5 mum/s. The maximum speed was about 1.5 to 2.0 mum/s. The movements were interrupted by resting periods of different lengths and frequency. Temperature, viscosity, pH, and the presence of yeast extract in the medium influenced the motility significantly; changes in glucose, calcium ions, and serum content were less effective. The movements were affected by iodoacetate, p-mercuribenzoate, and mitomycin C at inhibitory or subinhibitory concentrations. Sodium fluoride, sodium cyanide, dinitrophenol, chloramphenicol, puromycin, cholchicin, and cytochalasin B at minimal inhibitory concentrations did not affect motility. The movements were effectively inhibited by anti-M. pneumoniae antiserum. Studies with absorbed antiserum suggested that the surface components involved in motility are heat labile. The gliding of M. pneumoniae cells required an intact energy metabolism and the proteins involved seemed to have a low turnover. PMID:14925
In this study the enzymatic activity of Mycoplasma agalactiae MAG_5040, a magnesium-dependent nuclease homologue to the staphylococcal SNase was characterized and its antigenicity during natural infections was established. A UGA corrected version of MAG_5040, lacking the region encoding the signal peptide, was expressed in Escherichia coli as a GST fusion protein. Recombinant GST-MAG_5040 exhibits nuclease activity similar to typical sugar-nonspecific endo- and exonucleases, with DNA as the preferred substrate and optimal activity in the presence of 20 mM MgCl2 at temperatures ranging from 37 to 45°C. According to in silico analyses, the position of the gene encoding MAG_5040 is consistently located upstream an ABC transporter, in most sequenced mycoplasmas belonging to the Mycoplasmahominis group. In M. agalactiae, MAG_5040 is transcribed in a polycistronic RNA together with the ABC transporter components and with MAG_5030, which is predicted to be a sugar solute binding protein by 3D modeling and homology search. In a natural model of sheep and goats infection, anti-MAG_5040 antibodies were detected up to 9 months post infection. Taking into account its enzymatic activity, MAG_5040 could play a key role in Mycoplasma agalactiae survival into the host, contributing to host pathogenicity. The identification of MAG_5040 opens new perspectives for the development of suitable tools for the control of contagious agalactia in small ruminants. PMID:23469065
Cacciotto, Carla; Addis, Maria Filippa; Coradduzza, Elisabetta; Carcangiu, Laura; Nuvoli, Anna Maria; Tore, Gessica; Dore, Gian Mario; Pagnozzi, Daniela; Uzzau, Sergio; Chessa, Bernardo; Pittau, Marco; Alberti, Alberto
In this study the enzymatic activity of Mycoplasma agalactiae MAG_5040, a magnesium-dependent nuclease homologue to the staphylococcal SNase was characterized and its antigenicity during natural infections was established. A UGA corrected version of MAG_5040, lacking the region encoding the signal peptide, was expressed in Escherichia coli as a GST fusion protein. Recombinant GST-MAG_5040 exhibits nuclease activity similar to typical sugar-nonspecific endo- and exonucleases, with DNA as the preferred substrate and optimal activity in the presence of 20 mM MgCl2 at temperatures ranging from 37 to 45°C. According to in silico analyses, the position of the gene encoding MAG_5040 is consistently located upstream an ABC transporter, in most sequenced mycoplasmas belonging to the Mycoplasmahominis group. In M. agalactiae, MAG_5040 is transcribed in a polycistronic RNA together with the ABC transporter components and with MAG_5030, which is predicted to be a sugar solute binding protein by 3D modeling and homology search. In a natural model of sheep and goats infection, anti-MAG_5040 antibodies were detected up to 9 months post infection. Taking into account its enzymatic activity, MAG_5040 could play a key role in Mycoplasma agalactiae survival into the host, contributing to host pathogenicity. The identification of MAG_5040 opens new perspectives for the development of suitable tools for the control of contagious agalactia in small ruminants.
Cacciotto, Carla; Addis, Maria Filippa; Coradduzza, Elisabetta; Carcangiu, Laura; Nuvoli, Anna Maria; Tore, Gessica; Dore, Gian Mario; Pagnozzi, Daniela; Uzzau, Sergio; Chessa, Bernardo; Pittau, Marco; Alberti, Alberto
Lipoprotein LppQ, a predominant 48-kDa antigen, and its corresponding gene, lppQ, were characterized in Mycoplasma mycoides subsp. mycoides SC, the etiological agent of contagious bovine pleuropneumonia. The lppQ gene is specific to M. mycoides subsp. mycoides SC and was found in the type strain and in field strains isolated in Europe, Africa, and Australia, as well as in vaccinal strains.
Mycoplasma fermentans and other Mycoplasma species are colonizers of human mucosal surfaces and may be associated with human immunodeficiency virus infection. While many infectious agents have been described in different percentages of patients with Chronic Fatigue Syndrome (CFS), little is known about the prevalence of mycoplasmas and especially M. fermentans in CFS patients. A polymerase chain reaction (PCR)-based assay was
A Vojdani; P. C Choppa; C Tagle; R Andrin; B Samimi; C. W Lapp
To analyze the risk factors for HPV infection in the urethra, we examined the prevalence of various microorganisms, for example Neisseria gonorrhoeae, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasmahominis, Ureaplasma urealyticum, Ureaplasma parvum, Gardnerella vaginalis, and human papillomavirus (HPV) in Japanese male patients with urethritis, and investigated their sexual backgrounds. Rubbed samples obtained from the distal urethra and questionnaires regarding sexual activity and demographic information were collected from 176 participants. N. gonorrhoeae, C. trachomatis, M. genitalium, M. hominis, U. urealyticum, U. parvum, G. vaginalis, and HPV were detected in 19, 26, 18, 12, 12, 8.5, 14, and 20%, respectively, of all cases in this study. Multivariate logistic regression analysis indicated that more than 4 sexual partners within the last year and presence of N. gonorrhoeae and/or C. trachomatis and/or M. genitalium infections were independent risk factors for urethral HPV infection, with odds ratios of 3.85 (95% CI 1.49-9.94) and 2.41 (95% CI 1.03-5.61), respectively. It is likely that urethral HPV detection is associated with current sexual activity and the presence of N. gonorrhoeae, C. trachomatis, and/or M. genitalium infections. PMID:21213011
One of the main problems in cell culture is mycoplasma infection. It can extensively affect cell physiology and metabolism. As the applications of cell culture increase in research, industrial production and cell therapy, more concerns about mycoplasma contamination and detection will arise. This review will provide valuable information about: 1. the ways in which cells are contaminated and the frequency and source of mycoplasma species in cell culture; 2. the ways to prevent mycoplasma contamination in cell culture; 3. the importance of mycoplasma tests in cell culture; 4. different methods to identify mycoplasma contamination; 5. the consequences of mycoplasma contamination in cell culture and 6. available methods to eliminate mycoplasma contamination. Awareness about the sources of mycoplasma and pursuing aseptic techniques in cell culture along with reliable detection methods of mycoplasma contamination can provide an appropriate situation to prevent mycoplasma contamination in cell culture.
Mycoplasma infections are of great concern in avian medicine, because they cause economic losses in commercial poultry production. A multiplex polymerase chain reaction (PCR) was optimized to simultaneously detect four pathogenic species of avian mycoplasmas. Four sets of oligonucleotide primers specific forMycoplasma gallisepticum(MG),M. synoviae(MS),M. meleagridis(MM) andM. iowae(MI) were used in the test. By using agarose gel electrophoreses for detection of
Mycoplasma agalactiae is the main cause of contagious agalactia, a serious disease of sheep and goats, which has major clinical and economic impacts. We have developed a multilocus sequence typing (MLST) scheme using the sequenced genomes of the M. agalactiae strains PG2 and 5632. An MLST scheme based on the genes gltX, metS, gyrB, tufA and dnaA was designed and in total 3468 bp of sequence were analysed for each strain. MLST offers a highly discriminatory typing method for M. agalactiae and was capable of subdividing 53 strains into 17 distinct sequence types, largely according to geographical origin. MLST detected unexpected diversity in recent isolates from Spain, identifying two novel outliers, and enabled typing of novel Mongolian isolates for the first time. Genetic diversity in the sequenced regions was largely due to mutation, with recombination playing a much smaller role. A web-accessible database has been set up for this MLST scheme for M. agalactiae: http://pubmlst.org/magalactiae/. MLST offers a robust, objective molecular epidemiological tool for M. agalactiae that that enables interlaboratory comparison of data. PMID:21372188
McAuliffe, L; Gosney, F; Hlusek, M; de Garnica, M L; Spergser, J; Kargl, M; Rosengarten, R; Ayling, R D; Nicholas, R A J; Ellis, R J
In this study, the technical performance of culture, two commercially available polymerase chain reaction (PCR) tests, rapid plate agglutination (RPA) test, hemagglutination inhibition (HI) test, and eight commercially available enzyme-linked immunosorbent assays (ELISAs) were compared for the detection of avian mycoplasma infections from 3 days postinfection (d.p.i.) through 35 d.p.i. The tests were carried out on samples from specified pathogen-free layers that were infected at 66 wk of age with recent Mycoplasma synoviae (MS) and Mycoplasma gallisepticum (MG) field strains, MS and MG ATCC strains, and Mycoplasma imitans (MIM), respectively. Results showed a high percentage of positive samples in the homologous infected groups and a high percentage of negative samples (100%) in the uninfected and heterologous infected groups during 35 d.p.i. of both culture and PCR tests. For the group infected with the MG 15302 ATCC strain, serology was more sensitive than bacteriology. All MG and MS tests, with the exception of MG ELISA kit D showed a lower percentage of positive samples during 35 d.p.i. for the detection of the MG and MS ATCC strain infection compared with that of the field strains. Also, the number of cross-reactions (false positives) in the serologic tests was lower after infection with an ATCC strain than after an infection with the MG or MS field strain. Contradictory to other studies, the ELISAs and the RPA test using undiluted serum showed a relatively high number of false-positive results. The MG ELISAs (except ELISA kit D) showed more false-positive results (up to 37%) in the MIM-infected group than in the MS-infected groups. This was not unexpected, as MIM and MG have a close antigenic relationship. The results of the serologic tests in this study showed that a certain level of false-positive results can be expected in about any serologic test. Although the level of false-positive results varied between several serologic tests, this study showed that it is not advisable to rely completely on one test (system) only. PMID:16094832
Feberwee, A; Mekkes, D R; de Wit, J J; Hartman, E G; Pijpers, A
Production of ducks and geese in certain parts of the world is very important. Mycoplasma diseases cause significant losses to the duck and goose industry. This review summarizes the epidemiological, clinical, and pathomorphological characteristics of mycoplasma diseases of ducks and geese and the involvement of the various mycoplasma species in their pathogenesis. The role of mycoplasma infections in the development of clinical signs, pathological lesions, and mortality of challenged birds is demonstrated in challenge experiments. Transmission of mycoplasma in the ovary and eggs resulting in the reduction of egg production and an increase of embryo mortality has been shown in challenge experiments as well as in field studies. The susceptibility of many mycoplasma isolates of the most important mycoplasma species of duck and goose origin were tested and showed relatively high average minimum inhibitory concentrations of lincomycin, tilosin, oxytetracycline, chlortetracycline, and enrofloxacin but not for tiamulin. The successful treatment of mycoplasma infections with antibiotics in ducks and geese should be selected based on the minimum inhibitory concentration values against the mycoplasmas isolated from the flock. PMID:23091137
Isolation of mycoplasmas from the oropharynxes of 60 fantails reared under natural conditions at different zoological parks in Miyazaki prefecture was carried out. Mycoplasma columbinum, M. columborale and M. columbinasale were isolated from 28 (46.7%), 22 (36.7%), and 1 (1.7%) of 60 oropharynxes, respectively, but no Mycoplasma was isolated from 5 cloacas tested. Ureaplasma was not isolated from any of the 28 oropharynxes or 5 cloacas examined. We report that 41 (68.3%) of 60 fantails had one or two species of mycoplasmas in their oropharynxes, and make the first confirmation of M. columbinasale inhabiting a Japanese pigeon. PMID:9234221
Blastocystis hominis (B. hominis) is a parasite that often causes gastrointestinal symptoms in patients with immune deficiency and has a controversial pathogenicity in healthy people, although some symptoms are reported outside of the gastrointestinal system in healthy persons. Henoch-Schönlein Purpura (HSP) vasculitis is an acute autoimmune disease characterised by IgA storage of small vessels that is believed to include infectious factors in its aetiology. A 30-month follow-up with a boy diagnosed with HSP being treated with steroid therapy showed that he had recurrent symptoms within two days, and B. hominis was detected in the faecal analysis. His symptoms including rash, abdominal pain, and arthritis improved after treatment with steroid and co-trimaksazol. This paper is the first to present a case of HSP associated with B. hominis. PMID:23955912
Tutanç, Murat; Silfeler, Ibrahim; Ozgür, Tümay; Motor, Vicdan Köksald?; Kurto?lu, Ahmet Ibrahim
Blastocystis hominis and Endolimax nana exist as two separate parasitic organisms; however co-infection with the two individual parasites has been well documented. Although often symptomatic in immunocompromised individuals, the pathogenicity of the organisms in immunocompetent subjects causing gastrointestinal symptoms has been debated, with studies revealing mixed results. Clinically, both B. hominis and E. nana infection may result in acute or chronic diarrhea, generalized abdominal pain, nausea, vomiting, flatulence and anorexia. We report the case of a 24-year-old immunocompetent male presenting with chronic diarrhea and abdominal pain secondary to B. hominis and E. nana treated with metronidazole, resulting in symptom resolution and eradication of the organisms. Our case illustrates that clinicians should be cognizant of both B. hominis and E. nana infection as a cause of chronic diarrhea in an immunocompetent host. Such awareness will aid in a timely diagnosis and possible parasitic eradication with resolution of gastrointestinal symptoms.
Shah, Mitanshu; Tan, Christopher Bryan; Rajan, Dhyan; Ahmed, Shadab; Subramani, Krishnaiyer; Rizvon, Kaleem; Mustacchia, Paul
The presence of foreign contamination, especially of mycoplasmas, is a major hindrance in long term in vitro cultivation of Plasmodium falciparum and may be a source of false-positive results. Efforts have been made to control mycoplasma contamination by trypsinization of P. falciparum culture. Samples of accidentally contaminated cultures were used for this study. The presence of Mycoplasma orale in contaminated culture was ascertained by a species-specific PCR-based mycoplasma detection kit (Takara; Cat. No.6601). Trypsinization was carried out using trypsin-EDTA and the growth profile of P. falciparum was monitored for more than three weeks post-trypsinization. The studies were carried out with four different P. falciparum strains, various serum supplements and human erythrocytes belonging to different blood groups. It was interesting to observe that, irrespective of the different strains of P. falciparum and the variety of serum supplements and erythrocytes, mycoplasma contamination can successfully be removed from P. falciparum culture by trypsinization. No antibiotic except gentamicin, which is routinely used, was added to the medium. Results of this study indicate that the frequent appearance of mycoplasma in continuous long-term cultures of P. falciparum can be managed by trypsinization. PMID:23277231
We describe a 66-year-old woman with infective endocarditis due to Cardiobacterium hominis whose condition, complicated by severe aortic regurgitation and congestive heart failure, necessitated aortic valve replacement despite treatment with ceftriaxone followed by ciprofloxacin. The blood isolate of C. hominis produced ?-lactamase and exhibited high-level resistance to penicillin (MIC, ?256 ?g/ml) and reduced susceptibility to vancomycin (MIC, 8 ?g/ml).
The present study reports for the first time the completion of the life cycle of Cryptosporidium hominis in cell-free culture and multiplication of the parasite via qPCR. Individual life-cycle stages were characterised using Cryptosporidium-specific antibody staining (Sporo-Glo™) and fluorescent in situ hybridisation (FISH) staining on cultures inoculated with excysted oocysts and purified sporozoites. In both cultures, C. hominis successfully proliferated
Nawal Hijjawi; Annika Estcourt; Rongchang Yang; Paul Monis; Una Ryan
The male genitalia of Dermatobia hominis L. is described morphologically by the use of scanning electron microscopy (SEM) and compared, under light microscopy, with other Cuterebridae D. hominis differes from Metacuterebra apicalis (Guérin-Méneville) because it has longer gonopods, shorter parameter, a small crest and spines on the distiphallus a bellshaped gonopore with a prominent external outline, and surstyli with a spatulate distal region. PMID:2388247
Phytoplasmas and mycoplasmas are two groups of important pathogens in the bacterial class Mollicutes. Because of their economical and clinical importance, these obligate pathogens have attracted much research attention. However, difficulties involved in the empirical study of these bacteria, particularly the fact that phytoplasmas have not yet been successfully cultivated outside of their hosts despite decades of attempts, have greatly hampered research progress. With the rapid advancements in genome sequencing, comparative genome analysis provides a new approach to facilitate our understanding of these bacteria. In this study, our main focus is to investigate the evolution of gene content in phytoplasmas, mycoplasmas, and their common ancestor. By using a phylogenetic framework for comparative analysis of 12 complete genome sequences, we characterized the putative gains and losses of genes in these obligate parasites. Our results demonstrated that the degradation of metabolic capacities in these bacteria has occurred predominantly in the common ancestor of Mollicutes, prior to the evolutionary split of phytoplasmas and mycoplasmas. Furthermore, we identified a list of genes that are acquired by the common ancestor of phytoplasmas and are conserved across all strains with complete genome sequences available. These genes include several putative effectors for the interactions with hosts and may be good candidates for future functional characterization. PMID:22479625
NYC medium, primarily designed for isolation of pathogenic Neisseria, also readily supports the growth of large-colony mycoplasmas and T-mycoplasmas. When compared with A-3 agar medium and PPLO agar in clinical field trials, NYC medium performed equally as well as these mycoplasma-specific media in providing recovery of Mycoplasma species from female genital specimens. The transparent, highly-selective NYC medium permits direct, microscopic observation and presumptive identification of mycoplasmas, as well as Neisseria gonorrhoeae, without interference from contaminating saprophytes. As a single medium it can therefore be doubly useful in the diagnosis of gonorrhea and in the recognition of active or asymptomatic mycoplasma infections. Used in gonorrhea screening programs, the medium can be valuable in establishing the frequency of association of mycoplasmas with urogenital tract infection. Images
Faur, Yvonne C.; Weisburd, Martin H.; Wilson, Marion E.; May, Paul S.
Blastocystis hominis is probably the most common protozoan found in the human gut worldwide. In Taiwan, the prevalence of B. hominis infection is yet to be determined but is expected to be relatively higher among foreign workers. No data is available on the prevalence of B. hominis infection in long-term care facilities in Taiwan. This study included 713 subjects (552 residents and 161 care workers) from ten long-term care facilities in Taiwan who completed stool microscopic examinations with Merthiolate-iodine-formalin stain technique. The prevalence rate of blastocystosis was the highest among foreign and domestic care workers followed by residents (12.2%, 4.6%, and 2.7%, respectively). Older age (p = 0.04) and lower educational level (p = 0.008) were significantly associated with blastocystosis among care workers. Among residents, B. hominis infection was negatively associated with prolonged use of antibiotics within 3 months prior to examination (p = 0.05) and positively associated with tracheostomy in-place (p = 0.028). In conclusion, B. hominis infection was the most prevalent intestinal parasitic infection among both care workers and residents of long-term care facilities in Taiwan. Use of antibiotics was negatively associated with B. hominis infection among residents. Additionally, appropriate preventive measures should be implemented to older care workers with lesser educational attainment in order to reduce the risk of blastocystosis infection. PMID:19488784
Su, Fu-Hsiung; Chu, Fang-Yeh; Li, Chung-Yi; Tang, Hui-Fei; Lin, Yu-Shiang; Peng, Yu-Ju; Su, Yih-Ming; Lee, Shyh-Dye
Blastocystis hominis found in stool specimens has been the most frequently identified parasite among foreign workers from Southeast Asia in Taiwan since 1992. The prevalence of B. hominis was 14.1% in this study. In their quarantine physical examinations, 121 male Thai workers were examined hematologically and screened for stool parasites using the merthiolate-iodine-formaldehyde concentration method. Hematological values were compared in workers with and without a B. hominis infection. Multiple regressions were used to adjust for age. Those infected with any parasite other than B. hominis were excluded from further analysis. The workers infected with B. hominis had a lower leukocyte count (6.5+/-0.4 X 10(3)/microl) than those who were not (7.4+/-0.2 X 10(3)/microl). This was mainly caused by a reduced neutrophil count (3.2+/-0.4 vs 4.2+/-0.2 X 10(3)/microl). Hemoglobin (13.9+/-0.3 vs 14.5+/-0.1 g/dl) and hematocrit (41.4+/-0.6 vs 42.9+/-0.2%) were also reduced in B. hominis-positive workers. PMID:12743803
This study was designated to examine the pathogenicity of several strains of Mycoplasma gallisepticum (R, F, S?6, 227 and A5969) and laboratory derived substrains. Preliminary results indicated that the nine M. gallisepticum strains differed markedly in their pathogenicity for chickens. A comparison was made between various in vivo and in vitro methods for quantitative evaluation of pathogenicity. Reproducibility, convenience, and
Sharon Levisohn; M. J. Dykstra; M. Y. Lin; S. H. Kleven
Mycoplasma synoviae is a poultry pathogen causing respiratory disease and synovitis. An indirect enzyme-linked immunosorbent assay (ELISA) has previously been devised in our laboratory using the major membrane antigen MSPB of M. synoviae strain WVU 1853 as antigen. However, sera from chickens inoculated with the M. synoviae vaccine strain MS-H showed lower optical densities in the assay than chickens infected
Amir H. Noormohammadi; Glenn F. Browning; Jillian Jones; Kevin G. Whithear
Pathogenesis of Mycoplasma pneumoniae infection is considered to be in part attributed to excessive immune responses. Mycoplasma pneumoniae shows strong cytoadherence to host cells and this cytoadherence is thought to be involved in the progression of pneumonia. However, the interaction between the cytoadherence and the immune responses is not known in detail. In this study, we demonstrated that the induction of pro-inflammatory cytokines in the human monocyte cell line THP-1 is dependent on the property of cytoadherence of M. pneumoniae. A wild-type strain of M. pneumoniae with cytoadherence ability induced pro-inflammatory cytokines such as tumour necrosis factor-? and interleukin-1? (IL-1?). Whereas, heat-killed M. pneumoniae and cytoadherence-deficient mutants of M. pneumoniae caused significantly less production of pro-inflammatory cytokines than the wild-type strain. The wild-type strain induced pro-inflammatory cytokines in an endocytosis-independent manners, but the induction by heat-killed M. pneumoniae and cytoadherence-deficient mutants was dependent on endocytosis. Moreover, the wild-type strain induced caspase-1 production and ATP efflux, promoting the maturation of IL-1? and release of the pro-IL-1? precursor, whereas heat-killed M. pneumoniae and the cytoadherence-deficient mutants failed to induce them. These data suggest that the cytoadherence ability of M. pneumoniae activates immune responses and is involved in the pathogenesis of M. pneumoniae infection.
