Expression of the Arginine Deiminase Pathway Genes in Lactobacillus sakei Is Strain Dependent and Is Affected by the Environmental pH

PubMed Central

Rimaux, T.; Rivière, A.; Illeghems, K.; Weckx, S.; De Vuyst, L.

2012-01-01

The adaptation of Lactobacillus sakei to a meat environment is reflected in its metabolic potential. For instance, the ability to utilize arginine through the arginine deiminase (ADI) pathway, resulting in additional ATP, represents a competitive benefit. In L. sakei CTC 494, the arc operon (arcABCTDR) shows the same gene order and organization as that in L. sakei 23K, the genome sequence of which is known. However, differences in relative gene expression were found, and these seemed to be optimal in different growth phases, namely, the highest relative gene expression level was in the end exponential growth phase in the case of L. sakei CTC 494 and in the mid-exponential growth phase of L. sakei 23K. Also, the environmental pH influenced the relative expression level of the arc operon, as shown for L. sakei CTC 494, with the highest relative expression level occurring at the optimal pH for growth (pH 6.0). Deviations from this optimal pH (pH 5.0 and pH 7.0) resulted in an overall decline of the relative expression level of all genes of the arc operon. Furthermore, a differential relative expression of the individual genes of the arc operon was found, with the highest relative gene expression occurring for the first two genes of the arc operon (arcA and arcB). Finally, it was shown that some L. sakei strains were able to convert agmatine into putrescine, suggesting an operational agmatine deiminase pathway in these strains, a metabolic trait that is undesirable in meat fermentations. This study shows that this metabolic trait is most probably encoded by a previously erroneously annotated second putative arc operon. PMID:22544250

Lactobacillus sakei: A Starter for Sausage Fermentation, a Protective Culture for Meat Products

PubMed Central

Zagorec, Monique; Champomier-Vergès, Marie-Christine

2017-01-01

Among lactic acid bacteria of meat products, Lactobacillus sakei is certainly the most studied species due to its role in the fermentation of sausage and its prevalence during cold storage of raw meat products. Consequently, the physiology of this bacterium regarding functions involved in growth, survival, and metabolism during meat storage and processing are well known. This species exhibits a wide genomic diversity that can be observed when studying different strains and on which probably rely its multiple facets in meat products: starter, spoiler, or protective culture. The emerging exploration of the microbial ecology of meat products also revealed the multiplicity of bacterial interactions L. sakei has to face and their various consequences on microbial quality and safety at the end of storage. PMID:28878171

Primary metabolism in Lactobacillus sakei food isolates by proteomic analysis.

PubMed

McLeod, Anette; Zagorec, Monique; Champomier-Vergès, Marie-Christine; Naterstad, Kristine; Axelsson, Lars

2010-04-22

Lactobacillus sakei is an important food-associated lactic acid bacterium commonly used as starter culture for industrial meat fermentation, and with great potential as a biopreservative in meat and fish products. Understanding the metabolic mechanisms underlying the growth performance of a strain to be used for food fermentations is important for obtaining high-quality and safe products. Proteomic analysis was used to study the primary metabolism in ten food isolates after growth on glucose and ribose, the main sugars available for L. sakei in meat and fish. Proteins, the expression of which varied depending on the carbon source were identified, such as a ribokinase and a D-ribose pyranase directly involved in ribose catabolism, and enzymes involved in the phosphoketolase and glycolytic pathways. Expression of enzymes involved in pyruvate and glycerol/glycerolipid metabolism were also affected by the change of carbon source. Interestingly, a commercial starter culture and a protective culture strain down-regulated the glycolytic pathway more efficiently than the rest of the strains when grown on ribose. The overall two-dimensional gel electrophoresis (2-DE) protein expression pattern was similar for the different strains, though distinct differences were seen between the two subspecies (sakei and carnosus), and a variation of about 20% in the number of spots in the 2-DE gels was observed between strains. A strain isolated from fermented fish showed a higher expression of stress related proteins growing on both carbon sources. It is obvious from the data obtained in this study that the proteomic approach efficiently identifies differentially expressed proteins caused by the change of carbon source. Despite the basic similarity in the strains metabolic routes when they ferment glucose and ribose, there were also interesting differences. From the application point of view, an understanding of regulatory mechanisms, actions of catabolic enzymes and proteins, and

Isolation and identification of a novel bacterium, Lactobacillus sakei subsp. dgh strain 5, and optimization of growth condition for highest antagonistic activity.

PubMed

Tashakor, Amin; Hosseinzadehdehkordi, Mahshid; Emruzi, Zeynab; Gholami, Dariush

2017-05-01

In the present study, we isolated Lactobacillus sakei strain DGH5 from raw beef meat. This bacterium plays an inhibitory effect against food-spoiling bacteria and food-borne pathogens, including Listeria monocytogenes, a gram-positive and pathogenic bacterium. Lactobacillus sakei strain DGH5 was identified through both phenotypical and biochemical tests accompanied with 16S rRNA sequence analysis. Among all the sources of carbon, nitrogen and phosphorous forms, we selected the most potent compounds to optimize the condition for the highest antagonistic activity. Among the sugars, polygalacturonic acid demonstrated to improve the antagonistic activity. Ammonium nitrate demonstrated to be suitable nitrogen sources. Amongst phosphorous sources, disodium hydrogen phosphate had the greatest antagonistic effect. According to Taguchi's orthogonal array, temperature, disodium hydrogen phosphate and soy Peptone had significant effect on antagonistic activity. Furthermore, mean comparisons showed that the optimum conditions achieved at pH 6.0, 25 °C temperature, 1.5% (w/v) Na 2 HPO 4 and 0.5% (w/v) peptone. Copyright © 2017 Elsevier Ltd. All rights reserved.

Analysis of Lactobacillus sakei Mutants Selected after Adaptation to the Gastrointestinal Tracts of Axenic Mice▿ †

PubMed Central

Chiaramonte, Fabrizio; Anglade, Patricia; Baraige, Fabienne; Gratadoux, Jean-Jacques; Langella, Philippe; Champomier-Vergès, Marie-Christine; Zagorec, Monique

2010-01-01

We recently showed that Lactobacillus sakei, a natural meat-borne lactic acid bacterium, can colonize the gastrointestinal tracts (GIT) of axenic mice but that this colonization in the intestinal environment selects L. sakei mutants showing modified colony morphology (small and rough) and cell shape, most probably resulting from the accumulation of various mutations that confer a selective advantage for persistence in the GIT. In the present study, we analyzed such clones, issued from three different L. sakei strains, in order to determine which functions were modified in the mutants. In the elongated filamentous cells of the rough clones, transmission electron microscopy (TEM) analysis showed a septation defect and dotted and slanted black bands, suggesting the presence of a helical structure around the cells. Comparison of the cytoplasmic and cell wall/membrane proteomes of the meat isolate L. sakei 23K and of one of its rough derivatives revealed a modified expression for 38 spots. The expression of six oxidoreductases, several stress proteins, and four ABC transporters was strongly reduced in the GIT-adapted strain, while the actin-like MreB protein responsible for cell shaping was upregulated. In addition, the expression of several enzymes involved in carbohydrate metabolism was modified, which may correlate with the observation of modified growth of mutants on various carbon sources. These results suggest that the modifications leading to a better adaptation to the GIT are pleiotropic and are characterized in a rough mutant by a different stress status, a cell wall modification, and modified use of energy sources, leading to an improved fitness for the colonization of the GIT. PMID:20208026

Primary metabolism in Lactobacillus sakei food isolates by proteomic analysis

PubMed Central

2010-01-01

Background Lactobacillus sakei is an important food-associated lactic acid bacterium commonly used as starter culture for industrial meat fermentation, and with great potential as a biopreservative in meat and fish products. Understanding the metabolic mechanisms underlying the growth performance of a strain to be used for food fermentations is important for obtaining high-quality and safe products. Proteomic analysis was used to study the primary metabolism in ten food isolates after growth on glucose and ribose, the main sugars available for L. sakei in meat and fish. Results Proteins, the expression of which varied depending on the carbon source were identified, such as a ribokinase and a D-ribose pyranase directly involved in ribose catabolism, and enzymes involved in the phosphoketolase and glycolytic pathways. Expression of enzymes involved in pyruvate and glycerol/glycerolipid metabolism were also affected by the change of carbon source. Interestingly, a commercial starter culture and a protective culture strain down-regulated the glycolytic pathway more efficiently than the rest of the strains when grown on ribose. The overall two-dimensional gel electrophoresis (2-DE) protein expression pattern was similar for the different strains, though distinct differences were seen between the two subspecies (sakei and carnosus), and a variation of about 20% in the number of spots in the 2-DE gels was observed between strains. A strain isolated from fermented fish showed a higher expression of stress related proteins growing on both carbon sources. Conclusions It is obvious from the data obtained in this study that the proteomic approach efficiently identifies differentially expressed proteins caused by the change of carbon source. Despite the basic similarity in the strains metabolic routes when they ferment glucose and ribose, there were also interesting differences. From the application point of view, an understanding of regulatory mechanisms, actions of catabolic

Bio-protective potential of lactic acid bacteria: Effect of Lactobacillus sakei and Lactobacillus curvatus on changes of the microbial community in vacuum-packaged chilled beef.

PubMed

Zhang, Yimin; Zhu, Lixian; Dong, Pengcheng; Liang, Rongrong; Mao, Yanwei; Qiu, Shubing; Luo, Xin

2018-04-01

This study was to determine the bacterial diversity and monitor the community dynamic changes during storage of vacuum-packaged sliced raw beef as affected by Lactobacillus sakei and Lactobacillus curvatus . L. sakei and L. curvatus were separately incubated in vacuumed-packaged raw beef as bio-protective cultures to inhibit the naturally contaminating microbial load. Dynamic changes of the microbial diversity of inoculated or non-inoculated (control) samples were monitored at 4°C for 0 to 38 days, using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). The DGGE profiles of DNA directly extracted from non-inoculated control samples highlighted the order of appearance of spoilage bacteria during storage, showing that Enterbacteriaceae and Pseudomonas fragi emerged early, then Brochothrix thermosphacta shared the dominant position, and finally, Pseudomonas putida showed up became predominant. Compared with control, the inoculation of either L. sakei or L. curvatus significantly lowered the complexity of microbial diversity and inhibited the growth of spoilage bacteria (p<0.05). Interestingly, we also found that the dominant position of L. curvatus was replaced by indigenous L. sakei after 13 d for L. curvatus -inoculated samples. Plate counts on selective agars further showed that inoculation with L. sakei or L. curvatus obviously reduced the viable counts of Enterbacteraceae , Pseudomonas spp. and B. thermosphacta during later storage (p< 0.05), with L. sakei exerting greater inhibitory effect. Inoculation with both bio-protective cultures also significantly decreased the total volatile basic nitrogen values of stored samples (p<0.05). Taken together, the results proved the benefits of inoculation with lactic acid bacteria especially L. sakei as a potential way to inhibit growth of spoilage-related bacteria and improve the shelf life of vacuum-packaged raw beef.

Bio-protective potential of lactic acid bacteria: Effect of Lactobacillus sakei and Lactobacillus curvatus on changes of the microbial community in vacuum-packaged chilled beef

PubMed Central

Zhang, Yimin; Zhu, Lixian; Dong, Pengcheng; Liang, Rongrong; Mao, Yanwei; Qiu, Shubing

2018-01-01

Objective This study was to determine the bacterial diversity and monitor the community dynamic changes during storage of vacuum-packaged sliced raw beef as affected by Lactobacillus sakei and Lactobacillus curvatus. Methods L. sakei and L. curvatus were separately incubated in vacuumed-packaged raw beef as bio-protective cultures to inhibit the naturally contaminating microbial load. Dynamic changes of the microbial diversity of inoculated or non-inoculated (control) samples were monitored at 4°C for 0 to 38 days, using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). Results The DGGE profiles of DNA directly extracted from non-inoculated control samples highlighted the order of appearance of spoilage bacteria during storage, showing that Enterbacteriaceae and Pseudomonas fragi emerged early, then Brochothrix thermosphacta shared the dominant position, and finally, Pseudomonas putida showed up became predominant. Compared with control, the inoculation of either L. sakei or L. curvatus significantly lowered the complexity of microbial diversity and inhibited the growth of spoilage bacteria (p<0.05). Interestingly, we also found that the dominant position of L. curvatus was replaced by indigenous L. sakei after 13 d for L. curvatus-inoculated samples. Plate counts on selective agars further showed that inoculation with L. sakei or L. curvatus obviously reduced the viable counts of Enterbacteraceae, Pseudomonas spp. and B. thermosphacta during later storage (p< 0.05), with L. sakei exerting greater inhibitory effect. Inoculation with both bio-protective cultures also significantly decreased the total volatile basic nitrogen values of stored samples (p<0.05). Conclusion Taken together, the results proved the benefits of inoculation with lactic acid bacteria especially L. sakei as a potential way to inhibit growth of spoilage-related bacteria and improve the shelf life of vacuum-packaged raw beef. PMID:29059725

Effect of high intensity ultrasound on the fermentation profile of Lactobacillus sakei in a meat model system.

PubMed

Ojha, Kumari Shikha; Kerry, Joseph P; Alvarez, Carlos; Walsh, Des; Tiwari, Brijesh K

2016-07-01

The objective of this study was to investigate the efficacy of high intensity ultrasound on the fermentation profile of Lactobacillus sakei in a meat model system. Ultrasound power level (0-68.5 W) and sonication time (0-9 min) at 20 °C were assessed against the growth of L. sakei using a Microplate reader over a period of 24h. The L. sakei growth data showed a good fit with the Gompertz model (R(2)>0.90; SE<0.042). Second order polynomial models demonstrated the effect of ultrasonic power and sonication time on the specific growth rate (SGR, μ, h(-1)) and lag phase (λ, h). A higher SGR and a shorter lag phase were observed at low power (2.99 W for 5 min) compared to control. Conversely, a decrease (p<0.05) in SGR with an increase in lag phase was observed with an increase in ultrasonic power level. Cell-free extracts obtained after 24h fermentation of ultrasound treated samples showed antimicrobial activity against Staphylococcus aureus, Listeria monocytogenes, Escherichia coli and Salmonella typhimurium at lower concentrations compared to control. No significant difference (p<0.05) among treatments was observed for lactic acid content after a 24h fermentation period. This study showed that both stimulation and retardation of L. sakei is possible, depending on the ultrasonic power and sonication time employed. Hence, fermentation process involving probiotics to develop functional food products can be tailored by selection of ultrasound processing parameters. Copyright © 2016 Elsevier B.V. All rights reserved.

Purification and Characterization of an Arginine Aminopeptidase from Lactobacillus sakei

PubMed Central

Sanz, Yolanda; Toldrá, Fidel

2002-01-01

An arginine aminopeptidase (EC 3.4.11.6) that exclusively hydrolyzes basic amino acids from the amino (N) termini of peptide substrates has been purified from Lactobacillus sakei. The purification procedure consisted of ammonium sulfate fractionation and three chromatographic steps, which included hydrophobic interaction, gel filtration, and anion-exchange chromatography. This procedure resulted in a recovery rate of 4.2% and a 500-fold increase in specific activity. The aminopeptidase appeared to be a trimeric enzyme with a molecular mass of 180 kDa. The activity was optimal at pH 5.0 and 37°C. The enzyme was inhibited by sulfhydryl group reagents and several divalent cations (Cu2+, Hg2+, and Zn2+) but was activated by reducing agents, metal-chelating agents, and sodium chloride. The enzyme showed a preference for arginine at the N termini of aminoacyl derivatives and peptides. The Km values for Arg-7-amido-4-methylcoumarin (AMC) and Lys-AMC were 15.9 and 26.0 μM, respectively. The nature of the amino acid residue at the C terminus of dipeptides has an effect on hydrolysis rates. The activity was maximal toward dipeptides with Arg, Lys, or Ala as the C-terminal residue. The properties of the purified enzyme, its potential function in the release of arginine, and its further metabolism are discussed because, as a whole, it could constitute a survival mechanism for L. sakei in the meat environment. PMID:11916721

Proteomic and genetics insights on the response of the bacteriocinogenic Lactobacillus sakei CRL1862 during biofilm formation on stainless steel surface at 10°C.

PubMed

Pérez-Ibarreche, Mariana; Mendoza, Lucía M; Vignolo, Graciela; Fadda, Silvina

2017-10-03

Some lactic acid bacteria have the ability to form biofilms on food-industry surfaces and this property could be used to control food pathogens colonization. Lactobacillus sakei CR1862 was selected considering its bacteriocinogenic nature and ability to adhere to abiotic surfaces at low temperatures. In this study, the proteome of L. sakei CRL1862 grown either under biofilm on stainless steel surface and planktonic modes of growth at 10°C, was investigated. Using two-dimensional gel electrophoresis, 29 out of 43 statistically significant spots were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Ten proteins resulted up-regulated whereas 16 were down-regulated during biofilm formation. Differentially expressed proteins were found to belong to carbohydrate, nucleotide, aminoacid and lipid metabolisms as well as translation, peptide hydrolysis, cell envelope/cell wall biosynthesis, adaption to atypical conditions and protein secretion. Some proteins related to carbohydrate and nucleotide metabolisms, translation and peptide degradation were overexpressed whereas those associated to stress conditions were synthesized in lower amounts. It seems that conditions for biofilm development would not imply a stressful environment for L. sakei CRL1862 cells, directing its growth strategy towards glycolytic flux regulation and reinforcing protein synthesis. In addition, L. sakei CRL1862 showed to harbor nine out of ten assayed genes involved in biofilm formation and protein anchoring. By applying qRT-PCR analysis, four of these genes showed to be up regulated, srtA2 being the most remarkable. The results of this study contribute to the knowledge of the physiology of L. sakei CRL1862 growing in biofilm on a characteristic food contact surface. The use of this strain as green biocide preventing L. monocytogenes post-processing contamination on industrial surfaces may be considered. Copyright © 2017 Elsevier B.V. All rights reserved.

The kinetics of the arginine deiminase pathway in the meat starter culture Lactobacillus sakei CTC 494 are pH-dependent.

PubMed

Rimaux, T; Vrancken, G; Pothakos, V; Maes, D; De Vuyst, L; Leroy, F

2011-05-01

Lactobacillus sakei is frequently present as the dominant lactic acid bacterium in spontaneously fermented meat products, demonstrating its competitiveness in and adaptation to the meat environment. Since meat is generally low in carbohydrate content, the ability to utilize other energy sources to generate ATP, such as arginine via the arginine deiminase (ADI) pathway, represents a competitive benefit. In this study, the kinetics of growth and arginine conversion capabilities of Lb. sakei CTC 494 were analyzed, and a model was set up to describe the influence of pH on growth and arginine conversion. A series of in vitro batch fermentations using reconstituted MRS medium at different constant pH values (pH 4.50-pH 7.75) was performed. Arginine conversion through the ADI pathway, which was activated from the stationary growth phase on, resulted in the production of both citrulline and ornithine for all pH conditions tested. However, the pattern and the ratio of the end-products of the ADI pathway were influenced by pH. For certain pH values (between pH 5.0 and 6.5), a further conversion of citrulline into ornithine was found when all arginine was depleted. Characterization of responses of the ADI pathway in Lb. sakei CTC 494 to environmental conditions will allow a better understanding and control of this important starter culture in meat fermentations. Copyright © 2010 Elsevier Ltd. All rights reserved.

Modeling Bacteriocin Resistance and Inactivation of Listeria innocua LMG 13568 by Lactobacillus sakei CTC 494 under Sausage Fermentation Conditions

PubMed Central

Leroy, Frédéric; Lievens, Kristoff; De Vuyst, Luc

2005-01-01

In mixed cultures, bacteriocin production by the sausage isolate Lactobacillus sakei CTC 494 rapidly inactivated sensitive Listeria innocua LMG 13568 cells, even at low bacteriocin activity levels. A small fraction of the listerial population was bacteriocin resistant. However, sausage fermentation conditions inhibited regrowth of resistant cells. PMID:16269805

Industrial application of an antilisterial strain of Lactobacillus sakei as a protective culture and its effect on the sensory acceptability of cooked, sliced, vacuum-packaged meats.

PubMed

Bredholt, S; Nesbakken, T; Holck, A

2001-06-15

The application of a protective lactic acid bacterium (LAB) during the commercial production of cooked meat products is described. The LAB, a strain of Lactobacillus sakei, was previously isolated from cooked ham and inhibited growth of Listeria monocytogenes and Escherichia coli O157:H7 in this product. L. sakei was applied to the cooked products at a concentration of 10(5)-10(6) cfu/g immediately before slicing and vacuum-packaging using a hand-operated spraying bottle. The LAB strain inhibited growth of 10(3) cfu/g of a cocktail of three rifampicin resistant mutant L. monocytogenes strains both at 8 degrees C and 4 degrees C. Consumer acceptance tests of cooked ham and of servelat sausage, a Norwegian non-fermented cooked meat sausage, showed that control and inoculated products were equally acceptable. The products were still acceptable after storage for 28 days at 4 degrees C and, after opening the packages, for a further 5 days at 4 degrees C. The findings presented here confirm that the L. sakei strain is suitable for use as a protective culture and may technically easily be implemented in the commercial production of cooked meat products.

Genome Sequence of Lactobacillus sakei LK-145 Isolated from a Japanese Sake Cellar as a High Producer of d-Amino Acids

PubMed Central

Kato, Shiro

2017-01-01

ABSTRACT This announcement reports the complete genome sequence of strain LK-145 of Lactobacillus sakei isolated from a Japanese sake cellar as a potent strain for the production of large amounts of d-amino acids. Three putative genes encoding an amino acid racemase were identified. PMID:28818888

High-level expression of recombinant beta-galactosidases in Lactobacillus plantarum and Lactobacillus sakei using a Sakacin P-based expression system.

PubMed

Halbmayr, Elisabeth; Mathiesen, Geir; Nguyen, Thu-Ha; Maischberger, Thomas; Peterbauer, Clemens K; Eijsink, Vincent G H; Haltrich, Dietmar

2008-06-25

This work presents the cloning and expression of the genes encoding heterodimeric beta-galactosidases from Lactobacillus reuteri L103, Lactobacillus acidophilus R22, Lactobacillus plantarum WCFS1, and Lactobacillus sakei Lb790. These enzymes consist of two subunits of approximately 73 and 35 kDa, which are encoded by two overlapping genes, lacL and lacM, respectively. We have cloned these genes into the lactobacillal expression vectors pSIP403 and pSIP409, which are based on the sakacin P operon of L. sakei ( Sørvig et al. Microbiology 2005, 151, 2439- 2449 ), and expressed them in the host strains L. plantarum WCFS1 and L. sakei Lb790. Results varied considerably, ranging from 2.23 to 61.1 U/mg of beta-galactosidase activity, depending on the origin of the lacLM genes, the host strain, and the expression vector used. Highest expression levels were obtained in a laboratory cultivation of L. plantarum WCFS1 harboring the plasmid pEH3R containing the lacLM gene from L. reuteri L103. These cultivations yielded approximately 23 000 U of beta-galactosidase activity per liter, corresponding to the formation of roughly 100 mg of recombinant protein per liter of fermentation medium, and beta-galactosidase levels amounted to 55% of the total intracellular protein of the host organism. To further verify the suitability of this expression system, recombinant beta-galactosidase from L. reuteri was purified to apparent homogeneity. The properties of the purified enzyme were essentially identical with the properties of purified native beta-galactosidase from L. reuteri L103. The presented results lead the way to efficient overproduction of beta-galactosidase in a food-grade expression system, which is of high interest for applications in food industry.