The aim of this work was to discover if Mycoplasma fermentans, which is known to infect B cells, could be the cause of the raised ecto-5?-nucleotidase observed in the synovial fluid of rheumatoid arthritis patients. The ecto-5?-nucleotidase activity in the patients' serum has been shown to correlate with the erythrocyte sedimentation rate and DNA from the mycoplasma has been found in the synovial fluid. B lymphoblastoid cell lines were exposed to 16 strains of Mycoplasma fermentans and their ecto-5?-nucleotidase, CD73, was measured both biochemically and by mouse antibodies to human ecto 5?-nucleotidase using the fluorescence activated cell sorter. The type strain, PG 18, did not grow with the B cells. Some of the mycoplasmastrains (9/15) increased the cellular ecto-5?-nucleotidase activity from twice to 17 fold, and usually showed 5?-nucleotidase activity themselves. At least one strain, M106, induced human 5?-nucleotidase on the normally 5?-nucleotidase negative Daudi and Raji Burkitt's lymphoma cell lines, and increased sevenfold the 5?-nucleotidase on the monocyte/macrophage cell line THP-1. Growing the cells in aged medium increased the level of mycoplasma infection. This mycoplasma-induced enzyme showed a conformational change and an increase in activity with a glycosylation change involving mannose groups. The other group of strains, mostly of respiratory or cell culture origin, usually did not have any 5?-nucleotidase of their own and decreased the B-cell enzyme activity by about half. Electron microscopy and flow cytometry showed that the strain M106 was filamentous and could be found inside the B-cells. The 5?-nucleotidase-inducing strains of M. fermentans may be important in the aetiology of rheumatoid arthritis.
Mycoplasma contamination of the licensed anthrax vaccine administered to military personnel has been suggested as a possible cause of Persian Gulf illness. Vaccine samples tested by nonmilitary laboratories were negative for viable mycoplasma and mycoplasma DNA and did not support its survival. Mycoplasma contamination of anthrax vaccine should not be considered a possible cause of illness.
Homologous internal controls were used as competitor DNA in the polymerase chain reaction for the quantitative detection of mycoplasma DNA. PCR primer sets were designed on the basis of the most conserved nucleotide sequences of the 16S rRNA gene of mycoplasma species. Amplification of this gene was examined in five different mycoplasma species: Mycoplasma orale, M. hyorhinus, M. synoviae, M.
Maninder K. Sidhu; Abbas Rashidbaigi; Douglas Testa; Mei-June Liao
A portion of the putative Mycoplasma gallisepticum (MG) cytadhesin gene was identified and used as a diagnostic DNA probe. Degenerate oligonucleotide primers corresponding to conserved regions of the cytadhesin proteins from two human mycoplasmas, M. pneumoniae and M. genitalium, were synthesized for use in the polymerase chain reaction (PCR) on genomic MG DNA. A 583-base-pair MG DNA fragment was amplified and subsequently cloned and sequenced. The MG DNA fragment is predicted to encode a 193-amino-acid peptide. This peptide demonstrates significant homology to the expected portions of the two human mycoplasmal cytadhesin proteins. Used as a probe to study the distribution of this fragment in pathogenic and nonpathogenic avian mycoplasmas, the PCR product hybridized to genomic DNA from all seven MG strains tested. However, it failed to hybridize to M. synoviae, M. meleagridis, M. iowae, or M. gallinarum DNA. PMID:8363503
Dohms, J E; Hnatow, L L; Whetzel, P; Morgan, R; Keeler, C L
Summary The polymerase chain reaction (PCR) method was used to detect mycoplasma contamination in a panel of 42 continuous cell lines.\\u000a According to the microbiological cultivation assay on agar, 29 cell lines were chronically infected and 13 cell lines were\\u000a negative. Sets of outer and inner primers (nested double-step PCR) were applied which anneal to DNA sequences coding for conserved\\u000a regions
Anne Hopert; Cord C. Uphoff; Manfred Wirth; Hansjörg Hauser; Hans G. Drexler
Mycoplasmas are commensals and pathogens of different avian species, especially poultry and passeriforms. The role of mycoplasmas in raptors has not yet been completely determined, and especially not the possibility of vertical transmission. Therefore 424 raptor eggs were examined for the occurrence of mycoplasmas using culture, and 155 of these eggs with a Mycoplasma genus-specific polymerase chain reaction (PCR) assay.
M. Lierz; N. Hagen; N. Harcourt-Brown; S. J. Hernandez-Divers; D. Lüschow; H. M. Hafez
Mycoplasma bovis is a pathogen causing respiratory disease, otitis media, arthritis, mastitis, and a variety of other diseases in cattle worldwide. It is increasingly recognized by the veterinary and livestock communities as having an important impact on the health, welfare, and productivity of dairy and beef cattle. M. bovis diseases can be difficult to diagnose and control because of inconsistent disease expression and response to treatments and vaccines, and large gaps in our understanding of the epidemiology and pathophysiology of these diseases. There are limited data on which to base evidence-based decisions for treatment and control, and the literature contains differing clinical biases and opinions. This document is intended for veterinarians dealing with cattle and is focused on the cattle production systems of North America. The goal of the consensus statement panel was to encourage an evidence-based approach to M. bovis problems. The scientific literature was critically reviewed, including peer-reviewed journal articles and reviews obtained by database searches using the terms "Mycoplasma bovis" or "mycoplasma + cattle." Where other data were lacking, conference proceedings were reviewed as a source of expert opinion. PMID:21745245
Maunsell, F P; Woolums, A R; Francoz, D; Rosenbusch, R F; Step, D L; Wilson, D J; Janzen, E D
There are several mechanisms used by pathogenic bacteria to evade host defenses, including antigenic variation on their surfaces. Antigenic variation of cell surface lipoproteins has been reported in a number of Mycoplasma species (3). Sequencing of the Mycoplasma pneumoniae genome has re- vealed that it harbors 46 lipoprotein genes in six multigene families, all of unknown function (4, 7). These
The cytotoxicities of various strains of Mycoplasma mycoides subsp. mycoides small colony type (SC), the agent of contagious bovine pleuropneumonia (CBPP), were measured in vitro using embryonic calf nasal epithelial (ECaNEp) cells. Strains isolated from acute cases of CBPP induced high cytotoxicity in the presence of glycerol, concomitant with the release of large amounts of toxic H2O2 that were found to be translocated into the cytoplasms of the host cells by close contact of the Mycoplasmastrains with the host cells. Currently used vaccine strains also showed high cytotoxicity and high H2O2 release, indicating that they are attenuated in another virulence attribute. Strains isolated from recent European outbreaks of CBPP with mild clinical signs, which are characterized by a defect in the glycerol uptake system, released small amounts of H2O2 and showed low cytotoxicity to ECaNEp cells. M. mycoides subsp. mycoides SC strain PG1 released large amounts of H2O2 but was only slightly cytotoxic. PG1 was found to have a reduced capacity to bind to ECaNEp cells and was unable to translocate H2O2 into the bovine cells, in contrast to virulent strains that release large amounts of H2O2. Thus, an efficient translocation of H2O2 into host cells is a prerequisite for the cytotoxic effect and requires an intact adhesion mechanism to ensure a close contact between mycoplasmas and host cells. PMID:17998309
Bischof, Daniela F; Janis, Carole; Vilei, Edy M; Bertoni, Giuseppe; Frey, Joachim
Trichomonades from the mouth were studied by Steinberg who proposed to group them into three distinct types; namely, Trichomonas elongata, Trichomonas caudata, and Trichomonas flagellata. Doflein (3) regards them as probably identical with Trichomonas hominis. Opinions differ as to whether or not Trichomonas vaginalis Donné and Trichomonas hominis Grassi are the same species. Lynch, for instance, believes that they are the same species, while von Prowazek (4), Bensen (5), and others (6, 7) insist that they are different types. Bensen's view seems to be well supported by the difference alleged to be found between the mode of encystment in the two trichomonades, were it not for the fact that our knowledge about the so called cyst of trichomonades is still obscure. According to Alexeieff (8) many of the so called cysts were evidently blastomyces contained in the cell body of the trichomonas. An autogamy alleged to take place in cysts as described by Bohne and von Prowazek (9) has not been confirmed by Dobell (10). And Wenyon (11) contends that it has never been found possible to produce any development of these cysts outside the body on the warm stage as can be done with the cysts of Entamśba coli. Therefore, it is still premature to take the process of encystment into consideration as far as the classification of trichomonas is concerned. On the other hand, Rodenwaldt (12) seems to think that there are many species of trichomonas in the human intestines, and Wenyon has described a new trichomonas from the human intestines (Macrostoma mesnili Wenyon). Further cultural studies in the morphology and biology of these organisms must be carried out in order to solve these problems. In the light of modern investigations there are five subgenera to be included under the genus Trichomonas Donné. They are as follows: (1) Protrichomonas Alexeieff, with three anterior flagella, without an undulating membrane. (2) Trichomastix Biitschli) with three anterior flagella and a trailing flagellum (Schleppgeissel) without an undulating membrane. (3) Trichomonas Donné, with three anterior flagella and an undulating membrane. (4) Macrostoma Alexeieff, Amend, Wenyon (11), with three anterior flagella and an undulating membrane wedged in a deep groove (peristome). (5) Tetratrichomonas Parisi (13), with four anterior flagella and an undulating membrane. As far as our culture trichomonas from the human mouth is concerned, it has been shown that it is not strictly a trichomonas and that it should be classed under the subgenus Tetratrichomonas.
The cytotoxicities of various strains of Mycoplasma mycoides subsp. mycoides small colony type (SC), the agent of contagious bovine pleuropneumonia (CBPP), were measured in vitro using embryonic calf nasal epithelial (ECaNEp) cells. Strains isolated from acute cases of CBPP induced high cytotoxicity in the presence of glycerol, concomitant with the release of large amounts of toxic H2O2 that were found
Daniela F. Bischof; Carole Janis; Edy M. Vilei; Giuseppe Bertoni; Joachim Frey
Restriction enzyme fingerprinting of genomic DNA and Southern blots probed with subclones of the Mycoplasma pneumoniae cytadhesin P1 gene were used to characterize clinical isolates of M. pneumoniae. On the basis of the examination of 29 individual M. pneumoniae isolates, two distinct groups were established. Group 1, which displayed a 12-kilobase band following DNA digestion with HindIII, consisted of strain M129-B16 and three others obtained in the state of Washington during the 1960s. The remaining M. pneumoniae strains belonged to group 2, which lacked the 12-kilobase band and included samples from the 1940s, 1970s, and 1980s. This category also included the only M. pneumoniae strain isolated from the synovial fluid of an arthritic patient. Images
Rationale: As the smallest free-living bacteria and a frequent cause of respiratory infections, mycoplasmas are unique pathogens. Mice infected with Mycoplasma pulmonis can develop localized, life-long airway infection accompanied by persistent inflammation and remodeling. Objective: Because mast cells protect mice from acute septic peritonitis and gram-negative pneumonia, we hypothesized that they defend against mycoplasma infection. This study tests this hypothesis using mast cell–deficient mice. Methods: Responses to airway infection with M. pulmonis were compared in wild-type and mast cell–deficient KitW-sh/KitW-sh mice and sham-infected control mice. Measurements and Main Results: Endpoints include mortality, body and lymph node weight, mycoplasma antibody titer, and lung mycoplasma burden and histopathology at intervals after infection. The results reveal that infected KitW-sh/KitW-sh mice, compared with other groups, lose more weight and are more likely to die. Live mycoplasma burden is greater in KitW-sh/KitW-sh than in wild-type mice at early time points. Four days after infection, the difference is 162-fold. Titers of mycoplasma-specific IgM and IgA appear earlier and rise higher in KitW-sh/KitW-sh mice, but antibody responses to heat-killed mycoplasma are not different compared with wild-type mice. Infected KitW-sh/KitW-sh mice develop larger bronchial lymph nodes and progressive pneumonia and airway occlusion with neutrophil-rich exudates, accompanied by angiogenesis and lymphangiogenesis. In wild-type mice, pneumonia and exudates are less severe, quicker to resolve, and are not associated with increased angiogenesis. Conclusions: These findings suggest that mast cells are important for innate immune containment of and recovery from respiratory mycoplasma infection.
Xu, Xiang; Zhang, Dongji; Lyubynska, Natalya; Wolters, Paul J.; Killeen, Nigel P.; Baluk, Peter; McDonald, Donald M.; Hawgood, Samuel; Caughey, George H.
Mycoplasma synoviae (MS) is an important pathogen of chickens and turkeys. In recent years sequence analysis of the partial MS variable lipoprotein and hemagglutinin A (vlhA) gene PCR product has been utilized routinely for MS strain genotyping. Several PCR assays have been proposed for the amplification of the conserved upstream region of the MS vlhA gene; however, in several clinical instances the published assays failed to generate vlhA PCR products from confirmed MS-positive cases. These occurrences hindered our capability to genotype those cases. In silico analysis of the published MS vlhA PCRs raised concerns, which were addressed by the design of revised MS vlhA PCRs. The published and revised assays were tested for their relative sensitivity and specificity with laboratory and clinical MS-positive samples. One of the revised MS vlhA PCRs (revised Hong) was demonstrated to be more sensitive and specific, and amplified all clinical samples analyzed in this study. PMID:21313852
We investigated the mode of action underlying the anti-mycoplasma activity of cationic antimicrobial peptides (AMPs) using four known AMPs and Mycoplasma pulmonis as a model mycoplasma. Scanning electron microscopy revealed that the integrity of the M. pulmonis membrane was significantly damaged within 30min of AMPs exposure, which was confirmed by measuring the uptake of propidium iodine into the mycoplasma cells. The anti-mycoplasma activity of AMPs was found to depend on the binding affinity for phosphatidylcholine, which was incorporated into the mycoplasma membrane from the growth medium and preferentially distributed in the outer leaflet of the lipid bilayer. PMID:23994526
Park, Ho Jin; Kang, Ki Mo; Dybvig, Kevin; Lee, Bok Luel; Jung, Yong Woo; Lee, In Hee
A Mycoplasma synoviae (Ms) species-specific recombinant subclone, pMS156-20, of approximately 1.1 kbp was partially sequenced. Based on the sequence data, a pair of 25 base primers were synthesized to develop a Ms polymerase chain reaction (Afs-PCR). The primers amplified target DNA of approximately 1.1 kbp and had excellent specificity over a range of annealing temperature from 60 degrees C to 68 degrees C. The primers amplified 100 pg of 26 strains or isolates of Ms, but not three strains of M. gallisepticum (S6, K810, A5969), 15 other avian Mycoplasma species, and pUC8 plasmid. The minimum amount of target DNA detected by Ms-PCR was 10 fg, which was 100,000 times more sensitive than the dot blot methodology using Ms recombinant DNA probes. The specificity of Ms-PCR product detected by gel electrophoresis was confirmed by Southern blot hybridization using an internal probe derived from pMSl56-20. PMID:18671038
Background Mycoplasma fermentans has been associated with rheumatoid arthritis. Recently, it was detected in the joints and blood of patients with rheumatoid arthritis, but it is not clear yet how the bacteria enter the body and reach the joints. The purpose of this study was to determine the ability of M. fermentans to induce experimental arthritis in rabbits following inoculation of the bacteria in the trachea and knee joints. Methods P-140 and PG-18 strains were each injected in the knee joints of 14 rabbits in order to evaluate and compare their arthritogenicity. P-140 was also injected in the trachea of 14 rabbits in order to test the ability of the bacteria to reach the joints and induce arthritis. Results M. fermentans produced an acute arthritis in rabbits. Joint swelling appeared first in rabbits injected with P-140, which caused a more severe arthritis than PG-18. Both strains were able to migrate to the uninoculated knee joints and they were detected viable in the joints all along the duration of the experiment. Changes in the synovial tissue were more severe by the end of the experiment and characterized by the infiltration of neutrophils and substitution of adipose tissue by connective tissue. Rabbits intracheally injected with P-140 showed induced arthritis and the bacteria could be isolated from lungs, blood, heart, kidney, spleen, brain and joints. Conclusion M. fermentans induced arthritis regardless of the inoculation route. These findings may help explain why mycoplasmas are commonly isolated from the joints of rheumatic patients.
Contagious caprine pleuropneumonia (CCPP), one of the most serious and dramatic diseases of goats, is caused by Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae). This organism is very difficult to isolate and to correctly identify. In a previous report we described a method for the rapid detection and identification of M. capripneumoniae. This method is based on a PCR system by which a segment of the 16S rRNA gene from all mycoplasmas of the M. mycoides cluster can be amplified. The PCR product is then analyzed by restriction enzyme cleavage for the identification of M. capripneumoniae DNA. This system has now been further evaluated with respect to specificity and diagnostic efficacy for the identification and direct detection of the organism in clinical material. Identification by restriction enzyme analysis of amplified DNA from mycoplasmas of the M. mycoides cluster was verified for 55 strains, among which were 15 strains of M. capripneumoniae. The PCR was applied to clinical samples from the nose, ear, pharynx, pleural fluid, and lung tissue containing M. capripneumoniae or other mycoplasmas. As expected, mycoplasmas belonging to the M. mycoides cluster could be detected by the PCR. Restriction enzyme analysis of the PCR products could then be applied for the identification of M. capripneumoniae. Clinical samples and cultures containing M. capripneumoniae were dried on filter paper, to try an easier sample transport method, and were tested by PCR. M. capripneumoniae DNA could be detected in the dried specimens, but the sensitivity of the PCR test was reduced.
Bolske, G; Mattsson, J G; Bascunana, C R; Bergstrom, K; Wesonga, H; Johansson, K E
Contagious bovine pleuropneumonia and contagious caprine pleuropneumonia are two severe respiratory infections of ruminants due to infection by Mycoplasma mycoides subsp. mycoides SC (MmmSC) and Mycoplasma capricolum subsp. capripneumoniae (Mccp), respectively. They are included in the OIE list of notifiable diseases. Here we describe the development of rapid, sensitive, and specific real-time PCR assays for the detection and quantification of MmmSC and Mccp DNA. MmmSC PCR primers were designed after whole genome comparisons between the published sequence of MmmSC strain type PG1(T) and the sequence of an M. mycoides subsp. mycoides large colony strain. For Mccp, previously published conventional PCR primers were applied. SYBR green was used as a detection agent for both assays. The assays specifically detected the targeted species in both cultures and clinical samples, and no cross-amplifications were obtained from either heterologous mycoplasmastrain cultures or European field samples. The sensitivity of these new assays was estimated at 3-80 colony forming units per reaction and 4-80fg of DNA, representing a 2-3log increase in sensitivity compared to established conventional PCR tests. These new real-time PCR assays will be invaluable for application in various fields such as direct detection in diagnostic laboratories. PMID:18678244
Lorenzon, Sophie; Manso-Silván, Lucía; Thiaucourt, François
To determine the growth pattern and in vitro susceptibility of Blastocystis hominis to metronidazole (MTZ), garlic, ginger, white cumin, and black pepper. Stool specimens were collected from 16 irritable bowel syndrome (IBS) and 10 controls between July-November 2010. Stool microscopy and culture for B. hominis was performed. Drug susceptibility assays was done using 0.01 and 0.1 mg/ml of MTZ, garlic, ginger, white cumin, and black pepper. Effect was assessed on B. hominis culture after 48 h. Stool DNA was extracted using stool DNA extraction kit (Qiagen) and polymerase chain reaction (PCR) done using subtype-specific sequence-tagged-site primers. B. hominis genotype 3 and coinfection of 1 and 3 tended to grow well in culture compared to isolated type 1 infection. Exposed to MTZ at a concentration of 0.01 mg/ml, 38% (6/16) B. hominis from IBS did not grow in culture compared to 100% (10/10) of B. hominis from control (p = 0.001). When they were exposed to MTZ at 0.1 mg/ml, 56% (9/16) B. hominis from IBS did not grow in cultures compared to 100% (10/10) from control (p = 0.01). Forty-four percent (7/16) B. hominis from IBS did not grow in culture compared to 100% (10/10) B. hominis from control when exposed to garlic at a concentration of 0.01 mg/ml (p = 0.003) and following exposure to garlic at 0.1 mg/ml, 38% (6/16) B. hominis from IBS did not grow in cultures compared to 100% (10/10) from control (p = 0.001). B. hominis isolates from IBS had a cell count of 6,625 at a MTZ concentration of 0.01 mg/ml that reduced to 1,250 as MTZ concentration was increased to 0.1 mg/ml (p = 0.08). B. hominis from IBS with a mean cell count of 3 × 10(5) at baseline decreased to 1 × 10(4) when exposed to garlic at 0.01 mg/ml (p < 0.001) and to 1 × 10(3) (p < 0.001) when garlic was 0.1 mg/ml. B. hominis from IBS cell count decreased to 1 × 10(5) when exposed to white cumin at 0.01 mg/ml (p = 0.01) and to 1 × 10(5) (p < 0.001) when white cumin was 0.1 mg/ml. Exposed to black pepper at 0.1 mg/ml, cell count of B. hominis from IBS decreased to 1 × 10(5) (p = 0.01). B. hominis from IBS decreased to 1.3 × 10(5) exposed to ginger at 0.01 mg/ml (p = 0.001). B. hominis isolates were mostly genotypes 3, type 1 and 3 coinfection, and non-typeable B. hominis isolates. B. hominis isolates from IBS mostly genotype 1 demonstrated an increased sensitivity to garlic at 0.01 mg/ml with a B. hominis cell count of 3,714 compared to 6,142 when exposed to 0.01 mg/ml of MTZ. However, this sensitivity did not increase as garlic concentration was increased to 0.1 mg/ml, for B. hominis cell count was 6,000 compared to 1,428 as MTZ was increased to 0.1 mg/ml. PMID:21431384
Authors present the first laboratory and epidemiological results which reveal the circulation in a population of protozoan Blastocystis hominis and its implication in the determinations of some gastrointestinal troubles, with fever, diarrhea and constipation, intense intestinal meteorism, associated with abdominal pain and cramps. Out of the 3106 investigated patients, 9.7% presented B. hominis as a unique etiologic agent, with an increased prevalence in adults (74.3%) and women (65.3%). Blastocystis infection with clinical manifestations or its asymptomatic form is included among emergent diseases. PMID:16607843
We found 4 species of mosquitoes bearing eggs of the human botfly, Dermatobia hominis, in the Reserva Municipal de Trabiju, Pindamonhangaba, Săo Paulo, Brazil. The mosquitoes were simultaneously collected in landing-biting catches by 2 collectors. From a total of 6,902 specimens collected from January through April 2010, the 15 females carrying D. hominis eggs belonged to Aedes scapularis, Limatus durhamii, Onirion personatum, and Wyeomyia confusa. The first 3 species are new reports of phoresy among mosquitoes and the human botfly. PMID:22894123
Marchi, Marco Jacometto; Pereira, Petra Assis; de Menezes, Regiane Maria Tironi; Tubaki, Rosa Maria
Mycoplasma penetrans, a rare bacterium so far only found in HIV-infected persons, was isolated in the blood and throat of a non-HIV-infected patient with primary antiphospholipid syndrome (whose etiology and pathogenesis are unknown). PMID:10081687
Yáńez, A; Cedillo, L; Neyrolles, O; Alonso, E; Prévost, M C; Rojas, J; Watson, H L; Blanchard, A; Cassell, G H
Mycoplasma penetrans, a rare bacterium so far only found in HIV-infected persons, was isolated in the blood and throat of a non-HIV-infected patient with primary antiphospholipid syndrome (whose etiology and pathogenesis are unknown).