First Functional and Mutational Analysis of Group 3 N-Acetylneuraminate Lyases from Lactobacillus antri and Lactobacillus sakei 23K

PubMed Central

García-García, María Inmaculada; Gil-Ortiz, Fernando; García-Carmona, Francisco; Sánchez-Ferrer, Álvaro

2014-01-01

N-acetyl neuraminate lyases (NALs) catalyze the reversible aldol cleavage of N-acetyl neuraminic acid (Neu5Ac) to pyruvate and N-acetyl-D-mannosamine (ManNAc). Previous phylogenetic studies divided NALs into four different groups. Groups 1 and 2 have been well characterized at both kinetic and molecular levels, but no NAL from group 3 has been studied to date. In this work, a functional characterization of two group 3 members was performed using the recombinant NALs from Lactobacillus antri and Lactobacillus sakei 23K, revealing an optimal pH of between 6.0 and 7.0, low stability at basic pHs (>8.0), low optimal temperatures and, especially, low catalytic efficiency compared with their counterparts in group 1 and 2. The mutational analysis carried out showed that a plausible molecular reason for the low activity shown by Lactobacillus antri and Lactobacillus sakei 23k NALs compared with group 1 and 2 NALs could be the relatively small sugar-binding pocket they contain. A functional divergence analysis concluding that group 3 is more closely related to group 2 than to group 1. PMID:24817128

Metabolite profile of koji amazake and its lactic acid fermentation product by Lactobacillus sakei UONUMA.

PubMed

Oguro, Yoshifumi; Nishiwaki, Toshikazu; Shinada, Ryota; Kobayashi, Kazuya; Kurahashi, Atsushi

2017-08-01

The koji amazake is a traditional sweet Japanese beverage. It has been consumed for over a thousand years in Japan; nonetheless, little is yet known of the ingredients in koji amazake. Therefore, this study aimed to analyze the metabolites of koji amazake using a metabolomics approach. Additionally, we reformed the flavor of koji amazake by lactic acid fermentation (LAF-amazake) using Lactobacillus sakei UONUMA, which was isolated from snow caverns. The purpose of this article is to identify the ingredients in these beverages. In LAF-amazake and koji amazake, sugars, amino acids, organic acids, and vitamin B complex were determined in the two beverages, and over 300 compounds were detected in total. Thirteen saccharides were identified including two unknown trisaccharides, and there were no differences in these between the two beverages. In LAF-amazake, lactic acid, vitamin B2 (riboflavin), B3 (nicotinic acid and nicotinamide), and B6 (pyridoxine) were significantly increased as compared to koji amazake, whereas malate and glutamine decreased. These results suggested that LAF, malolactic fermentation, and glutamine deamidation occurred simultaneously in LAF-amazake. L. sakei UONUMA strains produced these vitamins. Moreover, it was surprising that acetylcholine, a well-known neurotransmitter, was newly generated in LAF-amazake. Here, we have succeeded in reforming the flavor of koji amazake and obtained these metabolic data on the two beverages. The present study could provide useful basic information for promoting functional analyses of koji amazake and LAF-amazake for human health. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Complete and Draft Genome Sequences of Nine Lactobacillus sakei Strains Selected from the Three Known Phylogenetic Lineages and Their Main Clonal Complexes.

PubMed

Loux, Valentin; Coeuret, Gwendoline; Zagorec, Monique; Champomier Vergès, Marie-Christine; Chaillou, Stéphane

2018-04-19

We present here the complete and draft genome sequences of nine Lactobacillus sakei strains, selected from the entire range of clonal complexes from the three known lineages of the species. The strains were chosen to provide a wide view of pangenomic and plasmidic diversity for this important foodborne species. Copyright © 2018 Loux et al.

Development and Optimization of a Fluorescent Differential Display PCR System for Analyzing the Stress Response in Lactobacillus sakei Strains

PubMed Central

Bonomo, Maria Grazia; Sico, Maria Anna; Grieco, Simona; Salzano, Giovanni

2009-01-01

Lactobacillus sakei is widely used as starter in the production process of Italian fermented sausages and its growth and survival are affected by various factors. We studied the differential expression of genome in response to different stresses by the fluorescent differential display (FDD) technique. This study resulted in the development and optimization of an innovative technique, with a high level of reproducibility and quality, which allows the identification of gene expression changes associated with different microbial behaviours under different growth conditions. PMID:22253979

Application of chemometrics to assess the influence of ultrasound frequency, Lactobacillus sakei culture and drying on beef jerky manufacture: Impact on amino acid profile, organic acids, texture and colour.

PubMed

Shikha Ojha, K; Granato, Daniel; Rajuria, Gaurav; Barba, Francisco J; Kerry, Joseph P; Tiwari, Brijesh K

2018-01-15

The effects of ultrasound (US) frequency, addition of Lactobacillus sakei culture and drying time on key nutritional (protein, amino acids, and organic acids) and physicochemical properties (texture and colour) of cultured and uncultured beef jerky were evaluated. Cultured and uncultured jerky samples were subjected to US frequencies of 25kHz, 33kHz and 45kHz for 30min prior to marination and drying. Principal component analysis demonstrated a significant effect of beef jerky processing conditions on physicochemical properties. Taurine content of jerky samples was found to increase with an increase in ultrasonic frequencies for cultured samples. No significant changes in colour values were observed for ultrasound pre-treated and control samples. Interactive effects of culture treatment, drying and ultrasonic frequency were observed. This study demonstrates that the nutritional profile of beef jerky can be improved through the incorporation of L. sakei. Copyright © 2017 Elsevier Ltd. All rights reserved.

The Use of Tomato Powder Fermented with Pediococcus pentosaceus and Lactobacillus sakei for the Ready-to-Cook Minced Meat Quality Improvement

PubMed Central

Juodeikiene, Grazina; Zadeike, Daiva; Viskelis, Pranas; Urbonaviciene, Dalia

2015-01-01

Summary In this study, the influence of lactic acid fermentation on the quality of tomato powder was evaluated. The effect of adding fermented tomato powder to ready-to-cook minced pork meat to improve its nutritional value and sensory characteristics was also analysed. The cell growth of Lactobacillus sakei (7.53 log CFU/g) was more intense in the medium containing tomato powder, compared to the growth of Pediococcus pentosaceus (6.35 log CFU/g) during 24 h of fermentation; however, higher acidity (pH=4.1) was observed in the tomato powder samples fermented with Pediococcus pentosaceus. The spontaneous fermentation of tomato powder reduced cell growth by 38% and pH values slightly increased to 4.17, compared to the fermentation with pure LAB. The lactofermentation of tomato powder increased the average β-carotene and lycopene mass fractions by 43.9 and 50.2%, respectively, compared with the nonfermented samples. Lycopene and β-carotene contents in the ready-to-cook minced pork meat were proportional to the added tomato powder (10 and 30%). After cooking, β-carotene and lycopene contents decreased, on average, by 24.2 and 41.2%, respectively. The highest loss (up to 49.2%) of carotenoids was found in samples with 30% nonfermented tomato powder. Tomato powder fermented with 10% Lactobacillus sakei KTU05-6 can be recommended as both a colouring agent and a source of lycopene in the preparation of ready-to-cook minced pork meat. PMID:27904345

Dextransucrase Expression Is Concomitant with that of Replication and Maintenance Functions of the pMN1 Plasmid in Lactobacillus sakei MN1

PubMed Central

Nácher-Vázquez, Montserrat; Ruiz-Masó, José A.; Mohedano, María L.; del Solar, Gloria; Aznar, Rosa; López, Paloma

2017-01-01

The exopolysaccharide synthesized by Lactobacillus sakei MN1 is a dextran with antiviral and immunomodulatory properties of potential utility in aquaculture. In this work we have investigated the genetic basis of dextran production by this bacterium. Southern blot hybridization experiments demonstrated the plasmidic location of the dsrLS gene, which encodes the dextransucrase involved in dextran synthesis. DNA sequencing of the 11,126 kbp plasmid (pMN1) revealed that it belongs to a family which replicates by the theta mechanism, whose prototype is pUCL287. The plasmid comprises the origin of replication, repA, repB, and dsrLS genes, as well as seven open reading frames of uncharacterized function. Lb. sakei MN1 produces dextran when sucrose, but not glucose, is present in the growth medium. Therefore, plasmid copy number and stability, as well as dsrLS expression, were investigated in cultures grown in the presence of either sucrose or glucose. The results revealed that pMN1 is a stable low-copy-number plasmid in both conditions. Gene expression studies showed that dsrLS is constitutively expressed, irrespective of the carbon source present in the medium. Moreover, dsrLS is expressed from a monocistronic transcript as well as from a polycistronic repA-repB-orf1-dsrLS mRNA. To our knowledge, this is the first report of a plasmid-borne dextransucrase-encoding gene, as well as the first time that co-transcription of genes involved in plasmid maintenance and replication with a gene encoding an enzyme has been established. PMID:29209293

Purification and Characterization of an X-Prolyl-Dipeptidyl Peptidase from Lactobacillus sakei

PubMed Central

Sanz, Yolanda; Toldrá, Fidel

2001-01-01

An X-prolyl-dipeptidyl peptidase has been purified from Lactobacillus sakei by ammonium sulfate fractionation and five chromatographic steps, which included hydrophobic interaction, anion-exchange chromatography, and gel filtration chromatography. This procedure resulted in a recovery yield of 7% and an increase in specificity of 737-fold. The enzyme appeared to be a dimer with a subunit molecular mass of approximately 88 kDa. Optimal activity was shown at pH 7.5 and 55°C. The enzyme was inhibited by serine proteinase inhibitors and several divalent cations (Cu2+, Hg2+, and Zn2+). The enzyme almost exclusively hydrolyzed X-Pro from the N terminus of each peptide as well as fluorescent and colorimetric substrates; it also hydrolyzed X-Ala at the N terminus, albeit at lower rates. Km s for Gly-Pro- and Lys-Ala-7-amido-4-methylcoumarin were 29 and 88 μM, respectively; those for Gly-Pro- and Ala-Pro-p-nitroanilide were 192 and 50 μM, respectively. Among peptides, β-casomorphin 1-3 was hydrolyzed at the highest rates, while the relative hydrolysis of the other tested peptides was only 1 to 12%. The potential role of the purified enzyme in the proteolytic pathway by catalyzing the hydrolysis of peptide bonds involving proline is discussed. PMID:11282638

Detection of different microenvironments and Lactobacillus sakei biotypes in Ventricina, a traditional fermented sausage from central Italy.

PubMed

Tremonte, Patrizio; Sorrentino, Elena; Pannella, Gianfranco; Tipaldi, Luca; Sturchio, Marina; Masucci, Armando; Maiuro, Lucia; Coppola, Raffaele; Succi, Mariantonietta

2017-02-02

The present study evaluated the physico-chemical and microbiological features of Ventricina, considering for the first time the presence of different compartments deriving from the technology of production. In fact meat pieces (pork muscle and fat cut into cubes of about 10-20cm 3 ), mixed with other ingredients and then stuffed into pig bladder, are still distinguishable at the end of the ripening. They appear delimited on the outside by the casing and inside by thin layers consisting of spices (mainly red pepper powder), salt and meat juices. Our results showed that the exterior (portion of the product in contact with the casing), the interstice (area between the different cubes of meat or fat) and the heart (the inner portion of meat cubes) had distinctive values of pH and a w , and a typical microbial progression, so that they can be considered as different ecological niches, here called microenvironments. The study of lactic acid bacteria population, performed with PCR-DGGE and sequence analysis targeting the V1-V3 region of the 16S rRNA gene (rDNA), highlighted the presence of a few species, including Lactobacillus sakei, Lb. plantarum, Weissella hellenica and Leuconostoc mesenteroides. The RAPD-PCR analysis performed on Lb. sakei, recognised as the predominant species, allowed the differentiation into three biotypes, with that characterised by the highest acidifying and proteolytic activities and the highest ability to grow in the presence of sodium chloride prevailing. This leading biotype, detectable in the interstice during the entire ripening period, was isolated in the microenvironments exterior and heart starting from the 30th d of ripening, and it was the sole biotype present at the end of the ripening. The analysis of microenvironments through the scanning electron microscopy (SEM) evidenced the presence of micro-channels, which could favour the microbial flow from the interstice to the exterior and the heart. Moreover, the SEM analysis allowed the

A Combined Model To Predict the Functionality of the Bacteriocin-Producing Lactobacillus sakei Strain CTC 494

PubMed Central

Leroy, Frédéric; De Vuyst, Luc

2003-01-01

The use of bacteriocin-producing lactic acid bacteria for improved food fermentation processes seems promising. However, lack of fundamental knowledge about the functionality of bacteriocin-producing strains under food fermentation conditions hampers their industrial use. Predictive microbiology or a mathematical estimation of microbial behavior in food ecosystems may help to overcome this problem. In this study, a combined model was developed that was able to estimate, from a given initial situation of temperature, pH, and nutrient availability, the growth and self-inhibition dynamics of a bacteriocin-producing Lactobacillus sakei CTC 494 culture in (modified) MRS broth. Moreover, the drop in pH induced by lactic acid production and the bacteriocin activity toward Listeria as an indicator organism were modeled. Self-inhibition was due to the depletion of nutrients as well as to the production of lactic acid. Lactic acid production resulted in a pH drop, an accumulation of toxic undissociated lactic acid molecules, and a shift in the dissociation degree of the growth-inhibiting buffer components. The model was validated experimentally. PMID:12571034

Growth of the Bacteriocin-Producing Lactobacillus sakei Strain CTC 494 in MRS Broth Is Strongly Reduced Due to Nutrient Exhaustion: a Nutrient Depletion Model for the Growth of Lactic Acid Bacteria

PubMed Central

Leroy, Frédéric; De Vuyst, Luc

2001-01-01

Although commercial MRS broth has been designed to allow excellent growth of lactobacilli, most of these bacteria are still subjected to a self-inhibiting process. The most likely explanation is the accumulation of lactic acid or other toxic end products and the depletion of nutrients. In this study, the self-inhibition of Lactobacillus sakei CTC 494 was analyzed in a kinetic way, and a nutrient depletion model was set up to describe the growth inhibition process. This simple model has considerable advantages compared to commonly used descriptive models such as the logistic growth equation. It offers a better fit and a more realistic description of the growth data by taking into account both growth inhibition due to lactic acid production and changes in growth rates due to nutrient depletion. Depending on the fermentation conditions, in MRS broth there appears to be a strong decrease of the specific growth rate over time. Some undefined compounds present in the complex nitrogen source of MRS broth appear to be of crucial importance because of their limited availability. Moreover, nutrient availability affects bacteriocin production through its effect on cell growth as well as on the bacteriocin production per cell. A plateau value for the bacteriocin production by L. sakei CTC 494 was observed. PMID:11571136

The Presence of Salt and a Curing Agent Reduces Bacteriocin Production by Lactobacillus sakei CTC 494, a Potential Starter Culture for Sausage Fermentation

PubMed Central

Leroy, Frédéric; de Vuyst, Luc

1999-01-01

The specific conditions in the batter of raw fermented sausages may reduce the efficiency of bacteriocin-producing starter cultures. In this work, using in vitro fermentation, we found that sodium chloride and sodium nitrite interfere with the growth of Lactobacillus sakei CTC 494, an organism which produces the antilisterial bacteriocin sakacin K. Because sakacin K production follows primary metabolite kinetics, a decrease in cell formation resulted in a decrease in sakacin K production as well. Sodium chloride dramatically influenced bacteriocin production by decreasing both biomass production and specific bacteriocin production. Sodium nitrite, however, had no effect on specific bacteriocin production and decreased bacteriocin production only because of its effect on cell growth. Moreover, sodium nitrite enhanced the toxic effect of lactic acid on bacterial growth. PMID:10583988

Neuroprotective effects of fermented soybean products (Cheonggukjang) manufactured by mixed culture of Bacillus subtilis MC31 and Lactobacillus sakei 383 on trimethyltin-induced cognitive defects mice.

PubMed

Go, Jun; Kim, Ji Eun; Kwak, Moon Hwa; Koh, Eun Kyoung; Song, Sung Hwa; Sung, Ji Eun; Kim, Dong Sup; Hong, Jin Tae; Hwang, Dae Youn

2016-07-01

This study aimed to investigate the beneficial effects of Cheonggukjang (CGK) manufactured by mixed culture of Bacillus subtilis MC31 and Lactobacillus sakei 383 on neurotoxic damages. The specific aspects of brain functions were measured in Institute for Cancer Research (ICR) mice that had been pretreated for 4 weeks with three difference doses of CGK before trimethyltin (TMT) treatment. The short- and long-term memory loss induced by TMT treatment was significantly improved in the CGK-pretreated group in a dose-dependent manner. The number of dead cells in the granule cell layer of the dentate gyrus was decreased in the TMT/CGK-cotreated group relative to the TMT/vehicle-treated group, whereas significant suppression of acetylcholinesterase (AChE) activity was observed in the same group. Additionally, a dose-dependent increase in nerve growth factor (NGF) concentration, activation of the NGF receptor signaling pathway including the TrkA high affinity receptor and p75(NTR) low affinity receptor, and decline in Bax/Bcl-2 level was measured in all TMT/CGK-treated groups, although a decrease in the active form of caspase-3 was observed in the TMT/H-CGK-treated group. Furthermore, superoxide dismutase (SOD) activity was enhanced in the TMT/CGK-treated group, whereas the level of malondialdehyde (MDA), a marker of lipid peroxidation, was 43-58% lower in the TMT/CGK-treated group than the TMT/vehicle-treated group. These results demonstrate that CGK fermented by mixed culture of B. subtilis and L. sakei could exert a wide range of beneficial activities for neurodegenerative diseases, including Alzheimer, Parkinson, and Huntington disease.

Low Temperature and Modified Atmosphere: Hurdles for Antibiotic Resistance Transfer?

PubMed

Van Meervenne, Eva; Van Coillie, Els; Van Weyenberg, Stephanie; Boon, Nico; Herman, Lieve; Devlieghere, Frank

2015-12-01

Food is an important dissemination route for antibiotic-resistant bacteria. Factors used during food production and preservation may contribute to the transfer of antibiotic resistance genes, but research on this subject is scarce. In this study, the effect of temperature (7 to 37°C) and modified atmosphere packaging (air, 50% CO2-50% N2, and 100% N2) on antibiotic resistance transfer from Lactobacillus sakei subsp. sakei to Listeria monocytogenes was evaluated. Filter mating was performed on nonselective agar plates with high-density inocula. A more realistic setup was created by performing modified atmosphere experiments on cooked ham using high-density and low-density inocula. Plasmid transfer was observed between 10 and 37°C, with plasmid transfer also observed at 7°C during a prolonged incubation period. When high-density inocula were used, transconjugants were detected, both on agar plates and cooked ham, under the three atmospheres (air, 50% CO2-50% N2, and 100% N2) at 7°C. This yielded a median transfer ratio (number of transconjugants/number of recipients) with an order of magnitude of 10(-4) to 10(-6). With low-density inocula, transfer was only detected under the 100% N2 atmosphere after 10-day incubation at 7°C, yielding a transfer ratio of 10(-5). Under this condition, the highest bacterial density was obtained. The results indicate that low temperature and modified atmosphere packaging, two important hurdles in the food industry, do not necessarily prevent plasmid transfer from Lactobacillus sakei subsp. sakei to Listeria monocytogenes.

Efficacies of garlic and L. sakei in wine-based marinades for controlling Listeria monocytogenes and Salmonella spp. in Chouriço de Vinho, a dry sausage made from wine-marinated pork.

PubMed

Linares, María Belén; Garrido, María Dolores; Martins, Conceição; Patarata, Luis

2013-05-01

Chouriço de vinho is made from roughly minced (10 to 30 mm) pork and fat, seasoned with a marinade made from wine, salt, garlic, and other facultative seasonings used according to the recipe of each producer. The batter is maintained at 4 to 7 ºC for 24 to 48 h. It is then stuffed into natural thin pork gut, cold smoked and matured at a low temperature for 1 to 4 wk. The effect of garlic used in wine-based marinade and a starter culture of indigenous Lactobacillus sakei on the behavior of Listeria monocytogenes and Salmonella spp. in the processing of chouriço was investigated. The garlic (as powder and fresh juice) was found to contribute (P < 0.05) to the control of both pathogens in broth. Garlic dose, as tested within the usual limits used for seasoning, did not impact the reduction of pathogens. Garlic-wine-based marinade and a starter culture of indigenous L. sakei contribute to controlling L. monocytogenes and Salmonella spp. in the processing of chouriço. Their presence was responsible for the loss of viability of L. monocytogenes and Salmonella spp. following 5 d of drying, even sooner than situations where no garlic was used. The results of the present work show that the use of a wine-based marinade with garlic has an important role in ensuring the safety of the product. © 2013 Institute of Food Technologists®

The acid tolerant L-arabinose isomerase from the food grade Lactobacillus sakei 23K is an attractive D-tagatose producer.

PubMed

Rhimi, Moez; Ilhammami, Rimeh; Bajic, Goran; Boudebbouze, Samira; Maguin, Emmanuelle; Haser, Richard; Aghajari, Nushin

2010-12-01

The araA gene encoding an L-arabinose isomerase (L-AI) from the psychrotrophic and food grade Lactobacillus sakei 23K was cloned, sequenced and over-expressed in Escherichia coli. The recombinant enzyme has an apparent molecular weight of nearly 220 kDa, suggesting it is a tetramer of four 54 kDa monomers. The enzyme is distinguishable from previously reported L-AIs by its high activity and stability at temperatures from 4 to 40 degrees C, and pH from 3 to 8, and by its low metal requirement of only 0.8 mM Mn(2+) and 0.8 mM Mg(2+) for its maximal activity and thermostability. Enzyme kinetic studies showed that this enzyme displays a high catalytic efficiency allowing D-galactose bioconversion rates of 20% and 36% at 10 and 45 degrees C, respectively, which are useful for commercial production of D-tagatose. 2010 Elsevier Ltd. All rights reserved.

Identification and characterization of lactic acid bacteria isolated from mixed pasture of timothy and orchardgrass, and its badly preserved silage.

PubMed

Tohno, Masanori; Kobayashi, Hisami; Nomura, Masaru; Uegaki, Ryuichi; Cai, Yimin

2012-04-01

In order to understand the relationship between lactic acid bacteria (LAB) species and silage fermentation, a total of 65 LAB strains isolated from mixed pasture of timothy (Phleum pratense L.) and orchardgrass (Dactylis glomerata L.), and its badly preserved silages were subjected to phenotypic and genetic analysis. According to these analyses, the isolates were divided into 13 groups, including Enterococcus gallinarum, Lactobacillus acidipiscis, L. coryniformis subsp. coryniformis, L. coryniformis subsp. torquens, L. curvatus, L. paraplantarum, L. plantarum subsp. argentoratensis, L. plantarum subsp. plantarum, L. sakei subsp. carnosus, Lactococcus garvieae, Lactococcus lactis subsp. cremoris, Leuconostoc pseudomesenteroides, Pediococcus acidilactici, Pediococcus pentosaceus, Weissella hellenica, Weissella paramesenteroides and Carnobacterium divergens. This is the first report to document that C. divergens, L. acidipiscis, L. sakei subsp. carnosus, L. garvieae, phenotypically novel L. lactis subsp. cremoris, E. gallinarum and W. hellenica are present in vegetative forage crops. L. plantarum group strains were most frequently isolated from the badly preserved silages. Some isolates showed a wide range of growth preferences for carbohydrate utilization, optimal growth pH and temperature in vitro, indicating that they have a high growth potential. These results are useful in understanding the diversity of LAB associated with decayed silage of timothy and orchardgrass. © 2011 The Authors. Animal Science Journal © 2011 Japanese Society of Animal Science.

Effects of glucose availability in Lactobacillus sakei; metabolic change and regulation of the proteome and transcriptome

PubMed Central

Mosleth, Ellen F.; Rud, Ida; Branco dos Santos, Filipe; Snipen, Lars; Liland, Kristian Hovde

2017-01-01

Effects of glucose availability were investigated in Lactobacillus sakei strains 23K and LS25 cultivated in anaerobic, glucose-limited chemostats set at high (D = 0.357 h-1) and low (D = 0.045 h-1) dilution rates. We observed for both strains a shift from homolactic towards more mixed acid fermentation when comparing high to low growth rates. However, this change was more pronounced for LS25 than for 23K, where dominating products were lactate>formate>acetate≥ethanol at both conditions. A multivariate approach was used for analyzing proteome and transcriptome data from the bacterial cultures, where the predictive power of the omics data was used for identifying features that can explain the differences in the end-product profiles. We show that the different degree of response to the same energy restriction revealed interesting strain specific regulation. An elevated formate production level during slow growth, more for LS25 than for 23K, was clearly reflected in correlating pyruvate formate lyase expression. With stronger effect for LS25, differential expression of the Rex transcriptional regulator and NADH oxidase, a target of Rex, indicated that maintainance of the cell redox balance, in terms of the NADH/NAD+ ratio, may be a key process during the metabolic change. The results provide a better understanding of different strategies that cells may deploy in response to changes in substrate availability. PMID:29099858

Isolation of Lactobacillus sakei strain KJ-2008 and its removal of characteristic malodorous gases under anaerobic culture conditions.