Yanez, A.; Cedillo, L.; Neyrolles, O.; Alonso, E.; Prevost, M. C.; Rojas, J.; Watson, H. L.; Blanchard, A.; Cassell, G. H.
The sequenced genomes of the two closely related bacteria Mycoplasma genitalium and Mycoplasma pneumoniae were compared with emphasis on genome organization and coding capacity. All the 470 proposed open reading frames (ORFs) of the smaller M.genitalium genome (580 kb) were contained in the larger genome (816 kb) of M.pneumoniae. There were some discrepancies in annotation, but inspection of the DNA
Ralf Himmelreich; Helga Plagens; Helmut Hilbert; Berta Reiner; Richard Herrmann
The genital mycoplasmas represent a complex and unique group of microorganisms that have been associated with a wide array of infectious diseases in adults and infants. The lack of conclusive knowledge regarding the pathogenic potential of Mycoplasma and Ureaplasma spp. in many conditions is due to a general unfamiliarity of physicians and microbiology laboratories with their fastidious growth requirements, leading to difficulty in their detection; their high prevalence in healthy persons; the poor design of research studies attempting to base association with disease on the mere presence of the organisms in the lower urogenital tract; the failure to consider multifactorial aspects of diseases; and considering these genital mycoplasmas only as a last resort. The situation is now changing because of a greater appreciation of the genital mycoplasmas as perinatal pathogens and improvements in laboratory detection, particularly with regard to the development of powerful molecular nucleic acid amplification tests. This review summarizes the epidemiology of genital mycoplasmas as causes of neonatal infections and premature birth; evidence linking ureaplasmas with bronchopulmonary dysplasia; recent changes in the taxonomy of the genus Ureaplasma; the neonatal host response to mycoplasma and ureaplasma infections; advances in laboratory detection, including molecular methods; and therapeutic considerations for treatment of systemic diseases.
Waites, Ken B.; Katz, Brenda; Schelonka, Robert L.
M. hyorhinis is considered one of the etiological agents of arthritis in sucking pigs, but recently as seen, some strains can produce pneumonia that could not be distinguished from the mycoplasmosis caused by M. hyopneumoniae. The study was conducted to research the presence of Mycoplasma hyorhinis (M. hyorhinis ) in different regions of the country from exudates of pig lungs with typical EP lesions. Exudates from 280 pig lungs with typical EP lesions were studied using molecular techniques such as PCR, real time PCR and amplification of the 16S-23S rRNA. It was detected that the 66% of the samples studied resulted positive to M. hyorhinis, and the presence of this species was detected in all the provinces. Amplification and studies on the intergenic region 16S-23S of M. hyorhinis rRNA demonstrated the existing variability among strains of a same species. This study is the first report on M. hyorhinis detection in Cuba.
Lobo, Evelyn; Poveda, Carlos; Gupta, Rakesh; Suarez, Alejandro; Hernandez, Yenney; Ramirez, Ana; Poveda, Jose B.
Mycoplasma pneumoniae, a common human respiratory pathogen, has been studied experimentally for years using intranasal inoculation of the golden Sytrian hamster. Because of recent evidence outlining the role in pulmonary immune development of particle size and depth of mycoplasma...
MICs of eight antibiotics were detected with 40 Chinese Mycoplasma pneumoniae isolates. Thirty-eight isolates (95%) were macrolide resistant. Each macrolide-resistant isolate harbored an A2063G or A2064G point mutation in the 23S rRNA gene. All 40 isolates (100%) were type I strains, but they might have originated from different clones.
BACKGROUND: Several methods have been reported for strain typing of Mycoplasma genitalium. The value of these methods has never been comparatively assessed. The aims of this study were: 1) to identify new potential genetic markers based on an analysis of short tandem repeat (STR) sequences in the published M. genitalium genome sequence; 2) to apply previously and newly identified markers
Liang Ma; Stephanie Taylor; Jřrgen S Jensen; Leann Myers; Rebecca Lillis; David H Martin
A new molecular assay, based on a rapid DNA extraction protocol, PCR, and hybridization to a specific probe in a nonradioactive microwell plate format was used to detect Mycoplasma pneumoniae in bronchoalveolar fluid specimens. The sensitivity of the assay was determined to be 10 to 100 organisms with M. pneumoniae reference strains. Specificity testing with different bacteria capable of producing
HARALD H. KESSLER; DEBORAH E. DODGE; KAREN PIERER; KAREN K. Y. YOUNG; YANHONG LIAO; BRIGITTE I. SANTNER; ERNST EBER; MARIA G. ROEGER; DORIS STUENZNER; BIRGIT SIXL-VOIGT; EGON MARTH
Three experimental strains of breeder chickens were accidentally exposed to Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS), presumably from a newly introduced group of leghorn-type pullets. The experimental strains subsequently became infected and were diagnosed positive for MG and MS by the serum plate agglutination (SPA) test and confirmed by the hemagglutination inhibition (HI) test and the polymerase chain reaction (PCR) of tracheal swabs. Treatment with 10 mg/kg enrofloxacin via drinking water for 14 days was elected. Before and after initiation of treatment, MG and MS were monitored for changes by SPA, HI, PCR, and culture, with sampling intervals ranging from 1 wk to 7 wk. MG and MS SPA, HI, PCR, and culture were performed at each sampling period, with the exception of weeks 1.0 and 6.5. Week 1.0 included SPA and His for MG and MS. Week 6.5 included PCR and culture for MG and MS. The MG and MS SPA results were positive throughout the 29-wk trial period. MG HI titers declined until the last sampling, whereas the MS HI titers did not decline significantly. PCR for MG yielded only one positive result, which occurred before treatment. MS PCR remained positive throughout the trial period. MG was never isolated from any sample; however, one MS organism was isolated during treatment. The treatment regimen was effective for MG on the basis of PCR results. Treatment with enrofloxacin did not eliminate SPA reactions during the 29-wk trial period. MG HI titers remained in the suspicious range throughout the remainder of the trial period. Four weeks after the treatment ended, MG HIs were reduced by approximately 40%, with MS HIs remaining high throughout the 29-wk period. PCR appeared to be a sensitive and specific test on the basis of correlation with HIs. On the basis of the isolation of MS during treatment and continued subsequent PCR positive reactions, the treatment for MS with enrofloxacin was not as efficacious as for MG. PMID:11417841
Stanley, W A; Hofacre, C L; Speksnijder, G; Kleven, S H; Aggrey, S E
The present report describes a method for establishment of colonies of Blastocystis hominis from single cells in soft agar. The percentage of colony-forming efficiency (% CFE = number of colonies grown \\/ number of\\u000a cells inoculated × 100) for the cultures was greatly improved by the addition of sodium thioglycollate. Five human Blastocystis isolates chosen for this study showed no
S. W. Tan; M. Singh; K. T. Thong; L. C. Ho; K. T. Moe; X. Q. Chen; G. C. Ng; E. H. Yap
Summary Morphological changes in the central vacuole during the growth in in vitro culture ofBlastocystis hominis were investigated by light and electron microscopy. Most cells in log phase and an early stationary phase showed a positive staining reaction in the central vacuole with PAS or Sudan black B stain, whereas cells in late stationary phase showed few positive reactions. Electron
Resistance of Sarcoptes scabiei to various topical therapies has been described, but clinical assessment of treatment failure is problematic and in-vitro assays are generally not available. We describe a simple in-vitro analysis used to evaluate the relative efficacy of a range of topical, oral, and herbal treatments available in Australia for the treatment of scabies. S. scabiei var. hominis mites
Recently, several investigators have reported large-bowel diarrhea in cats associated with intestinal trichomonad parasites. These reports have presumptively identified the flagellates as Pentatrichomonas hominis , an organism putatively capable of infecting the intestinal tracts of a number of mammalian hosts, including cats, dogs, and man. The purpose of the present study was to determine the identity of this recently recognized
Michael G. Levy; Jody L. Gookin; Matthew Poore; Adam J. Birkenheuer; Michael J. Dykstra; R. Wayne Litaker
...2009-01-01 false Detection of mycoplasma contamination... VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS...Procedures Â§ 113.28 Detection of mycoplasma contamination...conducting the mycoplasma detection test. (1...of a cell line or a sample of primary...
...2010-01-01 false Detection of mycoplasma contamination... VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS...Procedures Â§ 113.28 Detection of mycoplasma contamination...conducting the mycoplasma detection test. (1...of a cell line or a sample of primary...
Mycoplasma synoviae is a Gram positive bacteria lacking of cell wall that affects chickens and turkeys causing infection in the upper respiratory tract and in some cases arthritis, with economical impact to broiler breeders. Treatment and prevention of avian synovitis depend on knowledge of the infectious process. Secreted or surface-exposed proteins play a critical role in disease because they often mediate interactions between host and pathogen. In the present work, we sought to identify possible M. synoviae secreted proteins by cultivating the bacteria in a modified protein-free Frey medium. Using this approach, we were able to detect in the cell-free fraction a number of proteins that have been shown in other organisms to be secreted, suggesting that they may also be secreted by M. synoviae.
The variable surface antigens (Vsa) of the murine respiratory pathogen Mycoplasma pulmonis are associated with the virulence of the microorganism in the lung. In strain UAB CT, the antigens consist of an N-terminal region that is combined with one of seven different C-terminal variable regions comprised of tandem repeats. M. pulmonis producing a VsaA protein with about 40 tandem repeats
Mycoplasma pulmonis is a wall-less eubacterium belonging to the Mollicutes (trivial name, myco- plasmas) and responsible for murine respiratory diseases. The genome of strain UAB CTIP is composed of a single circular 963 879 bp chromosome with a G + C content of 26.6 mol%, i.e. the lowest reported among bacteria, Ureaplasma urealyticum apart. This genome contains 782 putative coding
Isabelle Chambaud; Roland Heilig; Stéphane Ferris; Valérie Barbe; Delphine Samson; Frédérique Galisson; I van Moszer; Kevin Dybvig; Henri Wróblewski; Alain Viari; Eduardo P. C. Rocha; Alain Blanchard
A species-specific 760-base pair (bp) BamHI to EcoRI DNA fragment (fMG-2) was isolated from a Mycoplasma gallisepticum (MG) genomic library constructed in plasmid pUC8. Based on the DNA sequence data of fMG-2, a pair of 25 base primers, designated amplification (Amp) left (L) and right (R) primers, was synthesized. When used in the polymerase chain reaction (PCR), the Amp L and R primers directed amplification of DNA of 16 MG strains yielding an expected 732-bp product, but did not amplify DNA of Escherichia coli, calf thymus, lambda phage, pUC8 plasmid, or 16 other species of avian mycoplasmas. As low as 10(-6) picogram of MG DNA, a fraction of the total chromosomal content of one cell, was detected following amplification by PCR. PCR amplification products were visualized by either ethidium bromide/ultraviolet exposure or hybridization with a 481-bp probe (fMG-3) prepared from the central region of fMG-2. PMID:2029263
Nascimento, E R; Yamamoto, R; Herrick, K R; Tait, R C
Several Mycoplasma species have had their genome completely sequenced, including four strains of the swine pathogen Mycoplasma hyopneumoniae. Nevertheless, little is known about the nucleotide sequences that control transcriptional initiation in these microorganisms. Therefore, with the objective of investigating the promoter sequences of M. hyopneumoniae, 23 transcriptional start sites (TSSs) of distinct genes were mapped. A pattern that resembles the ?70 promoter ?10 element was found upstream of the TSSs. However, no ?35 element was distinguished. Instead, an AT-rich periodic signal was identified. About half of the experimentally defined promoters contained the motif 5?-TRTGn-3?, which was identical to the ?16 element usually found in Gram-positive bacteria. The defined promoters were utilized to build position-specific scoring matrices in order to scan putative promoters upstream of all coding sequences (CDSs) in the M. hyopneumoniae genome. Two hundred and one signals were found associated with 169 CDSs. Most of these sequences were located within 100 nucleotides of the start codons. This study has shown that the number of promoter-like sequences in the M. hyopneumoniae genome is more frequent than expected by chance, indicating that most of the sequences detected are probably biologically functional.
Weber, Shana de Souto; Sant'Anna, Fernando Hayashi; Schrank, Irene Silveira
Strains of Mycoplasma mycoides subspecies capri (Mmc) are frequently isolated from goats with contagious agalactia, but they can also be recovered from herds that have shown no clinical signs of mycoplasmosis for several years. The present study was conducted in order to explore the potential genetic and antigenic differences existing between an Mmc strain isolated from an outbreak (septicaemic) and a strain isolated from the ear canal of a goat belonging to a herd with no recent episode of mycoplasmosis (carriage strain). The genomes of the two strains, compared by suppression subtractive hybridization, were shown to be poorly divergent. The two strains were inoculated into goats to produce specific antisera, but both induced fatal mycoplasmosis. These results indicate that septicaemic and carriage strains cannot be distinguished by their genetic background or by their pathogenic capacity under experimental conditions. PMID:20708197
Tardy, F; Maigre, L; Tricot, A; Poumarat, F; Nguyen, L; Le Grand, D
Mycoplasma mastitis is an emerging mastitis pathogen. Herd prevalence has increased over the past decade, and this increase parallels the increase in average dairy herd size. It has been documented that the importation of cattle into a herd can result in new cases of Mycoplasma disease in general and Mycoplasma mastitis specifically. Thus, expanding herds are likely to have a greater incidence of this disease. Transmission of the agent can result from either contact with diseased animals or with colonized or asymptomatically infected cattle. Initial transmission might occur via nose-to-nose contact and result in an outbreak of Mycoplasma mastitis, or it might occur during the milking time. This would suggest that new, incoming animals should be quarantined before being comingled with original herd animals. Quarantining does not seem to be a biosecurity strategy often practiced in control of Mycoplasma mastitis and may not be warranted in herds with excellent milking time hygiene practices. The ability to monitor for the incipient stages of an outbreak, often done through bulk tank milk culturing, is recommended. PMID:22664205
Mycoplasma pneumoniae is a significant cause of community-acquired pneumonia, which is often empirically treated with macrolides or azalides such as erythromycin or azithromycin. Recent studies have discovered the existence of macrolide-resistant strains within the population that have been mapped to mutations within the domain V region of the 23S rRNA gene. Currently, identification of these resistant strains relies on time-
Bernard J. Wolff; W. Lanier Thacker; Stephanie B. Schwartz; Jonas M. Winchell
Pheasants and partridges with signs of upper respiratory disease were cultured for mycoplasmas and were also examined for Mycoplasma gallisepticum and Mycoplasma synoviae using commercial polymerase chain reaction (PCR) kits. Sixty-two incidents of disease were investigated in pheasants and 12 in partridges. M. gallisepticum was detected by culture in only four and three incidents in pheasants and partridges, respectively, but with PCR a further 15 M. gallisepticum-positive incidents were detected in pheasants and another five in partridges. Several fast-growing Mycoplasma species, in particular Mycoplasma glycophilum, Mycoplasma gallinaceum and Mycoplasma pullorum, were isolated frequently and were thought to be impeding the isolation of M. gallisepticum by outgrowing it. Samples yielding M. gallisepticum isolates contained significantly fewer "contaminating" species and were exclusively from specimens submitted as whole heads rather than as swabs or as cultures from other laboratories. M. synoviae was not isolated and was detected in only one specimen by PCR. PMID:19184924
Mycoplasma infections are of great concern in avian medicine, because they cause economic losses in commercial poultry production. A multiplex polymerase chain reaction (PCR) was optimized to simultaneously detect four pathogenic species of avian mycoplasmas. Four sets of oligonucleotide primers specific for Mycoplasma gallisepticum (MG), M. synoviae (MS), M. meleagridis (MM) and M. iowae (MI) were used in the test. By using agarose gel electrophoreses for detection of the PCR-amplified DNA products, the sensitivity of detection was between 1 pg for MG, 1 pg for MS, 100 fg for MM and 100 pg for MI after 35 cycles of PCR. Similar sensitivity of these primers was achieved with broth cultures of these four organisms. PMID:9232620
Mycoplasma penetrans is a newly identified species of the genus Mycoplasma. It was first isolated from a urine sample from a human immunodeficiency virus (HIV)-infected patient. M. penetrans changes its surface antigen profile with high frequency. The changes originate from ON?OFF phase variations of the P35 family of surface membrane lipoproteins. The P35 family lipoproteins are major antigens recognized by the human immune system during M. penetrans infection and are encoded by the mpl genes. Phase variations of P35 family lipoproteins occur at the transcriptional level of mpl genes; however, the precise genetic mechanisms are unknown. In this study, the molecular mechanisms of surface antigen profile change in M. penetrans were investigated. The focus was on the 46-kDa protein that is present in M. penetrans strain HF-2 but not in the type strain, GTU. The 46-kDa protein was the product of a previously reported mpl gene, pepIMP13, with an amino-terminal sequence identical to that of the P35 family lipoproteins. Nucleotide sequencing analysis of the pepIMP13 gene region revealed that the promoter-containing 135-bp DNA of this gene had the structure of an invertible element that functioned as a switch for gene expression. In addition, all of the mpl genes of M. penetrans HF-2 were identified using the whole-genome sequence data that has recently become available for this bacterium. There are at least 38 mpl genes in the M. penetrans HF-2 genome. Interestingly, most of these mpl genes possess invertible promoter-like sequences, similar to those of the pepIMP13 gene promoter. A model for the generation of surface antigenic variation by multiple promoter inversions is proposed.
Actinobacillus species are usually not considered as being human pathogens apart from A. actinomycetemcomitans. However, single cases of human meningitis, septicemia, and empyema caused by Actinobacillus lignieresii have been reported in the literature. This is the first reported case of Actinobacillus hominis giving rise to pleural-empyema in a patient with carcinoma of the lung. The function of peripheral blood neutrophils, serum opsonic activity and specific precipitating antibodies were investigated. Neutrophils from the patient exhibited an enhanced oxidative burst response measured by chemiluminescence assay. Furthermore, the opsonic activity of the serum from the patient was higher than that of a healthy control person. Several precipitating antibodies to various antigens of Actinobacillus hominis were demonstrated in the serum of the patient by crossed immunoelectrophoresis. PMID:3196473
A non-axenic and an axenic isolate ofBlastocystis hominis have been induced to form cysts in vitro using an encystation medium. The morphology of the parasite at different time points was observed by scanning electron microscopy. In day-2 cultures the cysts were spherical and had a non-uniform, coarse outer surface around the body. A deep, pore-like opening was seen in some
K. Suresh; J. Howe; G. C. Ng; L. C. Ho; N. P. Ramachandran; A. K. Loh; E. H. Yap; M. Singh
Little is known about the emergence and spread of virulent subtypes of Cryptosporidium hominis, the predominant species responsible for human cryptosporidiosis. We conducted sequence analyses of 32 genetic loci of 53 C. hominis specimens isolated from a longitudinally followed cohort of children living in a small community. We identified by linkage disequilibrium and recombination analyses only limited genetic recombination, which occurred exclusively within the 60-kDa glycoprotein gene subtype IbA10G2, a predominant subtype for outbreaks in industrialized nations and a virulent subtype in the study community. Intensive transmission of virulent subtype IbA10G2 in the study area might have resulted in genetic recombination with other subtypes. Moreover, we identified selection for IbA10G2 at a 129-kb region around the 60-kDa glycoprotein gene in chromosome 6. These findings improve our understanding of the origin and evolution of C. hominis subtypes and the spread of virulent subtypes. PMID:24050210
In this study, reverse transcriptase PCR was employed to construct a transcriptional profile of Mycoplasma pneumoniae lipoprotein genes contained in six multigene families. Most genes were found to be expressed. Many truncated lipoprotein genes were expressed, often polycistronically with other truncated genes, indicating that these genes may still be functional.
Background and Purpose: Previous studies have suggested certain infections as potential risk factors for stroke. Chlamydia pneumoniae, an atypical respiratory pathogen, has been linked to atherosclerotic vascular diseases. Mycoplasma pneumoniae, another atypical respiratory micro-organism, can rarely cause stroke. We investigated whether serological markers of M. pneumoniae infection were associated with acute stroke or transient ischaemic attack (TIA) in elderly patients.