PubMed

Kim, Jeong-Dong; Kang, Kook-Hee

2004-12-01

A number of different sources, such as composts, leachates, and pig feces samples were collected from different pig farms in Korea. Several microorganisms were screened for their ability to deodorize the malodorous gases. As a result, a novel malodorous gas-deodorizing bacterial strain KJ-2008 was isolated due to the most abundant of nitrate-supplemented minimal media under anaerobic conditions. Crimp-sealed serum bottles containing nitrate-supplemented minimal medium (MM-NO(3)(-)) in airtight conditions were inoculated with KJ-2008. Nitrate concentration decreased rapidly after 20 h incubation and nitrite production reached almost zero during the time the experimental was carried out. Taxonomic identification including 16S rDNA base sequencing and phylogenetic analysis indicated that the isolate KJ-2008 had a 99.8% homology in its 16S rDNA base sequence with Lactobacillus sakei. Among the volatile fatty acids, acetic acid contained in large amounts in fresh piggery slurry decreased about 40% after 50 h incubation of the strain KJ-2008. n-Butyric acid, n-valeric acid, and iso-valeric acid gradually decreased, and iso-butyric acid and capronic acid dramatically eliminated at initial time with the treatment. Moreover, NH(3) removal efficiency reached a maximum of 98.5% after 50 h of incubation. The concentration of H(2)S did not change.

Molecular Characterization of tet(M) Genes in Lactobacillus Isolates from Different Types of Fermented Dry Sausage

PubMed Central

Gevers, Dirk; Danielsen, Morten; Huys, Geert; Swings, Jean

2003-01-01

The likelihood that products prepared from raw meat and milk may act as vehicles for antibiotic-resistant bacteria is currently of great concern in food safety issues. In this study, a collection of 94 tetracycline-resistant (Tcr) lactic acid bacteria recovered from nine different fermented dry sausage types were subjected to a polyphasic molecular study with the aim of characterizing the host organisms and the tet genes, conferring tetracycline resistance, that they carry. With the (GTG)5-PCR DNA fingerprinting technique, the Tcr lactic acid bacterial isolates were identified as Lactobacillus plantarum, L. sakei subsp. carnosus, L. sakei subsp. sakei, L. curvatus, and L. alimentarius and typed to the intraspecies level. For a selection of 24 Tcr lactic acid bacterial isolates displaying unique (GTG)5-PCR fingerprints, tet genes were determined by means of PCR, and only tet(M) was detected. Restriction enzyme analysis with AccI and ScaI revealed two different tet(M) allele types. This grouping was confirmed by partial sequencing of the tet(M) open reading frame, which indicated that the two allele types displayed high sequence similarities (>99.6%) with tet(M) genes previously reported in Staphylococcus aureus MRSA 101 and in Neisseria meningitidis, respectively. Southern hybridization with plasmid profiles revealed that the isolates contained tet(M)-carrying plasmids. In addition to the tet(M) gene, one isolate also contained an erm(B) gene on a different plasmid from the one encoding the tetracycline resistance. Furthermore, it was also shown by PCR that the tet(M) genes were not located on transposons of the Tn916/Tn1545 family. To our knowledge, this is the first detailed molecular study demonstrating that taxonomically and genotypically diverse Lactobacillus strains from different types of fermented meat products can be a host for plasmid-borne tet genes. PMID:12571056

Macedocin, a Food-Grade Lantibiotic Produced by Streptococcus macedonicus ACA-DC 198

PubMed Central

Georgalaki, Marina D.; Van den Berghe, Erika; Kritikos, Dimitrios; Devreese, Bart; Van Beeumen, Jozef; Kalantzopoulos, George; De Vuyst, Luc; Tsakalidou, Effie

2002-01-01

Streptococcus macedonicus ACA-DC 198, a strain isolated from Greek Kasseri cheese, produces a food-grade lantibiotic named macedocin. Macedocin has a molecular mass of 2,794.76 ± 0.42 Da, as determined by electrospray mass spectrometry. Partial N-terminal sequence analysis revealed 22 amino acid residues that correspond with the amino acid sequence of the lantibiotics SA-FF22 and SA-M49, both of which were isolated from the pathogen Streptococcus pyogenes. Macedocin inhibits a broad spectrum of lactic acid bacteria, as well as several food spoilage and pathogenic bacteria, including Clostridium tyrobutyricum. It displays a bactericidal effect towards the most sensitive indicator strain, Lactobacillus sakei subsp. sakei LMG 13558T, while the producer strain itself displays autoinhibition when it is grown under conditions that do not favor bacteriocin production. Macedocin is active at pHs between 4.0 and 9.0, and it retains activity even after incubation for 20 min at 121°C with 1 atm of overpressure. Inhibition of macedocin by proteolytic enzymes is variable. PMID:12450808

Intraspecies Genomic Diversity and Natural Population Structure of the Meat-Borne Lactic Acid Bacterium Lactobacillus sakei▿ †

PubMed Central

Chaillou, Stéphane; Daty, Marie; Baraige, Fabienne; Dudez, Anne-Marie; Anglade, Patricia; Jones, Rhys; Alpert, Carl-Alfred; Champomier-Vergès, Marie-Christine; Zagorec, Monique

2009-01-01

Lactobacillus sakei is a food-borne bacterium naturally found in meat and fish products. A study was performed to examine the intraspecies diversity among 73 isolates sourced from laboratory collections in several different countries. Pulsed-field gel electrophoresis analysis demonstrated a 25% variation in genome size between isolates, ranging from 1,815 kb to 2,310 kb. The relatedness between isolates was then determined using a PCR-based method that detects the possession of 60 chromosomal genes belonging to the flexible gene pool. Ten different strain clusters were identified that had noticeable differences in their average genome size reflecting the natural population structure. The results show that many different genotypes may be isolated from similar types of meat products, suggesting a complex ecological habitat in which intraspecies diversity may be required for successful adaptation. Finally, proteomic analysis revealed a slight difference between the migration patterns of highly abundant GapA isoforms of the two prevailing L. sakei subspecies (sakei and carnosus). This analysis was used to affiliate the genotypic clusters with the corresponding subspecies. These findings reveal for the first time the extent of intraspecies genomic diversity in L. sakei. Consequently, identification of molecular subtypes may in the future prove valuable for a better understanding of microbial ecosystems in food products. PMID:19114527

Development of a microbial time/temperature indicator prototype for monitoring the microbiological quality of chilled foods.

PubMed

Vaikousi, Hariklia; Biliaderis, Costas G; Koutsoumanis, Konstantinos P

2008-05-01

A time/temperature indicator (TTI) system based on the growth and metabolic activity of a Lactobacillus sakei strain was developed for monitoring food quality throughout the chilled-food chain. In the designed system, an irreversible color change of a chemical chromatic indicator (from red to yellow) progressively occurs due to the pH decline that results from microbial growth and metabolism in a selected medium. The relation of the TTI response (color change) to the growth and metabolic activity (glucose consumption, lactic acid production, pH decrease) of L. sakei was studied. In addition, the temperature dependence of the TTI kinetics was investigated isothermally in the range of 0 to 16 degrees C and modeled with a system of differential equations. At all temperatures tested, the pH and color changes of the TTI system followed closely the growth of L. sakei, with the endpoint (the time at which a distinct visual color change to the final yellow was observed) of the TTI coinciding with a population level of 10(7) to 10(8) CFU/ml. The endpoint decreased from 27 days at 0 degrees C to 2.5 days at 16 degrees C, yielding an activation energy of 97.7 kJ/mol, which was very close to the activation energy of the L. sakei growth rate in the TTI substrate (103.2 kJ/mol). Furthermore, experiments conducted on the effect of the inoculum level showed a negative linear relationship between the level of L. sakei inoculated in the system medium and the endpoint of the TTI. For example, the endpoint at 8 degrees C ranged from 6 to 2 days for inoculum levels of 10(1) and 10(6) CFU/ml, respectively. This relationship allows the easy adjustment of the TTI endpoint at a certain temperature according to the shelf life of the food product of concern by using an appropriate inoculum level of L. sakei. The microbial TTI prototype developed in the present study could be used as an effective tool for monitoring shelf life during the distribution and storage of food products that are

Development of a Microbial Time/Temperature Indicator Prototype for Monitoring the Microbiological Quality of Chilled Foods▿

PubMed Central

Vaikousi, Hariklia; Biliaderis, Costas G.; Koutsoumanis, Konstantinos P.

2008-01-01

A time/temperature indicator (TTI) system based on the growth and metabolic activity of a Lactobacillus sakei strain was developed for monitoring food quality throughout the chilled-food chain. In the designed system, an irreversible color change of a chemical chromatic indicator (from red to yellow) progressively occurs due to the pH decline that results from microbial growth and metabolism in a selected medium. The relation of the TTI response (color change) to the growth and metabolic activity (glucose consumption, lactic acid production, pH decrease) of L. sakei was studied. In addition, the temperature dependence of the TTI kinetics was investigated isothermally in the range of 0 to 16°C and modeled with a system of differential equations. At all temperatures tested, the pH and color changes of the TTI system followed closely the growth of L. sakei, with the endpoint (the time at which a distinct visual color change to the final yellow was observed) of the TTI coinciding with a population level of 107 to 108 CFU/ml. The endpoint decreased from 27 days at 0°C to 2.5 days at 16°C, yielding an activation energy of 97.7 kJ/mol, which was very close to the activation energy of the L. sakei growth rate in the TTI substrate (103.2 kJ/mol). Furthermore, experiments conducted on the effect of the inoculum level showed a negative linear relationship between the level of L. sakei inoculated in the system medium and the endpoint of the TTI. For example, the endpoint at 8°C ranged from 6 to 2 days for inoculum levels of 101 and 106 CFU/ml, respectively. This relationship allows the easy adjustment of the TTI endpoint at a certain temperature according to the shelf life of the food product of concern by using an appropriate inoculum level of L. sakei. The microbial TTI prototype developed in the present study could be used as an effective tool for monitoring shelf life during the distribution and storage of food products that are spoiled primarily by lactic acid

Characterization of Selected Lactobacillus Strains for Use as Probiotics

PubMed Central

Song, Minyu; Yun, Bohyun; Moon, Jae-Hak; Park, Dong-June; Lim, Kwangsei; Oh, Sejong

2015-01-01

The aim of this study was to evaluate the functional properties of lactic acid bacteria from various sources and to identify strains for use as probiotics. Ten Lactobacillus strains were selected and their properties such as bile tolerance, acid resistance, cholesterol assimilation activity, and adherence to HT-29 cells were assessed to determine their potential as probiotics. Lactobacillus sp. JNU 8829, L. casei MB3, L. sakei MA9, L. sakei CH8, and L. acidophilus M23 were found to show full tolerance to the 0.3% bile acid. All strains without L. acidophilus M23 were the most acid-tolerant strains. After incubating the strains at pH 2.5 for 2 h, their viability decreased by 3 Log cells. Some strains survived at pH 2.5 in the presence of pepsin and 0.3% bile acid. Lactobacillus sp. JNU 8829, L. acidophilus KU41, L. acidophilus M23, L. fermentum NS2, L. plantarum M13, and L. plantarum NS3 were found to reduce cholesterol levels by >50% in vitro. In the adhesion assay, Lactobacillus sp. JNU 8829, L. casei MB3, L. sakei MA9, and L. sakei CH8 showed higher adhesion activities after 2 h of co-incubation with the intestinal cells. The results of this comprehensive analysis shows that this new probiotic strain named, Lactobacillus sp. JNU 8829 could be a promising candidate for dairy products. PMID:26761878

Investigation of reduction and tolerance capability of lactic acid bacteria isolated from kimchi against nitrate and nitrite in fermented sausage condition.

PubMed

Paik, Hyun-Dong; Lee, Joo-Yeon

2014-08-01

Lactobacillus brevis KGR3111, Lactobacillus curvatus KGR 2103, Lactobacillus plantarum KGR 5105, and Lactobacillus sakei KGR 4108 isolated from kimchi were investigated for their potential to be used as starter culture for fermented sausages with the capability to reduce and tolerate nitrate/nitrite. The reduction capability of tested strains for nitrate was not dramatic. All tested strains, however, showed the capability to produce nitrite reductase with the reduction amount of 58.46-75.80 mg/l of NO(2)(-). L. brevis and L. plantarum showed nitrate tolerance with the highest number of 8.71 log cfu/ml and 8.81 log cfu/ml, and L. brevis and L. sakei exhibited nitrite tolerance with the highest number of 8.24 log cfu/ml and 8.25 log cfu/ml, respectively. As a result, L. brevis, L. plantarum, and L. sakei isolated from kimchi showed a tolerance against nitrate or nitrite with a good nitrite reduction capability, indicating the satisfaction of one of the selection criteria to be used as starter culture for fermented sausages. Copyright © 2014 Elsevier Ltd. All rights reserved.

Cloning strategies for heterologous expression of the bacteriocin enterocin A by Lactobacillus sakei Lb790, Lb. plantarum NC8 and Lb. casei CECT475.

PubMed

Jiménez, Juan J; Diep, Dzung B; Borrero, Juan; Gútiez, Loreto; Arbulu, Sara; Nes, Ingolf F; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2015-10-15

Bacteriocins produced by lactic acid bacteria (LAB) attract considerable interest as natural and nontoxic food preservatives and as therapeutics whereas the bacteriocin-producing LAB are considered potential probiotics for food, human and veterinary applications, and in the animal production field. Within LAB the lactobacilli are increasingly used as starter cultures for food preservation and as probiotics. The lactobacilli are also natural inhabitants of the gastrointestinal (GI) tract and attractive vectors for delivery of therapeutic peptides and proteins, and for production of bioactive peptides. Research efforts for production of bacteriocins in heterologous hosts should be performed if the use of bacteriocins and the LAB bacteriocin-producers is ever to meet the high expectations deposited in these antimicrobial peptides. The recombinant production and functional expression of bacteriocins by lactobacilli would have an additive effect on their probiotic functionality. The heterologous production of the bacteriocin enterocin A (EntA) was evaluated in different Lactobacillus spp. after fusion of the versatile Sec-dependent signal peptide (SP usp45 ) to mature EntA plus the EntA immunity gene (entA + entiA) (fragment UAI), and their cloning into plasmid vectors that permitted their inducible (pSIP409 and pSIP411) or constitutive (pMG36c) production. The amount, antimicrobial activity (AA) and specific antimicrobial activity (SAA) of the EntA produced by Lactobacillus sakei Lb790, Lb. plantarum NC8 and Lb. casei CECT475 transformed with the recombinant plasmids pSIP409UAI, pSIP411UAI and pMGUAI varied depending of the expression vector and the host strain. The Lb. casei CECT475 recombinant strains produced the largest amounts of EntA, with the highest AA and SAA. Supernatants from Lb. casei CECT (pSIP411UAI) showed a 4.9-fold higher production of EntA with a 22.8-fold higher AA and 4.7-fold higher SAA than those from Enterococcus faecium T136, the natural

Advantageous Direct Quantification of Viable Closely Related Probiotics in Petit-Suisse Cheeses under In Vitro Gastrointestinal Conditions by Propidium Monoazide - qPCR

PubMed Central

Villarreal, Martha Lissete Morales; Padilha, Marina; Vieira, Antonio Diogo Silva; Franco, Bernadette Dora Gombossy de Melo; Martinez, Rafael Chacon Ruiz; Saad, Susana Marta Isay

2013-01-01

Species-specific Quantitative Real Time PCR (qPCR) alone and combined with the use of propidium monoazide (PMA) were used along with the plate count method to evaluate the survival of the probiotic strains Lactobacillus acidophilus La-5 and Bifidobacterium animalis subsp. lactis Bb-12, and the bacteriocinogenic and potentially probiotic strain Lactobacillus sakei subsp. sakei 2a in synbiotic (F1) and probiotic (F2) petit-suisse cheeses exposed throughout shelf-life to in vitro simulated gastrointestinal tract conditions. The three strains studied showed a reduction in their viability after the 6 h assay. Bb-12 displayed the highest survival capacity, above 72.6 and 74.6% of the initial populations, respectively, by plate count and PMA-qPCR, maintaining population levels in the range or above 6 log CFU/g. The prebiotic mix of inulin and FOS did not offer any additional protection for the strains against the simulated gastrointestinal environment. The microorganisms' populations were comparable among the three methods at the initial time of the assay, confirming the presence of mainly viable and culturable cells. However, with the intensification of the stress induced throughout the various stages of the in vitro test, the differences among the methods increased. The qPCR was not a reliable enumeration method for the quantification of intact bacterial populations, mixed with large numbers of injured and dead bacteria, as confirmed by the scanning electron microscopy results. Furthermore, bacteria plate counts were much lower (P<0.05) than with the PMA-qPCR method, suggesting the accumulation of stressed or dead microorganisms unable to form colonies. The use of PMA overcame the qPCR inability to differentiate between dead and alive cells. The combination of PMA and species-specific qPCR in this study allowed a quick and unequivocal way of enumeration of viable closely related species incorporated into probiotic and synbiotic petit-suisse cheeses and under stress

Effect of starter cultures on survival of Listeria monocytogenes in Čajna sausage

NASA Astrophysics Data System (ADS)

Bošković, M.; Tadić, V.; Đorđević, J.; Glišić, M.; Lakićević, B.; Dimitrijević, M.; Baltić, M. Ž.

2017-09-01

The aim of the study was to evaluate the survival of Listeria monocytogenes during the production of Čajna sausage with short maturation time. Sausage batter was inoculated with three different serotypes 4b and serotype 1/2a of L. monocytogenes. Control sausages were without any starter culture added; the second batch was inoculated with strains of Lactobacillus sakei, Staphylococcus carnosus and Staphylococcus xylosus, and the third batch was inoculated with strains of Debaryomyces hansenii, Lactobacillus sakei, Pediococcus acidilactici, Pediococcus pentosaceus, Staphylococcus carnosus and Staphylococcus xylosus. After 18 days of ripening, L. monocytogenes was not detected in any of the sausages, but during this fermentation and drying, the numbers of this pathogen was lower in the sausages inoculated with starter cultures.

In vitro study of beneficial properties and safety of lactic acid bacteria isolated from Portuguese fermented meat products.

PubMed

Todorov, S D; Franco, B D G M; Wiid, I J

2014-09-01

Many lactic acid bacteria produce bacteriocins with a rather broad spectrum of inhibition, which could offer potential applications in food preservation. Bacteriocin production by starter cultures may bring advantage to these strains in competitive interactions with pathogenic bacteria from the food matrix. The objective of this study was to determine the safety of beneficial strains (Lactobacillus plantarum ST202Ch and ST216Ch, Enterococcus faecium ST211Ch, and Lactobacillus sakei ST22Ch, ST153Ch and ST154Ch) previously isolated from fermented meat products and characterised as bacteriocin producers. Auto-aggregation was strain-specific, and values of 28.97, 27.86 and 28.56% were recorded for L. sakei ST22Ch, ST153Ch and ST154Ch, respectively, 16.95 and 14.58% for L. plantarum ST202Ch and ST216Ch, respectively, and 12.77% for E. faecium ST211Ch. Various degrees of co-aggregation between 28.85 and 44.76% for Listeria monocytogenes 211 and 409, and between 23.60 to 34.96% for E. faecium ATCC 19443 were observed. According to the results of the diffusion method, the studied strains demonstrated susceptibility to penicillin G, ampicillin, amoxicillin, amoxicillin/clavulonic acid, imipenem, linezolid, and tetracycline. In addition, the susceptibility of the six strains to various non-antibiotic commercial drugs was examined. Production of β-galactosidase by L. sakei ST22Ch, ST153Ch and ST154Ch, L. plantarum ST202Ch and ST216Ch, and E. faecium ST211Ch was confirmed by employing sterile filter paper discs impregnated with o-nitrophenyl-β-D-galactopyranose. A statistically significant (P<0.001) inhibition of Mycobacterium tuberculosis growth by bacteriocins produced by L. plantarum ST202Ch (38.3%) and ST216Ch (48.6%), L. sakei ST153Ch (16.2%) and ST154Ch (16.1%), and E. faecium ST211Ch (21.7%) was observed. As determined by the polymerase chain reaction, the tested strains showed a low virulence gene profile.

Interactions of meat-associated bacteriocin-producing Lactobacilli with Listeria innocua under stringent sausage fermentation conditions.

PubMed

Leroy, Frédéric; Lievens, Kristoff; De Vuyst, Luc

2005-10-01

The kinetics of the antilisterial effect of meat-associated lactobacilli on Listeria innocua LMG 13568 were investigated during laboratory batch fermentations. During these fermentations, which were performed in a liquid meat simulation medium, a combination of process factors typical for European-style sausage fermentations was applied, such as a temperature of 20 degrees C and a representative pH and salting profile. Two bacteriocin-producing sausage isolates (Lactobacillus sakei CTC 494 and Lactobacillus curvatus LTH 1174), which have already proven efficacy in sausage trials, and one nonbacteriocinogenic, industrial strain (Lactobacillus sakei I), were evaluated. Staphylococcus carnosus 833 was included in the experiment because of its role in flavor and color development. When grown as a monoculture or upon cocultivation with L. sakei I and S. carnosus 833, L. innocua LMG 13568 developed slightly, despite the stress of low temperature, pH, lactic acid, salt, and nitrite. In contrast, when either of the bacteriocin producers was used, the L. innocua LMG 13568 population was rapidly inactivated with more than 3 log CFU ml(-1) after 2 days of fermentation. A bacteriocin-tolerant L. innocua LMG 13568 subpopulation (4 X 10(-4)) remained after bacteriocin inactivation. Thus, when the initial level of L. innocua LMG 13568 equaled 3 log CFU ml(-1), all cells were inactivated and no bacteriocin-tolerant cells were detected, even after 7 days of incubation. S. carnosus was not inactivated by the Lactobacillus bacteriocins and displayed slight growth.

Rapid Detection of Bacterial Antibiotic Resistance: Preliminary Evaluation of PCR Assays Targeting Tetracycline Resistance Genes

DTIC Science & Technology

2007-08-01

gonorrheae strain 2309 plasmid pOZ101; AF440277, Lactobacillus plantarum plasmid pMD5057; X75073, Neisseria meningitidis plasmid DNA for tet(M...tetracycline resistance tet(M) gene; AY057892, Staphylococcus aureus strain 1802 tetracycline resistance protein tet(M) gene; AY149596, Lactobacillus sakei

Adding Value to Goat Meat: Biochemical and Technological Characterization of Autochthonous Lactic Acid Bacteria to Achieve High-Quality Fermented Sausages

PubMed Central

Nediani, Miriam T.; García, Luis; Saavedra, Lucila; Martínez, Sandra; López Alzogaray, Soledad; Fadda, Silvina

2017-01-01

Quality and safety are important challenges in traditional fermented sausage technology. Consequently, the development of a tailored starter culture based on indigenous microbiota constitutes an interesting alternative. In the present study, spontaneously fermented goat meat sausages were created and analyzed using a physicochemical and microbiological approach. Thereafter 170 lactic acid bacteria (LAB) strains were isolated and preliminary characterized by phenotypic assays. The hygienic and technological properties, and growth and fermentative potential of isolates using a goat-meat-based culture medium were evaluated. All strains proved to have bioprotective features due to their acidogenic metabolism. Almost all grew optimally in meat environments. LAB isolates presented proteolytic activity against meat proteins and enriched amino acid contents of the goat-meat-based model. The most efficient strains were four different Lactobacillus sakei isolates, as identified by genotyping and RAPD analysis. L. sakei strains are proposed as optimal candidates to improve the production of fermented goat meat sausages, creating a new added-value fermented product. PMID:28513575

Adding Value to Goat Meat: Biochemical and Technological Characterization of Autochthonous Lactic Acid Bacteria to Achieve High-Quality Fermented Sausages.