Joseph Ngeh; Sandeep Gupta; Colin Goodbourn; Geraldine McElligott
The effects of length of incubation and urine osmolality on the survival of feline mycoplasmas and ureaplasmas and representative gram-positive and gram-negative bacteria in synthetic urine which approximated the osmolality of normal cat urine were investigated. Both Escherichia coli and Staphylococcus aureus withstood the effects of increasing osmotic pressure. In the most concentrated urine, significant decreases (P less than 0.001) in CFU were observed for E. coli at exposure times of 30 min and longer. S. aureus was not affected by longer exposure or increased osmotic strength. Both Mycoplasma felis and Mycoplasma gateae were affected adversely by longer exposure times and high osmotic strength (P less than 0.001). A Ureaplasma sp. was not adversely affected except at very high (greater than or equal to 2,980 mosM) osmotic strengths or after prolonged incubation (120 min) at relatively high (1,976 mosM) osmotic strengths (P less than 0.001). The failure of both M. felis and M. gateae to survive under osmotic conditions present in normal feline urine suggests that it is unlikely that these mycoplasmas are involved in urinary disorders in cats.
ology, rotational motility, and infection by a tailed bacteriophage, characteristics unknown among the 'classical' mycoplasmas, with which it shows no serological relation ship. It has therefore been placed in a new genus, Spiroplasma, as a new species, S. eitri (Sagl io et al., 1973). This organism is now well characterized (Cole et al., 1973; Bowyer and Calavan, 1974). Pathogenicity of
The effects of length of incubation and urine osmolality on the survival of feline mycoplasmas and ureaplasmas and representative gram-positive and gram-negative bacteria in synthetic urine which approximated the osmolality of normal cat urine were investigated. Both Escherichia coli and Staphylococcus aureus withstood the effects of increasing osmotic pressure. In the most concentrated urine, significant decreases (P less than 0.001) in CFU were observed for E. coli at exposure times of 30 min and longer. S. aureus was not affected by longer exposure or increased osmotic strength. Both Mycoplasma felis and Mycoplasma gateae were affected adversely by longer exposure times and high osmotic strength (P less than 0.001). A Ureaplasma sp. was not adversely affected except at very high (greater than or equal to 2,980 mosM) osmotic strengths or after prolonged incubation (120 min) at relatively high (1,976 mosM) osmotic strengths (P less than 0.001). The failure of both M. felis and M. gateae to survive under osmotic conditions present in normal feline urine suggests that it is unlikely that these mycoplasmas are involved in urinary disorders in cats. PMID:2056047
Mycoplasma growth factors in bovine serum fraction were separated by Sephadex G150 column chromatography and density ultracentrifugation. The major growth factor of bovine serum fraction eluted from the Sephadex column in the void volume. Its growth-supporting activity was greatly enhanced by the presence of bovine serum albumin in the mycoplasma culture media. Other investigators had previously identified the major growth factor in serum as an alpha-lipoprotein. Although density ultracentrifugation revealed the presence of traces of a high-density lipoprotein in bovine serum fraction, another, less dense component, isolated by ultracentrifugation (component 3) and containing cholesterol, cholesteryl esters, free fatty acids, triglycerides, and protein, but no lipoprotein, exhibited considerably more growth-supporting activity than did the high-density lipoprotein, thus indicating that at least two mycoplasma species do not require intact serum lipoprotein for growth. Both the high-density lipoprotein and component 3 exhibited maximum activity only in the presence of bovine serum albumin. A chloroform extract containing component 3 lipids combined with bovine serum albumin to form an effective, partially defined, less complex substitute for serum in mycoplasma culture media. Images
Farm rabbits are affected by respiratory and reproductive disorders that compromise their health and the productivity of the farm. In two earlier papers we highlighted the role of mycoplasmas in these processes and the present article evaluates the usefulness of autovaccines to control mycoplasmosis. On a commercial rabbitry with 800 females, 428 kits were vaccinated, leaving 3,622 as controls. A
Mycoplasma genitalium is an important cause of sexually transmitted infections that is gaining recognition and is an independent cause of acute and chronic nongonococcal urethritis in men. M. genitalium has been implicated as a possible causative factor in reactive arthritis. We report a case of reactive arthritis complicating M. genitalium urethritis in an HLA-B27-positive patient. PMID:24034901
Mycoplasma pneumoniae infections have extrapulmonary complications that involve the nervous system. The neurologic manifestations are diverse. Although the prognosis is usually favorable, the patients can undergo severe permanent sequelae. We present a young female adult with acute meningoencephalitis as a complication of a lower respiratory infection, which followed a benign course without neurologic sequelae. PMID:16193713
Del Castillo, Marcelo; D'Giano, Carlos; Goicoechea, María Teresa; Morello, Fernando; Salsamendi, Paz; Mora, Andrea
We report significant failure rates (28%, 95% confi- dence interval 15%-45%) after administering 1 g azithromycin to men with Mycoplasma genitalium-positive nongonococcal urethritis. In vitro evidence supported reduced susceptibility of M. genitalium to macrolides. Moxifloxacin administration resulted in rapid symptom res- olution and eradication of infection in all cases. These find- ings have implications for management of urethritis.
Catriona S. Bradshaw; Jorgen S. Jensen; Sepehr N. Tabrizi; Timothy R. H. Read; Suzanne M. Garland; Carol A. Hopkins; Lorna M. Moss; Christopher K. Fairley
"Candidatus Mycoplasma turicensis" infects felids. The pathogenesis of "Candidatus M. turicensis" chronic infection is poorly understood. The goals of the present study were to (1) induce reactivation of the infection in chronic carrier cats by attempted immunosuppression, (2) identify potential tissue sequestration using real-time TaqMan® PCR and (3) monitor the humoral immune response by DnaK enzyme-linked immunosorbent assay (ELISA). Ten specified pathogen-free cats that had ostensibly recovered from experimental "Candidatus M. turicensis" infection were used: five cats (group 1) received high dose methylprednisolone (attempted immunosuppression), while five cats served as untreated controls (group 2). Besides weekly blood samples, tissue samples were collected from bone marrow, kidney, liver and salivary glands at selected time points. The cats in group 1 had significantly lower lymphocyte counts and higher blood glucose levels after methylprednisolone administration than the controls. After methylprednisolone administration one blood and three tissue samples from cats in group 1 tested PCR-positive; before the administration, only one sample was positive. All other samples tested PCR-negative. All cats stayed seropositive; the antibody levels of the cats in group 1 showed a significant transient decrease after methylprednisolone administration. This is the first study to report the presence of "Candidatus M. turicensis" in tissues of chronically infected cats and the persistence of anti-feline hemoplasma antibodies in the absence of detectable bacteremia. Methylprednisolone administration did not lead to a significant reactivation of the infection. Our results enhance the knowledge of "Candidatus M. turicensis" infection pathogenesis and are clinically relevant to the prognosis of hemoplasma-infected cats.
Fifteen mycoplasma-free chickens were contact exposed to five chickens that had been experimentally infected with one of three different strains (two field strains and one laboratory strain) of Mycoplasma synoviae (MS). Culture and polymerase chain reaction (PCR) were positive by 3 days postinoculation (PI) in the experimentally infected birds. Lateral transmission was found by 7-14 days postexposure. Positive serum plate agglutination (SPA) results were detected 3-4 wk after positive culture and/or PCR in individual birds. By 42 days PI, all the birds in the groups exposed to field strain K1858 or K3344 had become infected as determined by culture and PCR, whereas only half of the birds in the group exposed to laboratory strain WUV1853 had become infected. Because of the unanticipated lack of seroconversion to hemagglutination inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) in infected chickens, the study was extended. Each group was split into two groups of 10 birds each, one of which was vaccinated with a live B1/LaSota Newcastle disease (ND) vaccine virus to determine if a viral respiratory challenge might incite a stronger antibody response to the mycoplasma infection. All the birds were tested for seroconversion 14 and 21 days later. Of the birds vaccinated for ND, a slightly greater number were MS positive by SPA than the nonvaccinated birds. This effect was not present 21 days after vaccination, and there was no significant difference in the MS HI results from these groups, suggesting that the viral respiratory infection had little direct impact on seroconversion. The virulent field strain (K3344) elicited a stronger MS antibody response than the other strains. All results from the MS ELISA were negative in all groups through 9 wk. Positive results from PCR analysis correlated well with culture results, whereas serologic tests did not detect MS infection for several weeks. Monitoring programs solely dependent on seroconversion may be inadequate for diagnosis and control of mycoplasma infections. PMID:9645313
Ewing, M L; Cookson, K C; Phillips, R A; Turner, K R; Kleven, S H
Mycoplasma arthritidis causes a severe polyarthritis under natural conditions in rats and under experimental conditions in both rats and mice. Although the disease itself has been extensively studied, M. arthritidis virulence factors remain uncharacterized. Comparison of relative arthritogenicity of 20 strains of M. arthritidis revealed that the strains tended to fall into two groups, a highly arthritogenic group, inducing maximum arthritis scores of > or = 11 in rats, and a low-virulence group, inducing maximum scores of < 6. Chromosomal DNA from the more highly arthritogenic strains possessed sequences that hybridized by Southern analysis with a probe prepared from lysogenic M. arthritidis bacteriophage MAV1, while DNA from low-virulence strains did not. One of the low-virulence strains, 158, was experimentally lysogenized with MAV1. Lysogenized 158 showed a significant increase in arthritogenicity over nonlysogenized 158. These data suggest that MAV1 carries a factor that is important in pathogenesis of M. arthritidis-induced arthritis of rats.
Voelker, L L; Weaver, K E; Ehle, L J; Washburn, L R
Background Mycoplasma gallisepticum is a major poultry pathogen and causes severe economic loss to the poultry industry. In mycoplasmas lipoproteins are abundant on the membrane surface and play a critical role in interactions with the host, but tools for exploring their molecular biology are limited. Results In this study we examined whether the alkaline phosphatase gene (phoA ) from Escherichia coli could be used as a reporter in mycoplasmas. The promoter region from the gene for elongation factor Tu (ltuf) and the signal and acylation sequences from the vlhA 1.1 gene, both from Mycoplasma gallisepticum , together with the coding region of phoA , were assembled in the transposon-containing plasmid pISM2062.2 (pTAP) to enable expression of alkaline phosphatase (AP) as a recombinant lipoprotein. The transposon was used to transform M. gallisepticum strain S6. As a control, a plasmid containing a similar construct, but lacking the signal and acylation sequences, was also produced (pTP) and also introduced into M. gallisepticum . Using a colorimetric substrate for detection of alkaline phosphatase activity, it was possible to detect transformed M. gallisepticum . The level of transcription of phoA in organisms transformed with pTP was lower than in those transformed with pTAP, and alkaline phosphatase was not detected by immunoblotting or enzymatic assays in pTP transformants, eventhough alkaline phosphatase expression could be readily detected by both assays in pTAP transformants. Alkaline phosphatase was shown to be located in the hydrophobic fraction of transformed mycoplasmas following Triton X-114 partitioning and in the membrane fraction after differential fractionation. Trypsin proteolysis confirmed its surface exposure. The inclusion of the VlhA lipoprotein signal sequence in pTAP enabled translocation of PhoA and acylation of the amino terminal cysteine moiety, as confirmed by the effect of treatment with globomycin and radiolabelling studies with [14?C]palmitate. PhoA could be identified by mass-spectrometry after separation by two-dimensional electrophoresis. Conclusion This is the first study to express PhoA as a lipoprotein in mycoplasmas. The pTAP plasmid will facilitate investigations of lipoproteins and protein translocation across the cell membrane in mycoplasmas, and the ease of detection of these transformants makes this vector system suitable for the simultaneous screening and detection of cloned genes expressed as membrane proteins in mycoplasmas.
Mycoplasma pneumoniae is primarily a respiratory pathogen but may affect exhibit a diverse range of presentations from asymptomatic infection to life threatening conditions. Myocarditis of varying severity is an unusual complication. We report a 6 year old with mycoplasma myocarditis, a rare age for such a presentation, and who responded well to treatment with no sequelae. Serological testing for Mycoplasma pneumoniae should be part of the routine work-up for myocarditis.
Formosa, Gouder M; Bailey, M; Barbara, C; Muscat, C; Grech, V
This study compares the antimicrobial susceptibility over time between two groups of Mycoplasma gallisepticum (MG) isolates from the same geographical area. Minimum inhibitory concentration of 13 antimicrobials was determined against two groups of MG isolates from chickens. Group 1 strains (n=22) were isolated in 2004-2005 while group 2 strains (n=7) were isolated in 2007-2008. Minimum inhibitory concentration 50 for group 1 versus group 2 was 4/4, 0.5/0.5, ? 0.031/? 64, ? 0.031/2, ? 0.031/0.125, 1/0.5, 1/1, ? 0.031/? 0.031, ? 0.031/2, ? 0.031/2, 1/4, ? 0.031/0.062, and 0.062/2 ?g/ml against gentamicin, spectinomycin, erythromycin, tilmicosin, tylosin, florfenicol, thiamphenicol, tiamulin, ciprofloxacin, enrofloxacin, chlortetracycline, doxycycline, and oxytetracycline, respectively. There was a statistically significant increase in resistance of group 2 to erythromycin, tilmicosin, tylosin, ciprofloxacin, enrofloxacin, chlortetracycline, doxycycline, and oxytetracycline. This dramatic increase in resistance against 8 antimicrobials belonging to three different families of antimicrobials in a relatively short period of time appears to be rare and of concern. The cause of this increased resistance observed in group 2 of MG isolates was not determined and should be further investigated. Monitoring of MG field strain susceptibility is highly recommended to implement successful treatment and prophylaxis programs in endemic areas. PMID:21382675
Mycoplasmas are commensals and pathogens of different avian species, especially poultry and passeriforms. The role of mycoplasmas in raptors has not yet been completely determined, and especially not the possibility of vertical transmission. Therefore 424 raptor eggs were examined for the occurrence of mycoplasmas using culture, and 155 of these eggs with a Mycoplasma genus-specific polymerase chain reaction (PCR) assay. This PCR was tested for its sensitivity and specificity, especially for use in a bird population of unknown mycoplasma status (prevalence and species). The size of the amplified PCR product was large (1013 base pairs) to enable use of the product for species differentiation by sequencing. Culture and PCR yielded only one positive result, in an egg of a Northern Goshawk (Accipiter gentilis). The isolate was identified as Mycoplasma lipofaciens using an immunobinding assay, as well as by sequencing part of its 16S rRNA gene. PMID:17479375
Lierz, M; Hagen, N; Harcourt-Brown, N; Hernandez-Divers, S J; Lüschow, D; Hafez, H M
Some species of the genus Mycoplasma code for the arginine deiminase pathway (ADI), which enables these bacteria to produce ATP from arginine by the successive reaction of three enzymes: arginine deiminase (ArcA), ornithine carbamoyltransferase (ArcB), and carbamate kinase (ArcC). It so far appears that independently isolated strains of Mycoplasma pneumoniae encode an almost identical truncated version of the ADI pathway in which the proteins ArcA and ArcB have lost their original enzymatic activities due to the deletion of significant regions of these proteins. To study the consequences of a functional ADI pathway, M. pneumoniae M129 was successfully transformed with the cloned functional arcA, arcB, and arcC genes from Mycoplasma fermentans. Enzymatic tests showed that while the M. pneumoniae ArcAB and ArcABC transformants possess functional arginine deiminase, ornithine carbamoyltransferase, and carbamate kinase, they were unable to grow on arginine as the sole energy source. Nevertheless, infection of a lung epithelial cell line, A549, with the M. pneumoniae transformants showed that almost 100% of the infected host cells were nonviable, while most of the lung cells infected with nontransformed M. pneumoniae were viable under the same experimental conditions. PMID:23897620
Rechnitzer, Hagai; Rottem, Shlomo; Herrmann, Richard
A total of 1700 male food handlers, above 20 years of age who came for health clearance certificate were the subjects of the present study. Health assessment questionnaire was filled out on each person including dietary habits, water supply, history of diarrhoeal disease. Clinical examination and stool samples collection in 3 alternative days were performed. The food handlers were divided into symptomatic (700) and saymptomatic (1000). Different concentration methods as well as test tube culture for Strongyloides larvae were done. Samples were preserved in PVA, trichrome stained slides were examined for protozoal parasites. Nineteen percent had intestinal parasites, G. lamblia, E. histolytica, A.. lumbricoides, S. mansoni, A. duodenale, T. trichura, H. nana, St. stercoralis, E. vermicularis and mixed infection & non-pathogenic; E. coli, I. Butschlii, C. mesnilli, E. nana, T. hominis and mixed infection. Blastocystis hominis was recovered from stools of 8.5% of symptomatic and 4% of asymptomatic. 2.4% symptomatic and 2% asymptomatic had B. hominis significant infection. B hominis was considered significant if > 5 organisms per HPF was counted. Significant infection was higher among symptomatic than asymptomatic persons with detectable faecal leucocytes especially eosinophils. The authors recommended that physicians as well as diagnostic parasitologists should be aware of the potential clinical significance of B. hominis especially, when present alone in significant number, otherwise positive cases must be considered as carriers and followed up for any ill effects. PMID:9257986
Sadek, Y; el-Fakahany, A F; Lashin, A H; el-Salam, F A
Mycoplasma bovis pneumonia in cattle has been epidemic in China since 2008. To investigate M. bovis pathogenesis, we completed genome sequencing of strain HB0801 isolated from a lesioned bovine lung from Hubei, China. The genomic plasticity was determined by comparing HB0801 with M. bovis strain ATCC® 25523™/PG45 from cow mastitis milk, Chinese strain Hubei-1 from lesioned lung tissue, and 16 other Mycoplasmas species. Compared to PG45, the genome size of HB0801 was reduced by 11.7 kb. Furthermore, a large chromosome inversion (580 kb) was confirmed in all Chinese isolates including HB0801, HB1007, a strain from cow mastitis milk, and Hubei-1. In addition, the variable surface lipoproteins (vsp) gene cluster existed in HB0801, but contained less than half of the genes, and had poor identity to that in PG45, but they had conserved structures. Further inter-strain comparisons revealed other mechanisms of gene acquisition and loss in HB0801 that primarily involved insertion sequence (IS) elements, integrative conjugative element, restriction and modification systems, and some lipoproteins and transmembrane proteins. Subsequently, PG45 and HB0801 virulence in cattle was compared. Results indicated that both strains were pathogenic to cattle. The scores of gross pathological assessment for the control group, and the PG45- and HB0801-infected groups were 3, 13 and 9, respectively. Meanwhile the scores of lung lesion for these three groups were 36, 70, and 69, respectively. In addition, immunohistochemistry detection demonstrated that both strains were similarly distributed in lungs and lymph nodes. Although PG45 showed slightly higher virulence in calves than HB0801, there was no statistical difference between the strains (P>0.05). Compared to Hubei-1, a total of 122 SNP loci were disclosed in HB0801. In conclusion, although genomic plasticity was thought to be an evolutionary advantage, it did not apparently affect virulence of M. bovis strains in cattle.
We explored the genetic basis for intraspecific variation in mycoplasmal sialidase activity that correlates with virulence, and its potentially advantageous linkage to nutrient catabolism. Polymorphism in N-acetylneuraminate scavenging and degradation genes (sialidase, N-acetylneuraminate lyase, N-acetylmannosamine kinase, N-acetylmannosamine-6-phosphate epimerase, N-acetylglucosamine-6-phosphate deacetylase, and glucosamine-6-phosphate deaminase) was evident among eight strains of the avian pathogen Mycoplasma synoviae. Most differences were single nucleotide polymorphisms, ranging from 0.34 ± 0.04 substitutions per 100 bp for N-acetylmannosamine kinase to 0.65 ± 0.03 for the single-copy sialidase gene nanI. Missense mutations were twice as common as silent mutations in nanI; 26% resulted in amino acids dissimilar to consensus; and there was a 12-base deletion near the nanI promoter in strain WVU1853T, supporting a complex genetic basis for differences in sialidase activity. Two strains had identical frameshifts in the N-acetylneuraminate lyase gene nanA, resulting in nonsense mutations, and both had downstream deletions in nanA. Such genetic lesions uncouple extracellular liberation of sialic acid from generation of fructose-6-phosphate and pyruvate via intracellular N-acetylneuraminate degradation. Retention of nanI by such strains, but not others in the M. synoviae phylogenetic cluster, is evidence that sialidase has an important non-nutritional role in the ecology of M. synoviae and certain other mycoplasmas.
Summary The in vitro growth of tumor cells infected with mycoplasmas was suppressed by macrophages pretreated with interferon (IFN), but the growth of mycoplasma-free tumor cells was not suppressed. Pretreatment of macrophages with IFN plus mycoplasmas or their soluble factors either simultaneously or sequentially, IFN first and mycoplasmas second, but not in the reverse order, was effective in activating macrophages
Immunohistochemistry was used to demonstrate secreted mucins MUC2, MUC5AC and MUC5B and membrane-bound mucin MUC4 in the pulmonary bronchioles of piglets experimentally infected with Mycoplasma hyopneumoniae. Conventional status, Landrace–Duroc cross-bred piglets, 13days of age, were randomised to two groups. One group (n=20) was infected by the intra-tracheal route with the SNU98703 strain of M. hyopneumoniae, and a group of 12
C. H. Kim; Y. Oh; K. Han; H. W. Seo; D. Kim; I. Kang; C. Chae
Day-old broilers chicks (n=200), free of Mycoplasma gallisepticum (MG) were randomly divided in to five groups designated as A, B, C, D, and E, comprising 40 birds each. At the age of one week, the birds in groups A, B, C, and D, were inoculated with a virulent strain of MG intra-tracheally with 0.1 mL of the PPLO (Pleuro Pneumonia
M. A. KHAN; M. S. KHAN; M. YOUNUS; T. ABBAS; I. KHAN; N. A. KHAN
The Vsa proteins are associated with the virulence of the murine respiratory pathogen Mycoplasma pulmonis. The antigens consist of a conserved N-terminal region that is combined with one of several different variable C-terminal regions comprised of tandem repeats. M. pulmonis strains that produce VsaA with about 40 tandem repeats do not adhere to polystyrene or erythrocytes and are highly resistant
Two conservative gene clusters, the S10 ribosomal protein region and one (of the two) set of rRNA genes, were split in a genome crossover rearrangement event in Mycoplasma gallisepticum. As a result of the rearrangement the major part of the S10 ribosomal protein cluster is located upstream of genes for 23S-5S rRNA (rrn23-5), but the genes infA-rpl36-rps13-rpoA-rpl17 are located immediately
Andrei Skamrov; Eugenia Feoktistova; Maria Goldman; Robert Beabealashvilli
Background Reactive arthritis (ReA) has been sporadically reported as triggered by Mycoplasma pneumoniae. This study examined the potential relationship between the acute M. pneumoniae infection and juvenile spondyloarthropathy (jSpA) in children.Patients and methods Twelve patients with ReA secondary to acute M. pneumoniae were examined. M. pneumoniae-specific IgM, IgG and IgA antibodies were serologically confirmed by enzyme-linked immunosorbent assay (ELISA) tests (Savyon Diagnost.,
Miroslav Harjacek; Jelena Ostojic; Oktavija Djakovic Rode
Furuncular myiasis caused by Dermatobia hominis is endemic throughout Central and South America. However, because of widespread travel, furuncular myiasis has become more common in North America. Misdiagnosis and mismanagement can occur owing to limited awareness of the condition outside endemic areas. We report a case of furuncular myiasis in an immigrant from El Salvador with magnetic resonance imaging findings. The case is unique because neuroimaging was obtained upon the clinical suspicion of calvarial osteomyelitis. Parasitic infestation should be included in the differential diagnosis of a new skin lesion in patients who have traveled to endemic areas. PMID:22910983
Vijay, Kanupriya; Kalapos, Paul; Makkar, Abhishek; Engbrecht, Brett; Agarwal, Amit
Two Cryptosporidium isolates from separate infants suffering from diarrhea were obtained from a hospital in Zhengzhou, China and were genotyped by PCR amplification and sequence analysis of the small-subunit ribosomal RNA (rRNA) (SSU rRNA), 70-kDa heat shock protein (HSP70), and actin genes. Further subtyping was performed by PCR amplification and sequence analysis of the 60-kDa glycoprotein (gp60) gene. Both the isolates were identified as Cryptosporidium hominis subtype IdA21, a rare subtype previously found only in a human immunodeficiency virus-infected child in South Africa and another child in Jordan.