PubMed

Nediani, Miriam T; García, Luis; Saavedra, Lucila; Martínez, Sandra; López Alzogaray, Soledad; Fadda, Silvina

2017-05-17

Quality and safety are important challenges in traditional fermented sausage technology. Consequently, the development of a tailored starter culture based on indigenous microbiota constitutes an interesting alternative. In the present study, spontaneously fermented goat meat sausages were created and analyzed using a physicochemical and microbiological approach. Thereafter 170 lactic acid bacteria (LAB) strains were isolated and preliminary characterized by phenotypic assays. The hygienic and technological properties, and growth and fermentative potential of isolates using a goat-meat-based culture medium were evaluated. All strains proved to have bioprotective features due to their acidogenic metabolism. Almost all grew optimally in meat environments. LAB isolates presented proteolytic activity against meat proteins and enriched amino acid contents of the goat-meat-based model. The most efficient strains were four different Lactobacillus sakei isolates, as identified by genotyping and RAPD analysis. L. sakei strains are proposed as optimal candidates to improve the production of fermented goat meat sausages, creating a new added-value fermented product.

Contribution of starter cultures to the proteolytic process of a fermented non-dried whole muscle ham product.

PubMed

Scannell, Amalia G M; Kenneally, Paul M; Arendt, Elke K

2004-06-01

Porcine longissimus dorsi muscles were cured by brine injection. Curing brine containing 15% (w/v) NaCl, 1.33% (w/v) glucose, 750 ppm sodium nitrite, and appropriate levels of either Lactobacillus sakei LAD, L. sakei LAD plus Kocuria varians FT4 (formally Micrococcus varians), L. sakei LAD plus papain and GDL (glucono-delta-lactone) plus K. varians FT4, was injected to the muscle at a pumping rate 15% w/v. The effect of these treatments on the proteolysis in the ham system was compared to a control ham, produced without starter culture and containing GDL acidulant to control pH and antibiotics to reduce the contribution of background microflora. Hydrolysis of sarcoplasmic and myofibrillar protein fractions was evaluated by SDS-PAGE and reverse phase-HPLC. Hams with different treatments were also investigated for differences in amino acid profile, protein and non-protein nitrogen level, colour, pH, water activity and moisture and microbiological evolution. There was no significant difference in the gross compositional analysis of any of the treatments compared to the control. There was no significant difference (p>0.05) in the protein content, non-protein nitrogen level, SDS-PAGE and free amino acid analysis between the control ham and ham inoculated with proteolytic starter culture. However, it was observed that hams containing starter cultures exhibited decreases in certain peptide fractions and corresponding increases in some free amino acids compared to the uninoculated control. It can be concluded that, while the principle mechanisms resulting in the proteolysis of this non-dried ham product involve the activity of endogeneous cathepsins, the addition of proteolytic starter cultures influence the amino acid profile thereby potentially enhancing the sensorial attributes of the ham. Copyright 2004 Elsevier B.V.

Effect of starter culture and turmeric on physico-chemical quality of carabeef pastirma.

PubMed

Maurya, P; Borpuzari, R N; Nath, D R; Nath, N C

2010-01-01

Carabeef samples were sliced, pressed, cured and divided into 6 groups. Starter cultures (Micrococcus varians M483 (MV), Staphylococcus carnosus (SC), Lactobacillus sakei (LS), M. varians M483+ Lb. sakei and Staph. carnosus + Lb. sakei) were inoculated at the dose of 10(6)-0(7)cfu/g and stored at 10 ± 1°C for 7 days. Uninoculated samples were maintained as control. Samples were then divided into 2 treatment groups. Samples of treatment 1 (T1) were smeared with a paste of turmeric followed by application of a thick layer of the paste of garlic, cumin, black pepper and red pepper whereas, samples of treatment 2 (T2) were applied with a thick layer of spices as above without turmeric. With the gradual fall in pH there was a reduction in water-holding capacity (WHC) of samples. The WHC of samples treated with SC+LS of T1 reduced to 6.3 ± 0.03 cm(2) and those inoculated with MV+LS of T2 to 6.2 ± 0.03 cm(2). The extract release volume (ERV) increased in all samples during storage. The least ERV of 11.7 and 11.6 ml were recorded in samples inoculated with MV of T1 and T2, respectively. The tyrosine (TV) and thiobarbituric acid (TBA) number of turmeric treated samples were significantly lower than non turmeric treated samples. The samples inoculated with LS had the least TV of 30.9 mg tyrosine/100 g of meat and TBA number of 0.06 mg manoladehyde/kg of meat. Samples inoculated with MV and LS of both T1 and T2 were better in physico-chemical qualities.

Competitiveness and antibacterial potential of bacteriocin-producing starter cultures in different types of fermented sausages.

PubMed

Ravyts, Frédéric; Barbuti, Silvana; Frustoli, Maria Angela; Parolari, Giovanni; Saccani, Giovanna; De Vuyst, Luc; Leroy, Frédéric

2008-09-01

Application of bacteriocin-producing starter cultures of lactic acid bacteria in fermented sausage production contributes to food safety. This is sometimes hampered by limited efficacy in situ and by uncertainty about strain dependency and universal applicability for different sausage types. In the present study, a promising antilisterial-bacteriocin producer, Lactobacillus sakei CTC 494, was applied as a coculture in addition to commercial fermentative starters in different types of dry-fermented sausages. The strain was successful in both Belgian-type sausage and Italian salami that were artificially contaminated with about 3.5 log CFU g(-1) of Listeria monocytogenes. After completion of the production process, this led to listerial reductions of up to 1.4 and 0.6 log CFU g(-1), respectively. In a control sausage, containing only the commercial fermentative starter, the reduction was limited to 0.8 log CFU g(-1) for the Belgian-type recipe, where pH decreased from 5.9 to 4.9, whereas an increase of 0.2 log CFU g(-1) was observed for Italian salami, in which the pH rose from 5.7 to 5.9 after an initial decrease to pH 5.3. In a Cacciatore recipe inoculated with 5.5 log CFU g(-1) of L. monocytogenes and in the presence of L. sakei CTC 494, there was a listerial reduction of 1.8 log CFU g(-1) at the end of the production process. This was superior to the effect obtained with the control sausage (0.8 log CFU g(-1)). Two commercial antilisterial cultures yielded reductions of 1.2 and 1.5 log CFU g(-1). Moreover, repetitive DNA sequence-based PCR fingerprinting demonstrated the competitive superiority of L. sakei CTC 494.

A synthetic promoter library for constitutive gene expression in Lactobacillus plantarum.

PubMed

Rud, Ida; Jensen, Peter Ruhdal; Naterstad, Kristine; Axelsson, Lars

2006-04-01

A synthetic promoter library (SPL) for Lactobacillus plantarum has been developed, which generalizes the approach for obtaining synthetic promoters. The consensus sequence, derived from rRNA promoters extracted from the L. plantarum WCFS1 genome, was kept constant, and the non-consensus sequences were randomized. Construction of the SPL was performed in a vector (pSIP409) previously developed for high-level, inducible gene expression in L. plantarum and Lactobacillus sakei. A wide range of promoter strengths was obtained with the approach, covering 3-4 logs of expression levels in small increments of activity. The SPL was evaluated for the ability to drive beta-glucuronidase (GusA) and aminopeptidase N (PepN) expression. Protein production from the synthetic promoters was constitutive, and the most potent promoters gave high protein production with levels comparable to those of native rRNA promoters, and production of PepN protein corresponding to approximately 10-15 % of the total cellular protein. High correlation was obtained between the activities of promoters when tested in L. sakei and L. plantarum, which indicates the potential of the SPL for other Lactobacillus species. The SPL enables fine-tuning of stable gene expression for various applications in L. plantarum.

Microbial Quality and Direct PCR Identification of Lactic Acid Bacteria and Nonpathogenic Staphylococci from Artisanal Low-Acid Sausages

PubMed Central

Aymerich, T.; Martín, B.; Garriga, M.; Hugas, M.

2003-01-01

Detection of six species of lactic acid bacteria and six species of gram-positive catalase-positive cocci from low-acid fermented sausages (fuets and chorizos) was assessed by species-specific PCR. Without enrichment, Lactobacillus sakei and Lactobacillus curvatus were detected in 11.8% of the samples, and Lactobacillus plantarum and Staphylococcus xylosus were detected in 17.6%. Enriched samples allowed the detection of L. sakei and S. xylosus in all of the samples (100%) and of Enterococcus faecium in 11.8% of the sausages. The percentages of L. curvatus, L. plantarum, Staphylococcus carnosus, and Staphylococcus epidermidis varied depending on the sausage type. L. curvatus was detected in 80% of fuets and in 57% of chorizos. L. plantarum was found in 50% of fuets and 100% of chorizos. S. epidermidis was detected in only 11.8% of fuets, and S. carnosus was detected in only 5.9% of chorizos. Lactococcus lactis, Staphylococcus warneri, and Staphylococcus simulans were not detected in any sausage type. From a microbiological point of view, 70.6% of the samples could be considered of high quality, as they had low counts of Enterobacteriaceae and did not contain any of the food-borne pathogens assayed. PMID:12902246

Dextrans produced by lactic acid bacteria exhibit antiviral and immunomodulatory activity against salmonid viruses.

PubMed

Nácher-Vázquez, Montserrat; Ballesteros, Natalia; Canales, Ángeles; Rodríguez Saint-Jean, Sylvia; Pérez-Prieto, Sara Isabel; Prieto, Alicia; Aznar, Rosa; López, Paloma

2015-06-25

Viral infections in the aquaculture of salmonids can lead to high mortality and substantial economic losses. Thus, there is industrial interest in new molecules active against these viruses. Here we describe the production, purification, and the physicochemical and structural characterization of high molecular weight dextrans synthesized by Lactobacillus sakei MN1 and Leuconostoc mesenteroides RTF10. The purified dextrans, and commercial dextrans with molecular weights ranging from 10 to 2000kDa, were assayed in infected BF-2 and EPC fish cell-line monolayers for antiviral activity. Only T2000 and dextrans from MN1 and RTF10 had significant antiviral activity. This was similar to results obtained against infectious pancreatic necrosis virus. However the dextran from MN1 showed ten-fold higher activity against hematopoietic necrosis virus than T2000. In vivo assays using the MN1 polymer confirmed the in vitro results and revealed immunomodulatory activity. These results together with the high levels of dextran production (2gL(-1)) by Lb. sakei MN1, indicate the compounds potential utility as an antiviral agent in aquaculture. Copyright © 2015 Elsevier Ltd. All rights reserved.

Behavior of Salmonella spp., Listeria monocytogenes, and Staphylococcus aureus in Chouriço de Vinho, a dry fermented sausage made from wine-marinated meat.

PubMed

Díez, J García; Patarata, L

2013-04-01

Portuguese chouriço de vinho is made by drying coarsely minced meat and fat that has been previously marinated with wine (usually red), salt, and garlic for 1 to 2 days at a low temperature (4 to 8 °C). This procedure may improve the microbiological safety of the product. The aim of this study was to evaluate the behavior of three pathogens in this product, Salmonella spp., Listeria monocytogenes, and Staphylococcus aureus, to establish the minimum period of drying and maturation necessary to render safe products. The pathogens were inoculated in the chouriço de vinho batter. A factorial design was used to study the following variables in the fermentation process: (i) the presence or absence of an indigenous Lactobacillus sakei starter culture; (ii) the presence or absence of fermentable carbohydrates; and (iii) the salt level (1.5 or 3%). The samples were analyzed 24 h after the preparation of the batter (at stuffing); after 7, 15, and 30 days of drying; and after 30 days of storage at 4 °C under vacuum. Under all of the conditions studied, the levels of the three pathogens decreased during the drying period. In the early stages of drying, the addition of L. sakei starter culture and/or carbohydrates resulted in lower levels of gram-positive pathogens. After 15 days of drying, populations of all pathogens decreased by ca. 2 log in all samples. At that sampling time, L. monocytogenes was undetectable in the chouriço de vinho with L. sakei starter culture and carbohydrates. The mean count of S. aureus after 15 days of drying was below 1 log CFU/g. After 30 days of drying, no pathogens were detected. The drying period could be shortened to 15 days when considering only the gram-positive pathogens studied and the use of a starter culture and carbohydrates. Due to the low infective dose of Salmonella spp., the product should be considered safe after 30 days, when this pathogen became undetectable.