Aims: To design and validate a polymerase chain reaction (PCR) assay targeting the 16S rRNA gene of Mycoplasma genitalium. Methods: Primers were designed that were complementary to the 16S rRNA gene sequence of M genitalium. After optimisation of the reaction conditions, the PCR was tested against nine M genitalium strains, a dilution series of M genitalium DNA, and a panel of common microorganisms. The PCR was also challenged in parallel with a published assay against 54 urine specimens from men with urethritis. Results: The expected 341 bp product was produced on amplification of material from all M genitalium strains and from none of the other microorganisms tested. The lower limit of detection was 50 genome copies. The new assay detected M genitalium DNA in nine of 54 men with urethritis, in comparison with eight positive specimens detected with the alternative PCR. Conclusions: This novel PCR targeting the M genitalium 16S rRNA gene has been optimised and now provides a sensitive and specific alternative or addition to the available MgPa gene targeting assays.
Eastick, K; Leeming, J P; Caul, E O; Horner, P J; Millar, M R
Adherence of Mycoplasma gallisepticum to erythrocytes was examined by colony immunoblotting, detergent phase fractionation, trypsin treatment, comparison of protein profiles, and comparison of erythrocyte-bound mycoplasma protein fractions of hemadsorption-positive and -negative mutants. The binding of M. gallisepticum to chicken or human erythrocytes was found to be mediated via surface-exposed membrane proteins undergoing high-frequency phase variation.
Athamna, A; Rosengarten, R; Levisohn, S; Kahane, I; Yogev, D
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) recently emerged as a technology for the identification of bacteria. In this study, we aimed to evaluate its applicability to human and ruminant mycoplasmal identification, which can be demanding and time-consuming when using phenotypic or molecular methods. In addition, MALDI-TOF MS was tested as a subtyping tool for certain species. A total of 29 main spectra (MSP) from 10 human and 13 ruminant mycoplasma (sub)species were included in a mycoplasma MSP database to complete the Bruker MALDI Biotyper database. After broth culture and protein extraction, MALDI-TOF MS was applied for the identification of 119 human and 143 ruminant clinical isolates that were previously identified by antigenic or molecular methods and for subcultures of 73 ruminant clinical specimens that potentially contained several mycoplasma species. MALDI-TOF MS resulted in accurate (sub)species-level identification with a score of ?1.700 for 96% (251/262) of the isolates. The phylogenetically closest (sub)species were unequivocally distinguished. Although mixtures of the strains were reliably detected up to a certain cellular ratio, only the predominant species was identified from the cultures of polymicrobial clinical specimens. For typing purposes, MALDI-TOF MS proved to cluster Mycoplasma bovis and Mycoplasma agalactiae isolates by their year of isolation and genome profiles, respectively, and Mycoplasma pneumoniae isolates by their adhesin P1 type. In conclusion, MALDI-TOF MS is a rapid, reliable, and cost-effective method for the routine identification of high-density growing mycoplasmal species and shows promising prospects for its capacity for strain typing. PMID:23903545
Pereyre, S; Tardy, F; Renaudin, H; Cauvin, E; Del Prá Netto Machado, L; Tricot, A; Benoit, F; Treilles, M; Bébéar, C
The aim of this study was to evaluate the sensitivity and the detection limit of a real-time polymerase chain reaction (Q-PCR) developed for the qualitative and quantitative detection of Mycoplasma gallisepticum. No cross-reactivity was observed with DNA from other important avian mycoplasmas, including Mycoplasma synoviae and Mycoplasma meleagridis. However, the Q-PCR could not distinguish between M. gallisepticum and Mycoplasma imitans.
Molecular analysis was conducted on 36 Mycoplasma gallisepticum DNA extracts from tracheal swab samples of commercial poultry in seven South African provinces between 2009 and 2012. Twelve unique M. gallisepticum genotypes were identified by polymerase chain reaction and sequence analysis of the 16S-23S rRNA intergenic spacer region (IGSR), M. gallisepticum cytadhesin 2 (mgc2), MGA_0319 and gapA genetic regions. The DNA sequences of these genotypes were distinct from those of M. gallisepticum isolates in a database composed of sequences from other countries, vaccine and reference strains. The most prevalent genotype (SA-WT#7) was detected in samples from commercial broilers, broiler breeders and layers in five provinces. South African M. gallisepticum sequences were more similar to those of the live vaccines commercially available in South Africa, but were distinct from that of F strain vaccine, which is not registered for use in South Africa. The IGSR, mgc2 or MGA_0319 sequences of three South African genotypes were identical to those of the ts-11 vaccine strain, necessitating a combination of mgc2 and IGSR targeted sequencing to differentiate South African wild-type genotypes from ts-11 vaccine. To identify and differentiate all 12 wild-types, mgc2, IGSR and MGA_0319 sequencing was required. Sequencing of gapA was least effective at strain differentiation. This research serves as a model for the development of an M. gallisepticum sequence database, and illustrates its application to characterize M. gallisepticum genotypes, select diagnostic tests and better understand the epidemiology of M. gallisepticum. PMID:23889487
Armour, Natalie K; Laibinis, Victoria A; Collett, Stephen R; Ferguson-Noel, Naola
In recent years polymerase chain reaction (PCR) assays have become widely used as methods to confirm the presence of Mycoplasma gallisepticum and Mycoplasma synoviae in poultry flocks, but there has been limited standardization of the protocols used. Thirteen laboratories from five different countries participated in an interlaboratory comparison of detection of M. gallisepticum and M. synoviae DNA by PCR in
Lipoprotein LppQ, a predominant 48-kDa antigen, and its corresponding gene, lppQ, were characterized in Mycoplasma mycoides subsp. mycoides SC, the etiological agent of contagious bovine pleuropneumonia. The lppQ gene is specific to M. mycoides subsp. mycoides SC and was found in the type strain and in field strains isolated in Europe, Africa, and Australia, as well as in vaccinal strains. LppQ is encoded as a precursor with a consensus sequence for prokaryotic signal peptidase II and a lipid attachment site. The leader sequence shows significant prominent transmembrane helix structure with a predicted outside-to-inside helix formation capacity. The N-terminal domain of the mature LppQ was shown to be surface exposed. It induced a strong, specific, early, and persistent immune response in naturally and experimentally infected animals. The C-terminal domain of LppQ possesses an integral membrane structure built up of repeated units, rich in hydrophobic and aromatic amino acids, which have a pore formation potential. A recombinant peptide representing the N-terminal domain of LppQ was obtained by site-directed mutagenesis of nine Mycoplasma-specific TGA (Trp) codons into universal TGG (Trp) codons and expression in Escherichia coli hosts. It was used for serodetection of cattle infected with M. mycoides subsp. mycoides SC, in which it was detected postinfection for significantly longer than conventional serological test reactions.
After a trip to Belize, a 25-year-old man noticed an erythematous papule on his upper right chest that enlarged over a 6-week period and formed a central aperture. The patient reported feeling movement and intermittent lancinating pains under the skin. The history and examination were consistent with cutaneous myiasis, likely secondary to the human botfly, Dermatobia hominis. The objective of reporting this case is to present a simple method of extraction of a botfly larva using a commercial venom extractor. The patient's upper chest was prepared, and an occlusive dressing was placed over the lesion for 30 minutes. The Extractor Pump (Sawyer Products, Safety Harbor, FL) was applied and activated, and the larva was rapidly extracted completely intact with no significant discomfort to the patient. The wound fully healed without complication. D hominis is a common etiology of cutaneous myiasis endemic to Belize. The larva burrows under the skin of mammals where it develops for a period of weeks before erupting and falling to the soil to pupate. The diagnosis and treatment of botfly infestation is pertinent to doctors in the United States as Central and South America are common travel destinations for North Americans. In this case, a commercially available venom extractor was demonstrated to be a safe, noninvasive, and painless method for botfly extraction in the field without use of hospital resources. PMID:23246347
Mycoplasma virus L2 is an enveloped, nonlytic virus containing double-stranded, superhelical DNA. The L2 virion contains about 7 to 8 major proteins identified by SDS-polyacrylamide gel electrophoresis, but the virion has no discernible capsid structure. It has been suggested that the L2 virion is a DNA-protein condensation surrounded by a lipid-protein membrane. The host for mycoplasma virus L2 is Acholeplasma laidlawii. A. laidlawii has no cell wall and contains a small genome, 1 x 10/sup 9/ daltons, which is two to three times smaller than that of most bacteria. Infection of A. laidlawii by L2 is nonlytic. The studies in this thesis show that L2 DNA synthesis begins at about 1 hour of infection and lasts throughout the infection. Viral DNA synthesis is inhibited by chloramphenicol, streptomycin, and novobiocin. Packaging of L2 DNA into progeny virus is also inhibited by chloramphenicol and novobiocin. It is concluded that protein synthesis and probably DNA gyrase activity are required for L2 DNA synthesis, and for packaging of L2 DNA into progeny virus. DNA-DNA hybridization studies demonstrate that L2 DNA integrates into the host cell during infection, and subsequent to infection the cells are mycoplasma virus L2 lysogens. The viral site of integration has been roughly mapped. L2 virus is restricted and modified by A. laidlawii strains JA1 and K2. The nature of the modification in strain K2 has been elucidated. Two L2 variants containing insertions in the viral DNA were identified in these studies. Restriction endonuclease cleavage maps of these variants have been determined. DNA from L2 and another isolate of L2, MV-Lg-L 172, are compared in these studies. 74 references, 33 figures, 6 tables. (ACR)
Swabs from the posterior vaginal fornix were obtained from 804 consecutive female patients visiting a large Dutch sexually transmitted diseases (STD) outpatient clinic. A detailed clinical history was obtained and complaints concerning the lower genital tract, such as vaginal discharge or vulval and vaginal irritation, were recorded. Patients were examined and the presence of non-physiological vaginal secretions was established by
Alex van Belkum; Cindy van der Schee; Willem I van der Meijden; Henri A Verbrugh; Hans J. F Sluiters
The human bot fly, Dermatobia hominis (Linnaeus Jr., 1781) is a major pest of livestock in Mexico, Central and South America. Myiasis caused by the larvae result in economic losses due to hide damage and reductions in weight gain and milk production. They have a broad host range which includes wildl...
PCR primers corresponding to the adhesin genes of Mycoplasma pneumoniae and Mycoplasma genitalium were shown to detect the corresponding organisms specifically. Absence of cross-reaction with seven other mollicute species and six unrelated bacterial species commonly found in humans was demonstrated. Positive control primers directed against human mitochondria1 DNA could be mixed with the Mycoplasma primers without loss of specificity or
We developed a duplex PCR assay targeting the hemagglutinin multigene families, vlhA and pMGA, of Mycoplasma synoviae and Mycoplasma gallisepticum, respectively. The assay proved to be specific and sensitive enough to justify its use for the simultaneous detection of the two major avian mycoplasma species from field isolates. PMID:15695715
Mardassi, B Ben Abdelmoumen; Mohamed, R Ben; Gueriri, I; Boughattas, S; Mlik, B
We developed a duplex PCR assay targeting the hemagglutinin multigene families, vlhA and pMGA, of Mycoplasma synoviae and Mycoplasma gallisepticum, respectively. The assay proved to be specific and sensitive enough to justify its use for the simultaneous detection of the two major avian mycoplasma species from field isolates.
Ben Abdelmoumen Mardassi, B.; Ben Mohamed, R.; Gueriri, I.; Boughattas, S.; Mlik, B.
The continual cell lines of bovine kidneys MDBK and AUBEK, and porcine kidneys RPD and IBRS, spontaneously infected with Mycoplasma arginini and Acholeplasma laidlawii, were decontaminated by the method of selective elimination. Two elimination procedures were modified to be used for the decontamination: one based on the reduction of infection by the light treatment of the cultures, the other based on the selection of mycoplasma-free cell population through cell clonation. On the basis of a long-continued control of the cell clones a methodical procedure of the preparation of mycoplasma-free cell lines was worked out. PMID:3090766
To correlate viability with attachment capacity, Mycoplasma gallisepticum cell harvested at different growth phases and treated by various agents were tested for their capacity to attach to human erythrocytes. The results show that viability per se is not essential for M. gallisepticum attachment to erythrocytes, as cells killed by ultraviolet irradiation anmd membranes isolated by lysing M. gallisepticum cells by various means retained attachment capacity. However, treatment of the mycoplasmas by protein-denaturing agents, such as heart, glutaraldehyde, or prolonged exposure to low pH, drastically affected or even abolished attachment, supporting the protein nature of the mycoplasma membrane components responsible for specific binding to the sialoglycoprotein receptors on the erythrocytes.
On October 28, 2011, the West Virginia Department of Health and Human Resources notified CDC of an increase in pneumonia cases among school-aged children in two rural counties. Mycoplasma pneumoniae was the suspected cause, based on epidemiology, clinical presentation, and testing of specimens sent to CDC. Three of six nasopharyngeal swabs were positive for M. pneumoniae in testing by quantitative real-time polymerase chain reaction (qPCR). The West Virginia Department of Health and Human Resources and CDC conducted an outbreak investigation to confirm the etiology of the outbreak, establish active case surveillance, and provide recommendations for treatment and containment. The investigation confirmed M. pneumoniae as the cause and identified 125 cases, including two caused by macrolide-resistant isolates. The outbreak was contained with public health interventions that included communicating to the public the importance of respiratory hygiene, providing hand sanitizer in schools, and informing health-care providers about macrolide resistance; antibiotic prophylaxis was not used. Despite the large number of cases and macrolide-resistant strains, no severe extrapulmonary manifestations (e.g., erythema multiforme) were reported. PMID:23076092
Mycoplasma genitalium is a member of genital mycoplasmas, which is emerging as an important causative agent of sexually transmitted infections both in males and females. The advent of polymerase chain reaction and other molecular methods have made studies on M. genitalium more feasible, which is otherwise a difficult organism to isolate. Besides Chlamydia trachomatis, M. genitalium is now an important and established cause of non gonococcal urethritis (NGU) in men, more so in persistent and recurrent NGU. Multiple studies have also shown a positive association of M. genitalium with mucopurulent cervicitis and vaginal discharge in females as well. The evidences for M. genitalium pelvic inflammatory diseases and infertility are quite convincing and indicate that this organism has potential to cause ascending infection. Lack of clear association with M. genitalium has been reported for bacterial vaginosis and adverse pregnancy outcomes. Diagnosis of M. genitalium infections is performed exclusively using nucleic acid amplification tests (NAATs), owing to poor or slow growth of bacterium in culture. Although there are no guidelines available regarding treatment, macrolide group of antimicrobials appear to be more effective than tetracyclines. The present review provides an overview of the epidemiology, pathogenesis, clinical presentation and management of sexually transmitted infections due to M. genitalium.
...Mycoplasma pneumoniae range from inapparent infection to mild or severe upper respiratory disease, ear infection, and bronchial pneumonia. (b) Classification. Class I (general controls). The device is exempt from the premarket notification...
The manual is intended to assist the laboratory worker who is involved in the isolation and identification of Mycoplasma species of human origin. The preparation of various media, rabbit antisera, complement-fixing antigen, and immunizing antigens are pre...
Mycoplasmas are most unusual self-replicating bacteria, possessing very small genomes, lacking cell wall components, requiring cholesterol for membrane function and growth, using UGA codon for tryptophan, passing through \\
Joel B. Baseman; Joseph G. Tully; Thomas Henry Huxley
The activity of oleuropein, a phenolic glycoside contained in olive oil, was investigated in vitro against Mycoplasmahominis, Mycoplasma fermentans, Mycoplasma pneumoniae and Mycoplasma pirum. Oleuropein inhibited mycoplasmas at concentrations from 20 to 320 mg\\/l. The MICs of oleuropein to M. pneumoniae, M. pirum, M. hominis and M. fermentans were 160, 320, 20 and 20 mg\\/l, respectively.
Pio Maria Furneri; Andreana Marino; Antonina Saija; Nicola Uccella; Giuseppe Bisignano
A total of 212 coagulase-negative Staphylococcus strains recovered prospectively during 119 surgeries for proven or suspected bone and joint infection (BJI) were identified by sodA sequencing. These strains were identified as 151 Staphylococcus epidermidis isolates, 15 S. warneri isolates, 14 S. capitis isolates, 9 S. hominis isolates, 6 S. lugdunensis isolates, 5 S. haemolyticus isolates, 4 S. caprae isolates, 4
V. Sivadon; M. Rottman; S. Chaverot; J.-C. Quincampoix; V. Avettand; P. de Mazancourt; L. Bernard; P. Trieu-Cuot; J.-M. Feron; A. Lortat-Jacob; P. Piriou; T. Judet; J.-L. Gaillard
Earlier studies implied a role for Mycoplasma arthritidis surface protein MAA2 in cytadherence and virulence and showed that it exhibited both size and phase variability. Here we report the further analysis of MAA2 and the cloning and sequencing of the maa2 gene from two M. arthritidis strains, 158p10p9 and H606, expressing two size variants of MAA2. Triton X-114 partitioning and metabolic labeling with [3H]palmitic acid suggested lipid modification of MAA2. Surface exposure of the C terminus was indicated by cleavage of monoclonal antibody-specific epitopes from intact cells by carboxypeptidase Y. The maa2 genes from both strains were highly conserved, consisting largely of six (for 158p10p9) or five (for H606) nearly identical, 264-bp tandem direct repeats. The deduced amino acid sequence predicted a largely hydrophilic, highly basic protein with a 29-amino-acid lipoprotein signal peptide. The maa2 gene was expressed in Escherichia coli from the lacZ promoter of vector pGEM-T. The recombinant product was approximately 3 kDa larger than the native protein, suggesting that the signal peptide was not processed in E. coli. The maa2 gene and upstream DNA sequences were cloned from M. arthritidis clonal variants differing in MAA2 expression state. Expression state correlated with the length of a poly(T) tract just upstream of a putative -10 box. Full-sized recombinant MAA2 was expressed in E. coli from genes derived from both ON and OFF expression variants, indicating that control of expression did not include alterations within the coding region. PMID:9596719
Washburn, L R; Weaver, K E; Weaver, E J; Donelan, W; Al-Sheboul, S
The polymerase chain reaction (PCR) has been used for the general detection ofMollicutes. 25Mycoplasma andAcholeplasma species were detected including important contaminants of cell cultures such asM. orale, M. arginini, M. hyorhinis, M. fermentans, A. laidlawii and additional human and animal mycoplasmas. PCR reactions were performed using a set of nested primers defined from conserved regions of the 16S rRNA gene.
Homologous internal controls were used as competitor DNA in the polymerase chain reaction for the quantitative detection of mycoplasma DNA. PCR primer sets were designed on the basis of the most conserved nucleotide sequences of the 16S rRNA gene of mycoplasma species. Amplification of this gene was examined in five different mycoplasma species: Mycoplasma orale, M. hyorhinus, M. synoviae, M. gallisepticum and M. pneumoniae. To evaluate the primers, a number of different cell lines were assayed for the detection of mycoplasma infections. All positive cell lines showed a distinct product on agarose gels while uninfected cells showed no DNA amplification. Neither bacterial nor eukaryotic DNA produced any cross-reaction with the primers used, thus confirming their specificity. Internal control DNA to be used for quantitation was constructed by modifying the sizes of the wild-type amplified products and cloning them in plasmid vectors. These controls used the same primer binding sites as the wild-type and the amplified products were differentiated by a size difference. The detection limits for all the mycoplasma species by competitive quantitative PCR were estimated to range from 4 to 60 genome copies per assay as determined by ethidium bromide-stained agarose gels. These internal standards also serve as positive controls in PCR-based detection of mycoplasma DNA, and therefore accidental contamination of test samples with wild-type positive controls can be eliminated. The quantitative PCR method developed will be useful in monitoring the progression and significance of mycoplasma in the disease process. PMID:7750739
A novel small haemoplasma was detected following cytological examination of blood smears from a splenectomized dog with haemic neoplasia. The 16S rRNA and rnpB genes of the organism were partially sequenced and a phylogenetic tree constructed. The organism was most closely related to the small feline haemoplasma, 'Candidatus Mycoplasma haemominutum' (94 % 16S rRNA gene nucleotide sequence identity; 75 %
Jane E. Sykes; Louise M. Ball; Nathan L. Bailiff; Michael M. Fry
The effect of fermented wheat germ extract (FWGE, Immunovet-HBM) was studied in chickens challenged with Mycoplasma gallisepticum. Ninety M. gallisepticum- and M. synoviae-free 3-wk-old chickens were exposed to aerosol infection of M. gallisepticum. One group (30 birds) was treated with FWGE, a second group with tiamulin, and a third group was untreated. The fourth group was exposed to PBS aerosol as a negative control. On d 9, all chickens were slaughtered and examined for the presence of gross and histological lesions, the presence of the challenge strain in the organs and specific antibodies in the serum. Body weight gains and feed conversion rates were recorded. In the groups treated with FWGE and with tiamulin, the chickens remained clinically healthy: their BW gains were 441.7 g and 446.8 g, respectively. Feed conversion ratios were 1.72 and 1.71 for FWGE- and tiamulin-treated birds, respectively. Control birds had BW gain of 480.8 g, and feed conversion ratio of 1.78. The numbers of birds with gross lesions (15 and 11, respectively) and lesion scores (25 and 25, respectively) of the FWGE- and tiamulin-treated groups were significantly lower than in the infected untreated group (25 birds, lesion score of 190). No mycoplasma was reisolated from brain, liver, spleen, heart, or kidneys of the FWGE-treated birds, and the number of mycoplasma isolations from the respiratory tract samples was less frequent (10) than from the infected untreated group (64). In addition, 35 samples from other internal organs were also positive. Twenty percent of the birds treated with FWGE showed serological response with a 5.0% reaction score, whereas in the infected untreated group, 83.3% of birds were reactors, with a 62.5% reaction score. PMID:15554060
Stipkovits, L; Lapis, K; Hidvégi, M; Kósa, E; Glávits, R; Resetár, A
The ability of Mycoplasma hyopneumoniae to persist despite fluoroquinolone treatments was investigated with pigs. Groups of specific-pathogen-free pigs were experimentally infected with M. hyopneumoniae strain 116 and treated with marbofloxacin at the therapeutic dose (TD) or half of the therapeutic dose (TD/2) for 3 days. Results showed that, despite tissue penetration of marbofloxacin, particularly in the trachea and the tracheal secretions, the treatments did not have any influence on M. hyopneumoniae recovery from tracheal swabs. Mycoplasmas were also isolated from inner organs and tissues such as liver, spleen, kidneys, and bronchial lymph nodes. Recontamination of pigs via environment could not explain mycoplasma persistence after medication, as decontamination of pigs and allocation to a new disinfected environment did not have any significant effect on the phenomenon. A significant decrease in the susceptibility level to marbofloxacin of 12 mycoplasma clones reisolated after the treatments (TD/2 and TD) was observed. Two point mutations were found in the ParC quinolone resistance-determining region (QRDR) of DNA topoisomerase IV (Ser80?Phe and Asp84?Asn), and one point mutation was observed just behind the QRDR of ParC (Ala116?Glu). This is the first time that mutations in a gene coding for topoisomerase IV have been described for M. hyopneumoniae after in vivo marbofloxacin treatments in experimentally infected pigs. However, development of resistance is not sufficient to explain M. hyopneumoniae persistence in vivo since (i) marbofloxacin concentrations were above the marbofloxacin MIC of the wild-type strain and (ii) mycoplasmas reisolated after a single injection of marbofloxacin did not display an increased marbofloxacin MIC.
Le Carrou, J.; Laurentie, M.; Kobisch, M.; Gautier-Bouchardon, A. V.
P48 is a 48 kDa monocytic differentiation/activation factor previously purified from the conditioned medium of the Reh human pre-B cell leukaemia cell line. It induces growth arrest and differentiation of HL-60 human promyelocytic leukaemia cells along the monocytic pathway and the production of the cytokines interleukin 1, tumour necrosis factor-alpha and interleukin 6 in human monocytes and monocytic cell lines. The cDNA for P48 was cloned from Reh cellular RNA using 3' reverse amplification of cDNA ends. Southern blot probing with P48 cDNA revealed hybridization with DNA from Reh and Molt-4 cells, but not with DNA from human peripheral blood mononuclear cells. Subsequent studies using PCR and Southern analysis revealed P48 sequences in DNA isolated from Mycoplasma fermentans but not M. hominis, M.iowae, M.synoviae or M.lypophilum. Although initial studies using Mycoplasma culture and hybridization techniques had failed to reveal Mycoplasma infection in our Reh and Molt-4 cell lines, subsequent PCR studies using Mycoplasma genus-specific rRNA primrs revealed Mycoplasma sequences in these cell lines. Using the P48 cDNA probe, we isolated a genomic clone from M. fermentans DNA which was found to be 98.5% identical with the P48 cDNA clone, and the deduced amino acid sequence agreed with N-terminal microsequencing data for P48 protein purified from the Reh cell line conditioned medium. The 5' end of the gene has a number of consensus sequences characteristic of prokaryotic genes, and the deduced amino acid sequence has a number of features suggesting that P48 is a lipoprotein. The P48 cDNA was expressed in pMAL in Escherichia coli, and the 60 kDa expressed fusion protein was found to react with anti-P48 antibodies on Western blots. This is consistent with a pMAL fusion protein representing the sum of the 42 kDa maltose-binding protein and 18 kDa of P48 recombinant protein, suggesting that native P48 has significant post-translational modification. Consistent with this, Northern blot studies revealed a single 1 kb transcript. The recombinant fusion protein was found to possess anti-proliferative activity against HL-60 cells, and antibodies against recombinant P48 were found to block the biological activity of native P48 isolated from conditioned medium. These studies demonstrate that P48, a molecule with immunomodulatory and haematopoietic differentiation activities, is derived from M. fermentans or a closely related species. P48 may be important in the pathophysiology of Mycoplasma infections and may be useful in dissecting the mechanisms involved in mammalian haematopoietic cell differentiation, immune function and cytokine biosynthesis.
Hall, R E; Agarwal, S; Kestler, D P; Cobb, J A; Goldstein, K M; Chang, N S
P48 is a 48 kDa monocytic differentiation/activation factor previously purified from the conditioned medium of the Reh human pre-B cell leukaemia cell line. It induces growth arrest and differentiation of HL-60 human promyelocytic leukaemia cells along the monocytic pathway and the production of the cytokines interleukin 1, tumour necrosis factor-alpha and interleukin 6 in human monocytes and monocytic cell lines. The cDNA for P48 was cloned from Reh cellular RNA using 3' reverse amplification of cDNA ends. Southern blot probing with P48 cDNA revealed hybridization with DNA from Reh and Molt-4 cells, but not with DNA from human peripheral blood mononuclear cells. Subsequent studies using PCR and Southern analysis revealed P48 sequences in DNA isolated from Mycoplasma fermentans but not M. hominis, M.iowae, M.synoviae or M.lypophilum. Although initial studies using Mycoplasma culture and hybridization techniques had failed to reveal Mycoplasma infection in our Reh and Molt-4 cell lines, subsequent PCR studies using Mycoplasma genus-specific rRNA primrs revealed Mycoplasma sequences in these cell lines. Using the P48 cDNA probe, we isolated a genomic clone from M. fermentans DNA which was found to be 98.5% identical with the P48 cDNA clone, and the deduced amino acid sequence agreed with N-terminal microsequencing data for P48 protein purified from the Reh cell line conditioned medium. The 5' end of the gene has a number of consensus sequences characteristic of prokaryotic genes, and the deduced amino acid sequence has a number of features suggesting that P48 is a lipoprotein. The P48 cDNA was expressed in pMAL in Escherichia coli, and the 60 kDa expressed fusion protein was found to react with anti-P48 antibodies on Western blots. This is consistent with a pMAL fusion protein representing the sum of the 42 kDa maltose-binding protein and 18 kDa of P48 recombinant protein, suggesting that native P48 has significant post-translational modification. Consistent with this, Northern blot studies revealed a single 1 kb transcript. The recombinant fusion protein was found to possess anti-proliferative activity against HL-60 cells, and antibodies against recombinant P48 were found to block the biological activity of native P48 isolated from conditioned medium. These studies demonstrate that P48, a molecule with immunomodulatory and haematopoietic differentiation activities, is derived from M. fermentans or a closely related species. P48 may be important in the pathophysiology of Mycoplasma infections and may be useful in dissecting the mechanisms involved in mammalian haematopoietic cell differentiation, immune function and cytokine biosynthesis. PMID:8921000
Hall, R E; Agarwal, S; Kestler, D P; Cobb, J A; Goldstein, K M; Chang, N S
Hemotrophic mycoplasmas, bacteria without cell walls whose niche is the erythrocytes of their hosts, have never been cultivated in vitro. Therefore, knowledge of their pathogenesis is fundamental. Mycoplasma suis infects pigs, causing either acute fatal hemolytic anemia or chronic low-grade anemia, growth retardation, and immune suppression. Recently, the complete genomes of two hemotrophic mycoplasma species, M. suis and M. haemofelis, were sequenced, offering new strategies for the analysis of their pathogenesis. In this study we implemented a proteomic approach to identify M. suis proteins during acute infection by using tandem mass spectrometry. Twenty-two percent of the predicted proteins encoded in M. suis strain KI_3806 were identified. These included nearly all encoded proteins of glycolysis and nucleotide metabolism. The proteins for lipid metabolism, however, were underrepresented. A high proportion of the detected proteins are involved in information storage and processing (72.6%). In addition, several proteins of different functionalities, i.e., posttranslational modification, membrane genesis, signal transduction, intracellular trafficking, inorganic ion transport, and defense mechanisms, were identified. In its reduced genome, M. suis harbors 65.3% (strain Illinois) and 65.9% (strain KI_3806) of the genes encode hypothetical proteins. Of these, only 6.3% were identified at the proteome level. All proteins identified in this study are present in both M. suis strains and are encoded in more highly conserved regions of the genome sequence. In conclusion, our proteome approach is a further step toward the elucidation of the pathogenesis and life cycle of M. suis as well as the establishment of an in vitro cultivation system.
Felder, Kathrin M.; Carranza, Paula M.; Gehrig, Peter M.; Roschitzki, Bernd; Barkow-Oesterreicher, Simon; Hoelzle, Katharina; Riedel, Katharina; Kube, Michael
Since 1970 Mycoplasma fermentans has been suspected of being associated with rheumatoid arthritis. However, this association has been difficult to prove, and this has been our goal. The distribution of M. fermentans was studied in the synovial fluid of patients suffering from different arthritides. Samples of synovial fluid were taken from patients with well-defined disease and a clear diagnosis. After removal of the inflammatory cells and hyaluran, they were treated with proteinase K and tested by a single or fully nested PCR with primers directed against part of the two 16S rRNA genes of M. fermentans. The product was sequenced automatically, by using an ALF Express automatic sequencer, to confirm the mycoplasma species and to identify the strain since the two genes were usually found to be polymorphic. This was also true of the type strain, strain PG18. M. fermentans was detected in 23 of 26 (88%) rheumatoid arthritis patients, and four different strains were found. It was also found in 7 of 8 (88%) of the nonrheumatoid inflammatory arthritis patient group, which consisted of one patient with reactive arthritis, one patient with pauciarticular juvenile chronic arthritis, two patients with gout, two patients with ankylosing spondylitis, and two patients with psoriatic arthritis, only one of whom was infected with M. fermentans. It was not detected in any of the 10 osteoarthritis patients. M. fermentans was therefore found to be a variable and very common organism in arthritic patients with inflammatory joint exudates and may well prove to be important in the etiology of the diseases.
Johnson, Sheena; Sidebottom, David; Bruckner, Felix; Collins, David
Few genetic systems for studying mycoplasmas exist, but transposon Tn916 has been shown to transpose into the genomes of some species and can be used as an insertional mutagen. In the current study, the ability of Enterococcus faecalis to serve as a donor for the conjugative transfer of transposon Tn916 into the genome of the avian pathogen Mycoplasma gallisepticum strain PG31 was examined. Transconjugants were obtained at a frequency of > or =6 x 10(-8) per recipient CFU. To determine the transposon insertion site, an oligonucleotide primer corresponding to the 3' end of Tn916 was designed for the purpose of directly sequencing genomic DNA without PCR amplification. Using the direct sequencing approach, Tn916 was shown to insert into any of numerous sites in the M. gallisepticum genome. This is the first report of conjugal transposition of Tn916 into the M. gallisepticum genome. The ability to determine transposon insertion sites in mycoplasmas by genomic sequencing has not been previously described and allows rapid sequence analysis of transposon-generated mutants. PMID:10964629
Ruffin, D C; van Santen, V L; Zhang, Y; Voelker, L L; Panangala, V S; Dybvig, K
Lipid-free polysaccharide fraction 2 extracted from Mycoplasma pneumoniae strain FH by Prescott et al. (J. Bacteriol. 91:2117-2115, 1966) was examined for its ability to cross-precipitate antibody from type-specific rabbit antipneumococcal sera types 1 to 34 inclusive. Cross-precipitation in type-specific pneumococcal anti-type 23 and anti-type 32 sera was examined in detail and could be attributed to a rhamnose-galactose-rich component of crude M. pneumoniae polysaccharide fraction 2 recovered from immunoprecipitates formed with anti-type 23 serum. Immunochemically isolated mycoplasma polysaccharide was found to contain glucose, galactose, rhamnose, and mannose in 1:14:5:4 molar proportions. Comparison of the ability of 6-O-alpha-L-rhamnosyl-D-glucose and free L-rhamnose to inhibit precepitation by homologous pneumococcal and heterologous mycoplasma polysaccharide antigens indicates a combining site specificity for anti-type 23 and anti-type 32 antibodies directed largely against the alpha-linked L-rhamnosyl determinants and the occurrence of alpha-L-rhamnosyl units in type 32 and M. pneumoniae polysaccharides. Hapten inhibition of the cross-precipitation of pneumococcal type 23 capsular polysaccharide in anti-type 32 serum helps to establish that cross-reactivity can be attributed to interaction of recurrent, alpha-L-rhamnosyl units of type 23 with anit-alpha-L-rhamnoside combining sites of anti-type 32 antibodies.
In order to study horizontal transmission of Mycoplasma synoviae an avian pathogen, a reverse transcription-polymerase chain reaction (RT-PCR) assay was developed to detect viable Mycoplasma in environment. The test was based on the RT-PCR of the 16S ribosomal RNA (rRNA) of Mycoplasma genus. Results showed that Mycoplasma 16S rRNA was stable up to 23h after cell death. Therefore, the test
The specific, sensitive and reliable detection of mycoplasma contamination in cell cultures is an important part of mycoplasma control. We have sought to develop and validate a method for mycoplasma detection which is sensitive and accurate, but also practical in the sense of time spent, costs, and applicability in the standard laboratory; finally, the method should be suitable for screening
Aim: Genital mycoplasmas are known as sexually transmitted agents, causing mainly urethritis, pelvic inflammatory disease, spontaneous abortion, pyelonephritis, infertility, still birth, low birth weight, neonatal meningititis, and neonatal pneumonia. Mycoplasma infections not only jeopardize fertility but also pose a risk for infertility treatment and resulting pregnancies. Diagnosis of genital mycoplasma infections by bacterial conventional methods is very difficult. The aim
57 The molecular genetic principles of mycoplasma adaptation to the adverse growth conditions are of special interest. Restricted biochemical capabilities of mycoplasmas that are the smallest prokaryotic organisms capable of independent reproduction by no means prevent their distribution in various biocenoses, persistence in higher eukaryotes, and circulation in the nature. However, the mechanisms used by mycoplasma are still little studied
V. M. Chernov; N. E. Mukhametshina; Yu. V. Gogolev; T. N. Nesterova; O. A. Chernova
Mycoplasma, the smallest self-replicating organism with a minimal metabolism and little genomic redundancy, is expected to be a close approximation to the minimal set of genes needed to sustain bacterial life. This study employs comparative evolutionary analysis of twenty Mycoplasma genomes to gain an improved understanding of essential genes. By analyzing the core genome of mycoplasmas, we finally revealed the
The length of the tandem repeat region of the Vsa protein of Mycoplasma pulmonis has previously been shown to modulate the susceptibility of mycoplasmas to killing by complement: cells that produce a short form of the Vsa protein are highly sensitive, and cells producing the long Vsa protein are resistant. In contrast to their differing susceptibilities to complement, the mycoplasmas
Different from most other host-specific mycoplasmas, Mycoplasma gallinarum has been isolated from various hosts, such as poultry, pig, cattle, and sheep. The wide distribution among different hosts, the low pathogenesis, and the weak host immunological responses suggest this mycoplasma has a unique ...
Hemotropic mycoplasmas are bacteria that infect erythrocytes and cause subclinical infections to life-threatening disease. We describe hemotropic mycoplasma infection in a free-ranging black howler monkey (Alouatta caraya). This is the first molecular detection of a hemotropic mycoplasma in a nonhuman primate from Brazil. PMID:23778631
Santos, Leonilda C; Cubilla, Michelle P; de Moraes, Wanderlei; Cubas, Zalmir S; Oliveira, Marcos J; Estrada, Marko; Leutenegger, Christian M; Sykes, Jane E; Lindsay, Leann L; Marcondes, Mary; Barros Filho, Ivan R; Biondo, Alexander W
Two conservative gene clusters, the S10 ribosomal protein region and one (of the two) set of rRNA genes, were split in a genome crossover rearrangement event in Mycoplasma gallisepticum. As a result of the rearrangement the major part of the S10 ribosomal protein cluster is located upstream of genes for 23S-5S rRNA (rrn23-5), but the genes infA-rpl36-rps13-rpoA-rpl17 are located immediately downstream of the isolated gene for 16S rRNA (rrn16). A new ribosomal protein cluster infA-rpl36-rps13-rpoA-rpl17-rps16-trmD-rpl19 was formed. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that this ribosomal protein cluster is an operon. PMID:11959450
Skamrov, Andrei; Feoktistova, Eugenia; Goldman, Maria; Beabealashvilli, Robert
Asthmatic patients frequently develop wheezing after respiratory tract infection. Mycoplasma pneumoniae causes respiratory tract infections in children and in adults. The purpose of this study was to determine the frequency of recovery of M. pneumoniae from patients with asthma. Seventy-seven patients with asthma and 88 persons without asthma or any other respiratory tract disease (controls) were included in the study. Ages ranged from 8 months to 31 years. Throat swabs were taken and deposited in egg yolk broth with methylene blue in order to isolate M. pneumoniae. The bacterium was identified using inhibition of growth with homologous antiserum. The isolation rate in patients with asthma was 24.7% while that in controls was 5.7% (P < .01). The results suggest that M. pneumoniae colonizes a higher percentage of patients with asthma than controls and could possibly have induced the wheezing. PMID:8424592
The mechanism of quinolone resistance in Mycoplasma genitalium remains poorly understood due to difficulties with in vitro culture, especially of clinical isolates. In this study, to confirm the association between mutations in topoisomerases and antimicrobial susceptibilities to quinolones, ciprofloxacin-resistant mutant strains were selected using the cultivable type strain ATCC 33530. Sequence analysis revealed that the mutant strains harbored mutations in topoisomerase IV: Gly81Cys in ParC, Pro261Thr in ParC, or Asn466Lys in ParE. The MICs of all quinolones tested against the mutant strains were 2- to 16-fold higher than those against the wild-type strain. No cross-resistance was observed with macrolides or tetracyclines. We determined the inhibitory activities of quinolones against DNA gyrase and topoisomerase IV in order to investigate the correlation between antimicrobial susceptibility and inhibitory activity against the target enzymes, considered the primary targets of quinolones. Furthermore, using enzymatic analysis, we confirmed that Gly81Cys in the ParC quinolone resistance-determining region (QRDR) contributed to quinolone resistance. This is the first study to isolate quinolone-resistant mutant strains of M. genitalium harboring substitutions in the parC or parE gene in vitro and to measure the inhibitory activities against the purified topoisomerases of M. genitalium.
Background Mycoplasma pneumoniae (Mpn) is a human pathogen that causes acute and chronic respiratory diseases and has been linked to many extrapulmonary diseases. Due to the lack of cell wall, Mpn is resistant to antibiotics targeting cell wall synthesis such as penicillin. During the last 10 years macrolide-resistant Mpn strains have been frequently reported in Asian countries and have been spreading to Europe and the United States. Therefore, new antibiotics are needed. In this study, 30 FDA-approved anticancer or antiviral drugs were screened for inhibitory effects on Mpn growth and selected analogs were further characterized by inhibition of target enzymes and metabolism of radiolabeled substrates. Results Sixteen drugs showed varying inhibitory effects and seven showed strong inhibition of Mpn growth. The anticancer drug 6-thioguanine had a MIC (minimum inhibitory concentration required to cause 90% of growth inhibition) value of 0.20 ?g ml-1, whereas trifluorothymidine, gemcitabine and dipyridamole had MIC values of approximately 2 ?g ml-1. In wild type Mpn culture the presence of 6-thioguanine and dipyridamole strongly inhibited the uptake and metabolism of hypoxanthine and guanine while gemcitabine inhibited the uptake and metabolism of all nucleobases and thymidine. Trifluorothymidine and 5-fluorodeoxyuridine, however, stimulated the uptake and incorporation of radiolabeled thymidine and this stimulation was due to induction of thymidine kinase activity. Furthermore, Mpn hypoxanthine guanine phosphoribosyl transferase (HPRT) was cloned, expressed, and characterized. The 6-thioguanine, but not other purine analogs, strongly inhibited HPRT, which may in part explain the observed growth inhibition. Trifluorothymidine and 5-fluorodeoxyuridine were shown to be good substrates and inhibitors for thymidine kinase from human and Mycoplasma sources. Conclusion We have shown that several anticancer and antiviral nucleoside and nucleobase analogs are potent inhibitors of Mpn growth and that the mechanism of inhibition are most likely due to inhibition of enzymes in the nucleotide biosynthesis pathway and nucleoside transporter. Our results suggest that enzymes in Mycoplasma nucleotide biosynthesis are potential targets for future design of antibiotics against Mycoplasma infection.
A second cytadhesin-like protein, MGC2, was identified in the avian respiratory pathogen Mycoplasma gallisepticum. The 912-nucleotide mgc2 gene encodes a 32.6-kDa protein with 40.9 and 31.4% identity with the M. pneumoniae P30 and M. genitalium P32 cytadhesins, respectively. Functional studies with reverse transcription-PCR, immunoblotting, double-sided immunogold labeling, and attachment inhibition assays demonstrated homology to the human mycoplasmal P30 and P32 cytadhesins. These findings suggest that there is a family of cytadhesin genes conserved among pathogenic mycoplasmas infecting widely divergent hosts. PMID:9632619
Hnatow, L L; Keeler, C L; Tessmer, L L; Czymmek, K; Dohms, J E
To develop a diagnostic tool to identify Mycoplasma pulmonis (M. pulmonis) in clinical isolates, we developed a polymerase chain reaction (PCR) assay using primers specific for the 16S-23S rRNA intergenic spacer region (SR) of M. pulmonis. One pair of PCR primers reacted specifically with two reference strains of M. pulmonis tested and seven samples isolated from naturally infected rats. The
H Takahashi-Omoe; K Omoe; S Matsushita; H Kobayashi; K Yamamoto
The intergenic region (IG) transcriptional activity of Mycoplasma hyopneumoniae strain 232 was studied via two-color microarrays and quantitative real-time polymerase chain reactions (RT-qPCR). Two types of microarrays were constructed, one consisting of PCR products and the other of synthesized oligonucleotides. The PCR-array consisted of 994 PCR products (probes) which covers 98% (683\\/698) of the total open reading frames (ORFs) of
The use of a buffer system based on N-[2-hydroxyethyl]piperazine-N?-[2-ethanesulfonic acid] (HEPES), in conjunction with standard Gourlay’s culture medium was investigated for the growth and maintenance of Mycoplasma mycoides subsp. mycoides SC vaccine strain T144. When the initial pH of the culture medium was adjusted to 8.0, 0.075 M HEPES–NaOH was found to be sufficient to prevent the pH falling below
Human immunodeficiency virus (HIV)-infected persons are commonly infected with Cryptosporidium species and Enterocytozoon bieneusi in both developed and developing countries, particularly patients with CD4+ cell counts below 200 cells/?L; 285 HIV-infected patients on highly active antiretroviral therapy (HAART) were enrolled in this study, and both stool and blood specimens were collected from participants. The stool specimens were analyzed and typed for E. bieneusi and Cryptosporidium spp. by polymerase chain reaction (PCR) and DNA sequencing. CD4 count was analyzed using flow cytometry. E. bieneusi and Cryptosporidium were detected in 18 (6.3%) and 4 (1.4%) patients, respectively. The E. bieneusi detected mostly belonged to a new genotype group that, thus far, has only been found in a few humans: genotype Nig4 in 2 patients and two new genotypes related to Nig4 in 12 patients. The Cryptosporidium detected included C. hominis (two patients), C. parvum (one patient), and C. felis (one patient), with the two C. hominis infections belonging to an unusual subtype family. Additional studies are required to determine whether some E. bieneusi genotypes and C. hominis subtypes are more prevalent in HIV patients on HAART. PMID:23629938
Akinbo, Frederick O; Okaka, Christopher E; Omoregie, Richard; Adamu, Haileeyesus; Xiao, Lihua
Naturally-occurring mycoplasmal conjunctivitis is described among 104 wild-caught, and initially seronegative, house finches (Carpodacus mexicanus) maintained in captivity for 12 wk during November 1995 through January 1996. Finches housed in three pens were monitored for clinical signs, and > or = 10 birds were euthanatized for necropsy and mycoplasma testing every 2 wk. Within 2 to 4 wk following initial detection of lesions, > 50% of the birds in each of three pens developed a debilitating disease characterized by mild to severe ocular swelling, conjunctivitis, and ocular and nasal discharge. Microscopic lesions in affected finches consisted of mild to severe lymphoplasmacytic inflammation with epithelial and lymphoid hyperplasia in conjunctivae, nasal turbinates, and trachea. Mycoplasma gallisepticum infection was confirmed by culture or polymerase chain reaction (PCR) in all birds with conjunctival lesions and in 43% of birds without lesions. An arbitrary primer PCR was used to confirm M. gallisepticum isolates as identical to a field strain previously associated with house finch conjunctivitis. Most birds (89%) with conjunctivitis developed a concurrent antibody response detectable by serum plate agglutination (SPA) within 2 wk of lesion development. Hemagglutination inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) tests were less sensitive than the SPA test. The clinical severity of this disease and high proportion of affected birds suggests that M. gallisepticum may have a negative impact on free-flying house finch populations. PMID:9577775
Luttrell, M P; Stallknecht, D E; Fischer, J R; Sewell, C T; Kleven, S H
Mycoplasma hyopneumoniae is a primary agent associated with mycoplasma pneumonia and the porcine respiratory disease complex (PRDC). Various reports have indicated that different strains of M. hyopneumoniae are circulating in the swine population. Lysates from lung swabs from naturally infected pigs of different ages were tested according to a new variable number of tandem repeats (VNTR) genetic typing method based on the polyserine repeat motif of the P146 lipoproteoadhesin, which can be applied directly on clinical material without isolation of M. hyopneumoniae. The aim was to determine the diversity of M. hyopneumoniae isolates from conventional farrow-to-finish pig farms located in different geographical areas of Serbia. PCR amplification was carried out using M. hyopneumoniae -specific designed, conserved primers (p146MH-L and p146MH-R) flanking the region encoding the repeat motif, followed by sequencing and cluster analysis. Five groups of M. hyopneumoniae with thirteen to twenty-four serine repeats were observed. Analysis of three samples from each farm indicated that the specific isolate is ubiquitous in pigs of different ages. Furthermore, seven clusters were observed within 27 tested samples. The results indicated a considerable diversity among M. hyopneumoniae field isolates in the swine population from conventional farrow-to-finish farms in Serbia and suggest close genetic relatedness of the corresponding isolates. PMID:20713321
Loop-mediated isothermal amplification (LAMP), a novel method of gene amplification, was employed in this study for detecting Mycoplasma hyopneumoniae in the respiratory tract or lungs of swine. The pathogen can be detected in LAMP reactions containing as few as 10 fg purified target DNA (10 copies of M. hyopneumoniae genome) within 30 min, which was comparable to real-time PCR. After 30-min reaction at 63 °C, the addition of a certain amount of dye (SYBR Green I and hydroxyl naphthol blue at a proper ratio) into the LAMP reaction system makes the results easily determined as positive or negative by visual inspection. In addition, the LAMP was able to distinguish between M. hyopneumoniae and other closely-related mycoplasmastrains, indicating a high degree of specificity. The LAMP assay was more simple and cheap, since the reaction could be completed under isothermal conditions and less laboratorial infrastructure are required. And, it was proven reliable for M. hyopneumoniae diagnosis of nasal swab and lung samples from the field. PMID:23184577
Li, Jiahe; Minion, F Chris; Petersen, Andrew C; Jiang, Fei; Yang, Sheng; Guo, Panpan; Li, Jinxiang; Wu, Wenxue
The genome of Mycoplasma arthritidis strain 158 has modified cytosine residues at AGCT sequences that render the DNA resistant to digestion with the AluI restriction endonuclease. The DNA methyltransferase responsible for the base modification has previously been designated MarI. From the complete genome sequence of M. arthritidis, we identify Marth_orf138 as a candidate marI gene. Marth_orf138 was cloned in Escherichia coli and its TGA codons converted to TGG. DNA isolated from E. coli cells expressing the modified Marth_orf138 gene was degraded by the AluI nuclease, indicating that Marth_orf138 does not code for MarI. However, the DNA from E. coli was found to have acquired resistance to the restriction endonuclease HhaI. Genomic DNA from M. arthritidis was also found to be resistant to HhaI (recognizes GCGC). The M. arthritidis isoschizomer of the HhaI DNA methyltransferase, coded by Marth_orf138, is designated MarII. Transformation of M. arthritidis was not significantly affected by modification of plasmid at HhaI sites, indicating that the mycoplasma lacks a restriction endonuclease that recognizes GCGC sites.
In order to develop a species-specific PCR for the detection of Mycoplasma genitalium, the sequence of 1,490 bases of the 16S rRNA gene was determined for M. genitalium G37 (type strain) and four Danish isolates of M. genitalium. The sequences of the four Danish strains, mutually different with respect to their MgPa gene, were 100% homologous, although they carried a single common base substitution compared to the type strain. Among members of the Mycoplasma pneumoniae phylogenetic cluster, M. genitalium showed the most-prominent homology to the 16S rRNA sequence of M. pneumoniae (98% homology). From regions showing the least homology to the M. pneumoniae 16S rRNA gene sequence, primers were chosen to amplify DNA from M. genitalium only. Two sets of primers were selected for their ability to detect <10 to 50 M. genitalium genome copies without cross-reactions with M. pneumoniae. The performance of these primers was compared to the performance of two pairs of primers amplifying parts of the MgPa adhesin gene; 1,030 randomly selected specimens submitted for Chlamydia trachomatis culture were screened with one of the 16S rRNA gene primer sets. A total of 41 specimens were found to be positive for this gene; 40 of these could be confirmed by one of the MgPa primer sets, whereas the other MgPa primer set detected only 21 positive specimens out of 40. These results indicate that estimates of the prevalence of M. genitalium in various populations using MgPa PCR primers could be incorrectly low if the PCR primers are located in variable regions of the MgPa gene.
Jensen, J?rgen Skov; Borre, Martin B.; Dohn, Birthe
Mycoplasmas are cell wall-less bacteria that evolved by drastic reduction of the genome size. Complete genome analysis of Mycoplasma pneumoniae revealed the presence of numerous copies of four distinct large M. pneumoniae repetitive elements (RepMPs). One copy each of RepMP2/3, RepMP4, and RepMP5 are localized within the P1 operon (MPN140 to MPN142 loci), and their involvement in sequence variation in adhesin P1 and adherence-related protein B/C has been documented. Here we analyzed a clinical strain of M. pneumoniae designated S1 isolated from a 1993 outbreak of respiratory infections in San Antonio, TX. Based on the type of RepMPs within the P1 operon, we classified clinical isolate S1 as type 2 with unique minor sequence variations. Hybridization with oligonucleotide arrays revealed sequence divergence in two previously unsuspected hypothetical genes (MPN137 and MPN138 loci). Closer inspection of this region revealed that the MPN137 and MPN138 loci harbored previously unrecognized unique RepMP1 sequences found only in M. pneumoniae. PCR and sequence analyses revealed a recombination event involving three RepMP1-containing genes that resulted in fusion of MPN137 and MPN138 reading frames and loss of all but a short fragment of another RepMP1-containing locus, MPN130. The multiple copies of unique RepMP1 elements spread throughout the chromosome could allow vast numbers of sequence variations in clinical strains. Comparisons of amino acid sequences showed the presence of leucine zipper motifs in MPN130 and MPN138 proteins in reference strain M129 and the absence of these motifs in the fused protein of S1. The presence of tandem leucine and other repeats points to possible regulatory functions of proteins encoded by RepMP1-containing genes.
Musatovova, Oxana; Kannan, T. R.; Baseman, Joel B.
Mycoplasma gallisepticum (MG) has caused an endemic upper respiratory and ocular infection in the eastern house finch (Carpodacus mexicanus) after the epidemic first described in 1994. The disease has been studied by a number of investigators at a population level and reports describe experimental infection in group-housed MG-free house finches. Because detailed observation and evaluation of individual birds in group-housed passerines is problematic, we studied individually housed house finches that were experimentally inoculated with the finch strain of MG in a controlled environment. To accomplish this, a study was conducted spanning the period of November 2001-April 2002 with 20 MG-free (confirmed by the rapid plate agglutination assay and polymerase chain reaction [PCR] assay) eastern house finches captured in the Cayuga Basin area of central New York (USA) in the summer of 2001. After a period of acclimatization and observation (12 wk), 20 finches were inoculated with a 0.05-ml aliquot of MG (3.24 x 10(5) colony-forming units/ml) via bilateral conjunctival sac instillations. Two additional finches acted as controls and were inoculated in the same manner with preservative-free sterile saline solution. After inoculation, all finches except the controls exhibited clinical signs of conjunctivitis within 2-6 days. The progression of the disease was evaluated by several methods, including PCR, behavioral observations, and physical examination including eye scoring, body weight, and body condition index. Over a period of 21 wk, MG-infected finches developed signs of disease and recovered (80%), developed signs of disease and progressed to become chronically infected (15%), or died (5%). We hypothesize that the high survival rate and recovery of these finches after infection was associated with the use of controlled environmental conditions, acclimatization, a high plane of nutrition, and low stocking (housing) density, all of which are factors documented to be important in the outcome of MG infections in domestic poultry and other species. PMID:15137491
Kollias, George V; Sydenstricker, Keila V; Kollias, Heidi W; Ley, David H; Hosseini, Parviez R; Connolly, Véronique; Dhondt, André A
Background Increasing evidence incriminates bacteria, especially Mycoplasma fermentans, as possible arthritogenic agents in humans. The purpose of this study was to investigate M. fermentans in the bloodstream of patients with rheumatoid arthritis. Methods Two hundred and nineteen blood samples from patients with rheumatoid arthritis, systemic lupus erythematosus, antiphospholipid syndrome, and healthy individuals were screened by bacterial culture and direct PCR in order to detect mycoplasmas; IgM and IgG against M. fermentans PG18 were also detected by ELISA and Immunoblotting assays in patients with rheumatoid arthritis and healthy individuals. Results Blood samples from patients with antiphospholipid syndrome and healthy individuals were negative for mycoplasma by culture or direct PCR. In blood samples from patients with systemic lupus erythematosus were detected by direct PCR M. fermentans in 2/50 (2%), M. hominis in 2/50 (2%) and U. urealyticum in 1/50 (0.5%). In patients with RA M. fermentans was detected by culture in 13/87 blood samples and in 13/87 by direct PCR, however, there was only concordance between culture and direct PCR in six samples, so M. fermentans was detected in 20/87(23%) of the blood samples from patients with RA by either culture or PCR. Antibody-specific ELISA assay to M. fermentans PG18 was done, IgM was detected in sera from 40/87 patients with RA and in sera of 7/67 control individuals, IgG was detected in sera from 48/87 RA patients and in sera from 7/67 healthy individuals. Antibody-specific immunoblotting to M. fermentans PG18 showed IgM in sera from 35/87 patients with RA and in sera from 4/67 healthy individuals, IgG was detected in sera from 34/87 patients and in sera from 5/67 healthy individuals. Conclusion Our findings show that only M. fermentans produce bacteremia in a high percentage of patients with RA. This finding is similar to those reported in the literature. IgM and IgG against M. fermentans PG18 were more frequent in patients with RA than healthy individuals.
The DNA recombination and repair machineries of Mycoplasma genitalium and Mycoplasma pneumoniae differ considerably from those of gram-positive and gram-negative bacteria. Most notably, M. pneumoniae is unable to express a functional RecU Holliday junction (HJ) resolvase. In addition, the RuvB homologues from both M. pneumoniae and M. genitalium only exhibit DNA helicase activity but not HJ branch migration activity in vitro. To identify a putative role of the RuvA homologues of these mycoplasmas in DNA recombination, both proteins (RuvA(Mpn) and RuvA(Mge), respectively) were studied for their ability to bind DNA and to interact with RuvB and RecU. In spite of a high level of sequence conservation between RuvA(Mpn) and RuvA(Mge) (68.8% identity), substantial differences were found between these proteins in their activities. First, RuvA(Mge) was found to preferentially bind to HJs, whereas RuvA(Mpn) displayed similar affinities for both HJs and single-stranded DNA. Second, while RuvA(Mpn) is able to form two distinct complexes with HJs, RuvA(Mge) only produced a single HJ complex. Third, RuvA(Mge) stimulated the DNA helicase and ATPase activities of RuvB(Mge), whereas RuvA(Mpn) did not augment RuvB activity. Finally, while both RuvA(Mge) and RecU(Mge) efficiently bind to HJs, they did not compete with each other for HJ binding, but formed stable complexes with HJs over a wide protein concentration range. This interaction, however, resulted in inhibition of the HJ resolution activity of RecU(Mge). PMID:22666500
Sluijter, Marcel; Estevăo, Silvia; Hoogenboezem, Theo; Hartwig, Nico G; van Rossum, Annemarie M C; Vink, Cornelis
PCR-based diagnostic tests using oligonucleotides specific to 16S rRNA were designed for the specific detection of the turkey pathogens Mycoplasma meleagridis and M. iowae. This method of detection was shown to be rapid, species specific, and unaffected by strain variation or the presence of other organisms. Detection of M. meleagridis in clinical samples by PCR was achieved and later confirmed by culture and growth inhibition. Definitive identification by culture and growth inhibition required up to 3 weeks, whereas positive results from PCR testing were obtained within a day and negative samples were confirmed within 4 days.
A high percentage of cell lines are chronically infected with various mycoplasma species. The addition of antibiotics that are particularly effective against these contaminants to the culture medium during a limited period of time is a simple, inexpensive, and very practical approach for decontaminating cell cultures. Here, we examined the effectiveness of the new antimycoplasma compound Plasmocin that has been employed routinely to cleanse chronically infected cell lines. In a first round of treatment 45 out of 58 (78%) mycoplasma-positive cell lines could be cured. In a second attempt using back-up cryopreserved original cells, four additional cell lines were cured; thus, the overall cure rate was 84%. Even if the mycoplasma contamination was not eradicated by Plasmocin, the parallel treatment with several other antibiotics (Baytril, BM-Cyclin, Ciprobay, MRA, or MycoZap) led to the cure of all 58 cell lines. The successful decontamination was permanent as mycoplasmas were no longer detected at day +14 posttreatment and at later time points as examined by PCR which is the most sensitive and specific mycoplasma detection method. Collectively, our results highlight certain antibiotics as effective antimycoplasma reagents and support the therapeutic rationale for their use in the eradication of this notorious cell culture contaminant.
Uphoff, Cord C.; Denkmann, Sabine-A.; Drexler, Hans G.
A latex agglutination test (LAT) has been developed for the diagnosis of contagious bovine pleuropneumonia (CBPP). The latex microspheres were coated with MmmSC polyclonal immunoglobulin G antiserum and detected MmmSC antigen in the serum of cattle infected with CBPP and in growth medium containing MmmSC. The specific antigen recognizsed by this test appeared to be the capsular polysaccharide (CPS). The LAT recognized all 23 strains of MmmSC examined in this study, with a sensitivity level of 2 ng of CPS, or the equivalent of 5 × 103 CFU, in a reaction volume of 0.03 ml. Therefore, rapid identification of MmmSC cultures should be possible. Agglutination was also observed with the related goat pathogens and “Mycoplasma mycoides” cluster members Mycoplasma mycoides subsp. mycoides large colony biotype (four of six strains positive) and Mycoplasma mycoides subsp. capri (three of six strains positive), in agreement with the suggestion that these latter two mycoplasmas may in fact represent a single species (although collectively exhibiting two capsular serotypes). Comparisons in diagnosis with the complement fixation test (CFT) were made by using African field sera from CBPP-infected cattle. After 2 (or 3) min of incubation, the test detected 55% (or 61%) of CFT-positive sera and 29% (or 40%) of CFT-negative sera, with an overall correlation in diagnosis of 62% (or 61%). The rates for false-positive diagnoses made by using “known” CBPP-negative sera from the United Kingdom were 3 or 13% after 2 or 3 min of incubation, respectively. The data agree with previous findings that some CBPP CFT-negative misdiagnoses may occur due to “antibody eclipsing” by excess circulating antigen. The LAT combines low cost and high specificity with ease of application in the field, without the need for any specialist training or equipment.
Few mycoplasmal polysaccharides have been described and little is known about their role in pathogenesis. The infection of mice with Mycoplasma pulmonis has been utilized in many in vivo and in vitro studies to gain a better understanding of host-pathogen interactions during chronic respiratory infection. Although alveolar macrophages have a primary role in host defence, M. pulmonis is killed inefficiently in vitro. One antiphagocytic factor produced by the mycoplasma is the family of phase- and size-variable Vsa lipoproteins. However, bacteria generally employ multiple strategies for combating host defences, with capsular polysaccharide often having a key role. We show here that mutants lacking the EPS-I polysaccharide of M. pulmonis exhibit increased susceptibility to binding and subsequent killing by alveolar macrophages. These results give further insight into how mycoplasmas are able to avoid the host immune system and sustain a chronic infection. PMID:23190331
Shaw, Brandon M; Daubenspeck, James M; Simmons, Warren L; Dybvig, Kevin
Crude lipoprotein-containing fractions obtained from sera of three different animal species were tested, in combination with bovine serum in Mycoplasma pneumoniae culture medium. All sera yielded at least one lipoprotein-containing component which was considerably more effective in promoting mycoplasma growth than the unfractionated serum sample from which it was derived. The very low activity of certain whole-serum samples tested in this investigation suggests that toxic substances may be present in whole serum which are not contained in the lipoprotein preparations. The greatest activity appeared in the high-density lipoprotein-containing components of bovine and horse sera and the low-density lipoprotein-containing components of human serum. The high degree of growth-supporting activity of these crude lipoprotein-containing serum components suggests that they may be useful as serum substitutes in mycoplasma culture media.
Experimental arthritis in rabbits was induced by M. pneumoniae. We compared it with the arthritis produced by well known animal arthritogenic agents (M. pulmonis and M. arthritidis). Mycoplasmas were detected in the knee joint by different techniques. M. pneumoniae and M. pulmonis produced an acute arthritis that resolved in 2 weeks while, M. arthritidis produced a chronic arthritis. Live M. pneumoniae and M. pulmonis were recovered from the joint during all the experiments. No live M. arthritidis was detected. Live mycoplasmas play an important role in acute arthritis. A similar inflammatory pattern was shown by M. pneumoniae and M. pulmonis. This animal model could be helpful in the study of arthritis induced by a human pathogen mycoplasma. PMID:1578446
Background & Objective: Viral respiratory infections (VRI) have been commonly associated with exacerbation of wheezing in asthmatic children. Mycoplasma pneumoniae (MP) causes many respiratory syndromes that clinically mimic VRI. Due to the nature of the organism, cultures are of no practical value and the diagnosis is usually made by serology. Only a few studies have associated mycoplasma infections with acute exacerbations of wheezing in the asthmatic patient. This study was an attempt to assess the incidence of recent mycoplasma infections in patients with status asthmaticus and to review their laboratory, clinical and radiological findings. Methods: Retrospective review of all patients admitted to PICU over 12 month period with status asthmaticus. Recent mycoplasma infection was determined utilizing the Immunocard Mycoplasma Enzyme Immunoassay (EIA) for detection of MP IgM antibodies (Meredian Diagnostics, Inc., Cincinnati, OH) Results; The records of 44 patients were reviewed. 9 were excluded because MP tests were never obtained during hospitalization. 15/35 (42%) were MP Positive. There were no statistically significant differences (P>0.05) in length of hospitalization (LOH), ICU days, duration of continuous albuterol aerosol hours (cont. Nebs hrs.), days on O2 (02 days) or WBC between the two groups, however patients who were mycoplasma positive were treated with a macrolide antibiotic in addition to their standard asthma therapy. Patients with evidence of recent MP infection were more likely to have one or more infiltrates on their CXR (13/15 vs 7/20; P= 0.002). Conclusion: Our study suggests that recent MP infections play a significant role in exacerbations of asthma and occurrence of status asthmaticus in children. The presence of infiltrates on CXR in status asthmaticus warrants tests for MP.
This article describes the first detection of Cytauxzoon felis, using molecular techniques, in a naturally infected domestic cat from Brazil, South America. Coinfection with 'Candidatus Mycoplasma haemominutum' was also found. The molecular identification of the piroplasmid species was performed by Polymerase Chain Reaction (PCR) and sequencing analysis. A 284 pb fragment of the gene encoding the 18S ribosomal RNA region was amplified and showed 99% identity with other C. felis strains from North America. In addition, PCR-RFLP (restriction fragment length polymorphism) analysis, which amplifies a 595 bp fragment of the gene encoding 16S ribosomal RNA of some bacterial species, identified the co-infecting species as 'Candidatus M. haemominutum'. PMID:23856727
Maia, Leticia Mendes Pupio; Cerqueira, Aloysio de Mello Figueiredo; de Barros Macieira, Daniel; de Souza, Aline Moreira; Moreira, Namir Santos; da Silva, Adrianna Vieira; Messick, Joanne Belle; Ferreira, Renata Fernandes; Almosny, Nádia Regina Pereira
In the bacterial world, methylation is most commonly associated with restriction-modification systems that provide a defense mechanism against invading foreign genomes. In addition, it is known that methylation plays functionally important roles, including timing of DNA replication, chromosome partitioning, DNA repair, and regulation of gene expression. However, full DNA methylome analyses are scarce due to a lack of a simple methodology for rapid and sensitive detection of common epigenetic marks (ie N6-methyladenine (6 mA) and N4-methylcytosine (4 mC)), in these organisms. Here, we use Single-Molecule Real-Time (SMRT) sequencing to determine the methylomes of two related human pathogen species, Mycoplasma genitalium G-37 and Mycoplasma pneumoniae M129, with single-base resolution. Our analysis identified two new methylation motifs not previously described in bacteria: a widespread 6 mA methylation motif common to both bacteria (5?-CTAT-3?), as well as a more complex Type I m6A sequence motif in M. pneumoniae (5?-GAN7TAY-3?/3?-CTN7ATR-5?). We identify the methyltransferase responsible for the common motif and suggest the one involved in M. pneumoniae only. Analysis of the distribution of methylation sites across the genome of M. pneumoniae suggests a potential role for methylation in regulating the cell cycle, as well as in regulation of gene expression. To our knowledge, this is one of the first direct methylome profiling studies with single-base resolution from a bacterial organism.
Llorens-Rico, Veronica; Delgado, Javier; Fang, Gang; Spittle, Kristi; Clark, Tyson A.; Schadt, Eric; Turner, Stephen W.; Korlach, Jonas; Serrano, Luis
Mycoplasma genitalium is an important sexually transmitted pathogen that affects both men and women. In genital-mucosal tissues, it initiates colonization of epithelial cells by attaching itself to host cells via several identified bacterial ligands and host cell surface receptors. We have previously shown that a mutant form of M. genitalium lacking methionine sulfoxide reductase A (MsrA), an antioxidant enzyme which converts oxidized methionine (Met(O)) into methionine (Met), shows decreased viability in infected animals. To gain more insights into the mechanisms by which MsrA controls M. genitalium virulence, we compared the wild-type M. genitalium strain (G37) with an msrA mutant (MS5) strain for their ability to interact with target cervical epithelial cell lines (HeLa and C33A) and THP-1 monocytic cells. Infection of epithelial cell lines with both strains revealed that MS5 was less cytotoxic to HeLa and C33A cell lines than the G37 strain. Also, the MS5 strain was more susceptible to phagocytosis by THP-1 cells than wild type strain (G37). Further, MS5 was less able to induce aggregation and differentiation in THP-1 cells than the wild type strain, as determined by carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling of the cells, followed by counting of cells attached to the culture dish using image analysis. Finally, MS5 was observed to induce less proinflammatory cytokine TNF-? by THP-1 cells than wild type G37 strain. These results indicate that MsrA affects the virulence properties of M. genitalium by modulating its interaction with host cells. PMID:22558404
Das, Kishore; De la Garza, Georgina; Maffi, Shivani; Saikolappan, Sankaralingam; Dhandayuthapani, Subramanian
Four genetic Mycoplasma gallisepticum (MG) polymerase chain reactions (PCRs) (16s rRNA PCR, three newly developed PCR methods that target surface protein genes [mgc2, LP (nested) and gapA (nested)]) were compared for analytical specificity and sensitivity and for diagnostic sensitivity (Se) and specificity of detection from tracheal swabs. The licensed MG DNA Test Kit Flock Chek test (IDEXX, Laboratories, Inc., Westbrook, ME) was as well evaluated for the diagnostic specificity and sensitivity of detection from tracheal swabs. Analytical specificity was evaluated for the four generic PCR methods using a panel of DNA samples from microorganisms that may be isolated from the trachea of commercial poultry and other fowl. PCR methods mgc2, nLP, and ngapA only amplified DNA from MG, whereas 16S rRNA PCR amplified DNA from MG and Mycoplasma imitans. The analytical sensitivity of the four generic PCR methods expressed in color-changing units (CCU)/amplification reaction was estimated for each PCR method and ranged from 4 to 400 CCU/reaction; the sensitivities of single PCR methods 16S rRNA and mgc2 were estimated at 40 CCU/reaction, the nLP at 400 CCU/reaction, and the ngapA at 4 CCU/reaction. The diagnostic sensitivity and specificity of MG detection from tracheal swab pools, as compared to isolation from choanal cleft swabs, was evaluated for the five PCR methods using three groups of birds exposed to vaccine strains ts-11 and 6/85 and to challenge strain R. All PCR methods were able to detect the vaccine strains and the challenge strain R directly from tracheal swabs, indicating that PCR primers from the different methods amplified divergent MG strains. Isolation and PCR results correlated satisfactorily among the three experimentally infected groups, with agreement values (k) ranging from 0.52 to 1.00. The ngapA, IDEXX, and mgc2 PCRs showed the best sensitivity (Se) ratios for detection of M. gallisepticum strains as compared to isolation. Compared to the ngapA and IDEXX PCR methods, the mgc2 PCR has a faster turnaround time, since this test consists of a single amplification reaction and the amplification product is detected by gel electrophoresis. Therefore, among the PCR methods evaluated in this study, the mgc2 PCR is the method of choice to further validate in the field. PMID:15839425
García, Maricarmen; Ikuta, Nilo; Levisohn, Sharon; Kleven, S H
Mycoplasma synoviae is a poultry pathogen causing respiratory disease and synovitis. A number of serological assays have been developed for diagnosis of M. synoviae infection; however, they lack sensitivity and/or are prone to false-positive reactions. Using a combination of PCR and expression cloning, four overlapping regions (regions 1-4) of the surface antigen MSPB of M. synoviae WVU-1853 were expressed in a bacterial expression system. Immunostaining of the resultant polypeptides with chicken sera raised against different M. synoviae strains, or Mycoplasma gallisepticum S6, suggested that region 4 contained a highly antigenic and species-specific domain (amino acids 212-317) [corrected] of MSPB. A fusion protein of region 4 was expressed in the pMAL expression system and purified from cold-osmotic-shock fluids of Escherichia coli cells for use in an indirect ELISA. The potential of the purified antigen for detection of M. synoviae antibodies was assessed with sera obtained from chickens experimentally infected with different strains of M. synoviae or M. gallisepticum, or from uninoculated chickens. All the sera from M. synoviae-inoculated chickens provided higher absorbance values than those from M. gallisepticum-inoculated or uninoculated chickens. Chickens inoculated with M. synoviae 86079/7NS had detectable increases of serum anti-MSPB immunoglobulins at day 7 after inoculation. These studies have identified the most antigenic region of one of the major species-specific surface proteins of M. synoviae, and shown the potential of this protein as a serodiagnostic reagent. PMID:10463175
Noormohammadi, A H; Markham, P F; Markham, J F; Whithear, K G; Browning, G F
Mycoplasma genitalium is associated with acute and chronic urethritis in men. Existing data on infection in women are limited and inconsistent but suggest that M. genitalium is associated with urethritis, cervicitis, pelvic inflammatory disease, and possibly female infertility. Data are inconclusive regarding the role of M. genitalium in adverse pregnancy outcomes and ectopic pregnancy. Available data suggest that azithromycin is superior to doxycycline in treating M. genitalium infection. However, azithromycin-resistant infections have been reported in 3 continents, and the proportion of azithromycin-resistant M. genitalium infection is unknown. Moxifloxacin is the only drug that currently seems to uniformly eradicate M. genitalium. Detection of M. genitalium is hampered by the absence of a commercially available diagnostic test. Persons with persistent pelvic inflammatory disease or clinically significant persistent urethritis or cervicitis should be tested for M. genitalium, if possible. Infected persons who have not previously received azithromycin should receive that drug. Persons in whom azithromycin therapy fails should be treated with moxifloxicin. PMID:22080266
Manhart, Lisa E; Broad, Jennifer M; Golden, Matthew R
Mycoplasma gallisepticum, a cell wall-less bacterium, must confront the problem of colloid osmotic swelling. Cell volume was determined by optical density and intracellular water measurements. Transmembrane pH and electrical gradients were determined by the distribution of the weak acid benzoate and lipophilic cation tetraphenylphosphonium respectively. Cells incubated in sodium chloride without glucose exhibited a progressive fall in ATP over several hours. When ATP fell below 40 uM the cells swelled, leaked protein and became permeable to inulin. Subsequent addition of glucose induced shrinkage and restored the original permeability properties. Energized cells exhibited an electrochemical gradient of protons of up to 130 mV, inside negative and alkaline. The proton-ATPase inhibitor dicyclohexylcarbodiimide (DCCD), which collapsed the chemical and electrical components of the proton gradient, induced rapid swelling despite high ATP levels thus implicating the proton gradient in volume regulation. Either the pH gradient or the membrane potential could maintain volume. Energy-dependent sodium efflux in exchange for protons was demonstrated in sodium-loaded cells using radioactive sodium and 9-aminoacridine fluorescence to follow sodium and proton translocation respectively.
Mycoplasma genitalium is a human pathogen associated with several sexually transmitted diseases. Proteomic technologies, along with other methods for global gene expression analysis, play a key role in understanding the mechanisms of bacterial pathogenesis and physiology. The proteome of M. genitalium, model of a minimal cell, has been extended using a combination of different proteomic approaches and technologies. The total proteome of this microorganism has been analyzed using gel-based and gel-free approaches, achieving the identification of 85.3% of the predicted ORFs. In addition, a comprehensive analysis of membrane subproteome has been performed. For this purpose, the TX-114 soluble fraction has been analyzed as well as the surface proteins, using cell-surface protein labeling with CyDye. Finally, the serological response of M. genitalium-infected patients and healthy donors has been analyzed to identify proteins that trigger immunological response. Here, we present the most extensive M. genitalium proteome analysis (85.3% of predicted ORFs), a comprehensive M. genitalium membrane analysis, and a study of the human serological response to M. genitalium. PMID:22582988
Párraga-Nińo, Noemí; Colomé-Calls, Nuria; Canals, Francesc; Querol, Enrique; Ferrer-Navarro, Mario
An ongoing outbreak of conjunctivitis in free-ranging house finches (Carpodacus mexicanus) began in 1994 in the eastern United States. Bacterial organisms identified as Mycoplasma gallisepticum (MG) were isolated from lesions of infected birds. MG was also isolated from a blue jay (Cyanocitta cristata) that contracted conjunctivitis after being housed in a cage previously occupied by house finches with conjunctivitis, and from free-ranging American goldfinches (Carduelis tristis) in North Carolina in 1996. To investigate the molecular epidemiology of this outbreak, we produced DNA fingerprints of MG isolates by random amplification of polymorphic DNA (RAPD). We compared MG isolates from songbirds examined from 1994 through 1996 in 11 states, representing three host species, with vaccine and reference strains and with contemporary MG isolates from commercial poultry. All MG isolates from songbirds had RAPD banding patterns identical to each other but different from other strains and isolates tested. These results indicate that the outbreak of MG in songbirds is caused by the same strain, which suggests a single source; the outbreak is not caused by the vaccine or reference strains analyzed; and MG infection has not been shared between songbirds and commercial poultry. PMID:9284386
An ongoing outbreak of conjunctivitis in free-ranging house finches (Carpodacus mexicanus) began in 1994 in the eastern United States. Bacterial organisms identified as Mycoplasma gallisepticum (MG) were isolated from lesions of infected birds. MG was also isolated from a blue jay (Cyanocitta cristata) that contracted conjunctivitis after being housed in a cage previously occupied by house finches with conjunctivitis, and from free-ranging American goldfinches (Carduelis tristis) in North Carolina in 1996. To investigate the molecular epidemiology of this outbreak, we produced DNA fingerprints of MG isolates by random amplification of polymorphic DNA (RAPD). We compared MG isolates from songbirds examined from 1994 through 1996 in 11 states, representing three host species, with vaccine and reference strains and with contemporary MG isolates from commercial poultry. All MG isolates from songbirds had RAPD banding patterns identical to each other but different from other strains and isolates tested. These results indicate that the outbreak of MG in songbirds is caused by the same strain, which suggests a single source; the outbreak is not caused by the vaccine or reference strains analyzed; and MG infection has not been shared between songbirds and commercial poultry.
Forty-two strains of Mycoplasmahominis (including PG21), 2 strain of Mycoplasma fermentans (Pg18 and K7), 1 strain of Mycoplasma pneumoniae (strain m129) were investigated for their susceptibilities to Citrus bergamia essential oil and to its major components (limonene, linalyl acetate and linalool). C. bergamia essential oil inhibited mycoplasmas at concentrations from 0.5 to 1% (MIC value as % v/v). M. hominis showed MIC(50) values of 0.5% and MIC(90) values of 1%; M. pneumoniae showed a MIC value of 0.5% while M. fermentans strains were inhibited by MIC values of 1%. M. pneumoniae and M. hominis shared the same susceptibility to linalyl acetate, with MIC values of 0.015% (corresponding to MIC(50) and MIC(90) for M. hominis); M. fermentans strains were less susceptible with MIC values of 0.12%. Among the major components tested, linalool showed higher activity against M. pneumoniae and M. fermentans (MIC values of 0.015 and 0.06%, respectively) but was less active against M. hominis (MIC(50) and MIC(90) values of both 1%); limonene was active against M. pneumoniae (MIC value of 0.03%) but was less active against M. fermentans (MIC values of 1%) and M. hominis (both MIC(50) and MIC(90) values of ?4%). The results indicated that C. bergamia essential oil and its major components had shown an interesting in vitro antimycoplasmal activity. PMID:22465092
Furneri, Pio Maria; Mondello, Luigi; Mandalari, Giuseppina; Paolino, Donatella; Dugo, Paola; Garozzo, Adriana; Bisignano, Giuseppe
Three hemotropic mycoplasmas have been identified in pet cats: Mycoplasma haemofelis, "Candidatus Mycoplasma haemominutum," and "Candidatus Mycoplasma turicensis." The way in which these agents are transmitted is largely unknown. Thus, this study aimed to investigate fleas, ticks, and rodents as well as saliva and feces from infected cats for the presence of hemotropic mycoplasmas, to gain insight into potential transmission routes for these agents. DNA was extracted from arthropods and from rodent blood or tissue samples from Switzerland and from salivary and fecal swabs from two experimentally infected and six naturally infected cats. All samples were analyzed with real-time PCR, and some positive samples were confirmed by sequencing. Feline hemotropic mycoplasmas were detected in cat fleas and in a few Ixodes sp. and Rhipicephalus sp. ticks collected from animals but not in ticks collected from vegetation or from rodent samples, although the latter were frequently Mycoplasma coccoides PCR positive. When shedding patterns of feline hemotropic mycoplasmas were investigated, "Ca. Mycoplasma turicensis" DNA was detected in saliva and feces at the early but not at the late phase of infection. M. haemofelis and "Ca. Mycoplasma haemominutum" DNA was not amplified from saliva and feces of naturally infected cats, despite high hemotropic mycoplasma blood loads. Our results suggest that besides an ostensibly indirect transmission by fleas, direct transmission through saliva and feces at the early phase of infection could play a role in the epizootiology of feline hemotropic mycoplasmas. Neither the investigated tick nor the rodent population seems to represent a major reservoir for feline hemotropic mycoplasmas in Switzerland. PMID:17468284
Variable-number tandem-repeat (VNTR) analysis has been developed for the causative agent of contagious bovine pleuropneumonia using the genome of Mycoplasma mycoides subspecies mycoides small colony (M.m.m. SC) PG1. Genome analysis identified 60 VNTRs within the M.m.m. SC genome; however, screening of these VNTRs with a panel of strains identified only three VNTRs that gave allelic variation. Testing of three VNTRs against 39 strains of diverse geographical origin gave 12 different VNTR profiles groups. VNTR analysis may represent a new rapid tool for subtyping M.m.m. SC isolates. PMID:17956424
McAuliffe, Laura; Ayling, Roger D; Nicholas, Robin A J
A case of furuncular myiasis was reported for the first time in a 29-year-old young Taiwanese traveler returning from an ecotourism in Peru. Furuncle-like lesions were observed on the top of his head and he complained of crawling sensations within his scalp. The invasive larva of botfly, Dermatobia hominis, was extruded from the furuncular lesion of the patient. Awareness of cutaneous myiasis for clinicians should be considered for a patient who has a furuncular lesion and has recently returned from a botfly-endemic area. PMID:23620844
Hu, Je-Ming; Wang, Chih-Chien; Chao, Li-Lian; Lee, Chung-Shinn; Shih, Chien-Ming; Telford, Sam R
The p36 protein of Mycoplasma hyopneumoniae is a cytosolic protein carrying species-specific antigenic determinants. Based on the genomic sequence of the reference strain ATCC 25934, primers were designed for PCR amplification of the p36-encoding gene (948 bp). These primers were shown to be specific to M. hyopneumoniae since no DNA amplicons could be obtained with other mycoplasma species and pathogenic bacteria that commonly colonize the porcine respiratory tract. The amplified p36 gene was subcloned into the pGEX-4T-1 vector to be expressed in Escherichia coli as a fusion protein with glutathione S-transferase (GST). The GST-p36 recombinant fusion protein was purified by affinity chromatography and cut by thrombin, and the enriched p36 protein was used to immunize female BALB/c mice for the production of anti-p36 monoclonal antibodies (MAbs). The polypeptide specificity of the nine MAbs obtained was confirmed by Western immunoblotting with cell lysates prepared from the homologous strain. Cross-reactivity studies of the anti-p36 MAbs towards two other M. hyopneumoniae reference strains (ATCC 25095 and J strains) and Quebec field strains that had been isolated in culture suggested that these anti-p36 MAbs were directed against a highly conserved epitope, or closely located epitopes, of the p36 protein. No reactivity was demonstrated against other mycoplasma species tested. Clinical signs and lesions suggestive of enzootic pneumonia were reproduced in specific-pathogen-free pigs infected experimentally with a virulent Quebec field strain (IAF-DM9827) of M. hyopneumoniae. The bacteria could be recovered from lung homogenates of pigs that were killed after the 3-week observation period by both PCR and cultivation procedures. Furthermore, the anti-p36 MAbs permitted effective detection by indirect immunofluorescence of M. hyopneumoniae in frozen lung sections from experimentally infected pigs. However, attempts to use the recombinant p36 protein as an antigen in an indirect enzyme-linked immunosorbent assay for the detection of antibodies in sera from convalescent pigs showed no correlation with clinical and pathological findings.
Caron, J.; Sawyer, N.; Moumen, B. Ben Abdel; Bouh, K. Cheikh Saad; Dea, S.
A total of 195 Mycoplasma pneumoniae strains were isolated from 2,462 clinical specimens collected between April 2002 and March 2004 from pediatric outpatients with respiratory tract infections. Susceptibilities to six macrolide antibiotics (ML), telithromycin, minocycline, levofloxacin, and sitafloxacin were determined by the microdilution method using PPLO broth. A total of 183 M. pneumoniae isolates were susceptible to all agents and had excellent MIC90s in the following order: 0.00195 ?g/ml for azithromycin and telithromycin, 0.0078 ?g/ml for clarithromycin, 0.0156 ?g/ml for erythromycin, 0.0625 ?g/ml for sitafloxacin, 0.5 ?g/ml for minocycline, and 1 ?g/ml for levofloxacin. Notably, 12 ML-resistant M. pneumoniae strains were isolated from patients with pneumonia (10 strains) or acute bronchitis (2 strains). These strains showed resistance to ML with MICs of ?1 ?g/ml, except to rokitamycin. Transition mutations of A2063G or A2064G, which correspond to A2058 and A2059 in Escherichia coli, in domain V on the 23S rRNA gene in 11 ML-resistant strains were identified. By pulsed-field gel electrophoresis typing, these strains were classified into groups I and Vb, as described previously (A. Cousin-Allery, A. Charron, B. D. Barbeyrac, G. Fremy, J. S. Jensen, H. Renaudin, and C. Bebear, Epidemiol. Infect. 124:103-111, 2000). These findings suggest that excessive usage of MLs acts as a trigger to select mutations on the corresponding 23S rRNA gene with the resultant occurrence of ML-resistant M. pneumoniae. Monitoring ML susceptibilities for M. pneumoniae is necessary in the future.
A total of 195 Mycoplasma pneumoniae strains were isolated from 2,462 clinical specimens collected between April 2002 and March 2004 from pediatric outpatients with respiratory tract infections. Susceptibilities to six macrolide antibiotics (ML), telithromycin, minocycline, levofloxacin, and sitafloxacin were determined by the microdilution method using PPLO broth. A total of 183 M. pneumoniae isolates were susceptible to all agents and had excellent MIC90s in the following order: 0.00195 microg/ml for azithromycin and telithromycin, 0.0078 microg/ml for clarithromycin, 0.0156 microg/ml for erythromycin, 0.0625 microg/ml for sitafloxacin, 0.5 microg/ml for minocycline, and 1 microg/ml for levofloxacin. Notably, 12 ML-resistant M. pneumoniae strains were isolated from patients with pneumonia (10 strains) or acute bronchitis (2 strains). These strains showed resistance to ML with MICs of >or=1 microg/ml, except to rokitamycin. Transition mutations of A2063G or A2064G, which correspond to A2058 and A2059 in Escherichia coli, in domain V on the 23S rRNA gene in 11 ML-resistant strains were identified. By pulsed-field gel electrophoresis typing, these strains were classified into groups I and IIb [corrected] as described previously (A. Cousin-Allery, A. Charron, B. D. Barbeyrac, G. Fremy, J. S. Jensen, H. Renaudin, and C. Bebear, Epidemiol. Infect. 124:103-111, 2000). These findings suggest that excessive usage of MLs acts as a trigger to select mutations on the corresponding 23S rRNA gene with the resultant occurrence of ML-resistant M. pneumoniae. Monitoring ML susceptibilities for M. pneumoniae is necessary in the future. PMID:15917525
Chlamydia pneumoniae, Chlamydia trachomatis and Chlamydia psittaci were detected at low frequencies (<20%) among 69 pulmonary mucosa-associated lymphoid tissue (MALT) lymphomas, 30 other lymphoproliferative disorders (LPD) and 44 non-LPD. The incidence of individual Chlamydiae was generally higher in MALT lymphoma than non-LPD, although not reaching statistical significance. Mycoplasma pneumoniae DNA was not detected.
Chanudet, E; Adam, P; Nicholson, A G; Wotherspoon, A C; Ranaldi, R; Goteri, G; Pileri, S A; Ye, H; Muller-Hermelink, H K; Du, M-Q
Chlamydia pneumoniae, Chlamydia trachomatis and Chlamydia psittaci were detected at low frequencies (<20%) among 69 pulmonary mucosa-associated lymphoid tissue (MALT) lymphomas, 30 other lymphoproliferative disorders (LPD) and 44 non-LPD. The incidence of individual Chlamydiae was generally higher in MALT lymphoma than non-LPD, although not reaching statistical significance. Mycoplasma pneumoniae DNA was not detected. PMID:17876330
Chanudet, E; Adam, P; Nicholson, A G; Wotherspoon, A C; Ranaldi, R; Goteri, G; Pileri, S A; Ye, H; Müller-Hermelink, H K; Du, M-Q
A novel immunoregulatory factor, designated IL-X, is described which has been isolated from Mycoplasma. The subject invention also concerns polynucleotides which encode IL-X. IL-X protein is a growth factor for EBV transformed human B lymphocytes and for murine helper T lymphocytes. Also taught are methods of raising antibodies to IL-X, and cloning of IL-X.
Mycoplasma gallisepticum (MG) has caused an endemic upper respiratory and ocular infection in the eastern house finch (Carpodacus mexicanus) after the epidemic first described in 1994. The disease has been studied by a number of investigators at a population level and reports describe experimental infection in group-housed MG-free house finches. Because detailed observation and evaluation of individual birds in group
George V. Kollias; Keila V. Sydenstricker; Heidi W. Kollias; David H. Ley