Indigenous Starter Cultures to Improve Quality of Artisanal Dry Fermented Sausages from Chaco (Argentina).

PubMed

Palavecino Prpich, Noelia Z; Castro, Marcela P; Cayré, María E; Garro, Oscar A; Vignolo, Graciela M

2015-01-01

Lactic acid bacteria (LAB) and coagulase negative cocci (CNC) were isolated from artisanal dry sausages sampled from the northeastern region of Chaco, Argentina. In order to evaluate their performance in situ and considering technological features of the isolated strains, two mixed selected autochthonous starter cultures (SAS) were designed: (i) SAS-1 (Lactobacillus sakei 487 + Staphylococcus vitulinus C2) and (ii) SAS-2 (L. sakei 442 + S. xylosus C8). Cultures were introduced into dry sausage manufacturing process at a local small-scale facility. Microbiological and physicochemical parameters were monitored throughout fermentation and ripening periods, while sensory attributes of the final products were evaluated by a trained panel. Lactic acid bacteria revealed their ability to colonize and adapt properly to the meat matrix, inhibiting the growth of spontaneous microflora and enhancing safety and hygienic profile of the products. Both SAS showed a beneficial effect on lipid oxidation and texture of the final products. Staphylococcus vitulinus C2, from SAS-1, promoted a better redness of the final product. Sensory profile revealed that SAS addition preserved typical sensory attributes. Introduction of these cultures could provide an additional tool to standardize manufacturing processes aiming to enhance safety and quality while keeping typical sensory attributes of regional dry fermented sausages.

Neem (Azadirachta indica A. Juss) Oil: A Natural Preservative to Control Meat Spoilage

PubMed Central

Del Serrone, Paola; Toniolo, Chiara; Nicoletti, Marcello

2015-01-01

Plant-derived extracts (PDEs) are a source of biologically-active substances having antimicrobial properties. The aim of this study was to evaluate the potential of neem oil (NO) as a preservative of fresh retail meat. The antibacterial activity of NO against Carnobacterium maltaromaticum, Brochothrix thermosphacta, Escherichia coli, Pseudomonas fluorescens, Lactobacillus curvatus and L. sakei was assessed in a broth model system. The bacterial growth inhibition zone (mm) ranged from 18.83 ± 1.18 to 30.00 ± 1.00, as was found by a disc diffusion test with 100 µL NO. The bacterial percent growth reduction ranged from 30.81 ± 2.08 to 99.70 ± 1.53 in the broth microdilution method at different NO concentrations (1:10 to 1:100,000). Viable bacterial cells were detected in experimentally-contaminated meat up to the second day after NO treatment (100 µL NO per 10 g meat), except for C. maltaromaticum, which was detected up to the sixth day by PCR and nested PCR with propidium monoazide (PMA™) dye. In comparison to the previously published results, C. maltaromaticum, E. coli, L. curvatus and L. sakei appeared more susceptible to NO compared to neem cake extract (NCE) by using a broth model system. PMID:28231186

Neem (Azadirachtaindica A. Juss) Oil: A Natural Preservative to Control Meat Spoilage.

PubMed

Del Serrone, Paola; Toniolo, Chiara; Nicoletti, Marcello

2015-01-09

Plant-derived extracts (PDEs) are a source of biologically-active substances having antimicrobial properties. The aim of this study was to evaluate the potential of neem oil (NO) as a preservative of fresh retail meat. The antibacterial activity of NO against Carnobacterium maltaromaticum , Brochothrix thermosphacta, Escherichia coli , Pseudomonas fluorescens , Lactobacillus curvatus and L. sakei was assessed in a broth model system . The bacterial growth inhibition zone (mm) ranged from 18.83 ± 1.18 to 30.00 ± 1.00, as was found by a disc diffusion test with 100 µL NO. The bacterial percent growth reduction ranged from 30.81 ± 2.08 to 99.70 ± 1.53 in the broth microdilution method at different NO concentrations (1:10 to 1:100,000). Viable bacterial cells were detected in experimentally-contaminated meat up to the second day after NO treatment (100 µL NO per 10 g meat), except for C. maltaromaticum , which was detected up to the sixth day by PCR and nested PCR with propidium monoazide (PMA™) dye. In comparison to the previously published results, C. maltaromaticum , E. coli , L. curvatus and L. sakei appeared more susceptible to NO compared to neem cake extract (NCE) by using a broth model system.

Use of hydrolysates from Atlantic cod (Gadus morhua L.) viscera as a complex nitrogen source for lactic acid bacteria.

PubMed

Aspmo, Stein Ivar; Horn, Svein Jarle; Eijsink, Vincent G H

2005-07-01

Hydrolysates of cod viscera were tested as an alternative to commonly used complex nitrogen sources (peptones and/or extracts) for the type strains of the lactic acid bacteria Lactococcus lactis, Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus casei, Lactobacillus sakei and Pediococcus pentosaceus. Comparative studies with MRS-like media containing different nitrogen sources showed that all the fish hydrolysates performed equally well or better than commercial extracts/peptones for all selected lactic acid bacteria.

The secreted L-arabinose isomerase displays anti-hyperglycemic effects in mice.

PubMed

Rhimi, Moez; Bermudez-Humaran, Luis G; Huang, Yuan; Boudebbouze, Samira; Gaci, Nadia; Garnier, Alexandrine; Gratadoux, Jean-Jacques; Mkaouar, Héla; Langella, Philippe; Maguin, Emmanuelle

2015-12-21

The L-arabinose isomerase is an intracellular enzyme which converts L-arabinose into L-ribulose in living systems and D-galactose into D-tagatose in industrial processes and at industrial scales. D-tagatose is a natural ketohexose with potential uses in pharmaceutical and food industries. The D-galactose isomerization reaction is thermodynamically equilibrated, and leads to secondary subproducts at high pH. Therefore, an attractive L-arabinose isomerase should be thermoactive and acidotolerant with high catalytic efficiency. While many reports focused on the set out of a low cost process for the industrial production of D-tagatose, these procedures remain costly. When compared to intracellular enzymes, the production of extracellular ones constitutes an interesting strategy to increase the suitability of the biocatalysts. The L-arabinose isomerase (L-AI) from Lactobacillus sakei was expressed in Lactococcus lactis in fusion with the signal peptide of usp45 (SP(Usp45)). The L-AI protein and activity were detected only in the supernatant of the induced cultures of the recombinant L. lactis demonstrating the secretion in the medium of the intracellular L. sakei L-AI in an active form. Moreover, we showed an improvement in the enzyme secretion using either (1) L. lactis strains deficient for their two major proteases, ClpP and HtrA, or (2) an enhancer of protein secretion in L. lactis fused to the recombinant L-AI with the SP(Usp45). Th L-AI enzyme secreted by the recombinant L. lactis strains or produced intracellularly in E. coli, showed the same functional properties than the native enzyme. Furthermore, when mice are fed with the L. lactis strain secreting the L-AI and galactose, tagatose was produced in vivo and reduced the glycemia index. We report for the first time the secretion of the intracellular L-arabinose isomerase in the supernatant of food grade L. lactis cultures with hardly display other secreted proteins. The secreted L-AI originated from the food

Dissecting the taxonomic heterogeneity within Propionibacterium acnes: proposal for Propionibacterium acnes subsp. acnes subsp. nov. and Propionibacterium acnes subsp. elongatum subsp. nov.

PubMed

Dekio, Itaru; Culak, Renata; Misra, Raju; Gaulton, Tom; Fang, Min; Sakamoto, Mitsuo; Ohkuma, Moriya; Oshima, Kenshiro; Hattori, Masahira; Klenk, Hans-Peter; Rajendram, Dunstan; Gharbia, Saheer E; Shah, Haroun N

2015-12-01

Propionibacterium acnes subsp. acnes subsp. nov. and Propionibacterium acnes subsp. elongatum subsp. nov. are described. These emanate from the three known phylotypes of P. acnes, designated types I, II and III. Electron microscopy confirmed the filamentous cell shape of type III, showing a striking difference from types I/II, which were short rods. Biochemical tests indicated that, in types I/II, either the pyruvate, l-pyrrolidonyl arylamidase or d-ribose 2 test was positive, whereas all of these were negative among type III strains. Matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectra, which profile mainly their ribosomal proteins, were different between these two groups. Surface-enhanced laser-desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) spectra of all phylotypes revealed a specific protein biomarker that was overexpressed in type III strains compared with types I/II only when grown aerobically. Reference strains had high whole-genome similarity between types I (>91 %) and II (>75 %), but a considerably lower level of 72 % similarity with type III. recA and gyrB sequence dendrograms confirmed the distant relatedness of type III, indicating the presence of two distinct centres of variation within the species P. acnes. On the other hand, cellular fatty acid profiles and 16S rRNA gene sequence relatedness (>99.3 %) circumscribed the species. Thus, we propose two subspecies, Propionibacterium acnes subsp. acnes subsp. nov. for types I/II and Propionibacterium acnes subsp. elongatum subsp. nov. for type III. The type strain of Propionibacterium acnes subsp. acnes is NCTC 737T ( = ATCC 6919T = JCM 6425T = DSM 1897T = CCUG 1794T), while the type strain of Propionibacterium acnes subsp. elongatum is K124T ( = NCTC 13655T = JCM 18919T).

Production of a Suitable Antibody and an Enzyme Immunoassay Kit for the Field Detection of T-2 Toxin

DTIC Science & Technology

1983-10-25

Known to naturally occur on plants including food products. £> These trichothecenes are highly toxic to eukaryotic cells and are anong the most...Activities and Detection of Naturally O* Occurring 12,13-Epoxy-9-Trichothecenes," Clin. Tbxicol., 5, pp. 495-515. E* 3. Y. Ueno, M. Makajima, K. Sakei, K...North-Holland, N.Y. -13- i 6. S. V. Pathre, C. J. Mirocha (1977) Assay Methods for Trichothecenes and J; Review of Their Natural Occurence in: J. V

Effect of proteolytic starter cultures as leavening agents of pizza dough.

PubMed

Pepe, O; Villani, F; Oliviero, D; Greco, T; Coppola, S

2003-08-01

Lactic acid bacteria (LAB) and yeasts were selected on the basis of in vitro proteolytic activity against wheat gluten protein and then assayed as leavening agents for pizza dough. Trials were carried out to compare a proteolytic starter (Prt(+)), consisting of Lactobacillus sakei T56, Weissella paramesenteroides A51 and Candida krusei G271, and a non-proteolytic starter (Prt(-)), consisting of Lb. sakei T58, W. paramesenteroides A58 and Saccharomyces cerevisiae T22. The proteolytic activity of the starter cultures was monitored immediately after mixing of the dough and throughout the fermentation process. The proteolytic activity was assessed by analysing the salt-soluble protein (SSP) and the dioxane-soluble protein (DSP) fractions of the pizza dough by discontinuous SDS-PAGE. Only the Prt(+) starter exhibited considerable qualitative and quantitative changes in the electrophoretic patterns of the protein fractions extracted. After the fermentation, the Prt(+) and Prt(-) doughs were tested to evaluate the influence of the proteolytic activity on the mechanical properties of the dough before and after baking. Indications emerged suggesting an influence of the proteolytic activity on the viscoelasticity of pizza dough. The pizza dough with Prt(+) strains showed an increase in viscous properties during the fermentation as compared with the Prt(-) dough. Moreover, an increase in the firmness of the crumb was observed in Prt(+) baked pizza dough.

Campylobacter fetus subsp. testudinum subsp. nov., isolated from humans and reptiles.

PubMed

Fitzgerald, Collette; Tu, Zheng Chao; Patrick, Mary; Stiles, Tracy; Lawson, Andy J; Santovenia, Monica; Gilbert, Maarten J; van Bergen, Marcel; Joyce, Kevin; Pruckler, Janet; Stroika, Steven; Duim, Birgitta; Miller, William G; Loparev, Vladimir; Sinnige, Jan C; Fields, Patricia I; Tauxe, Robert V; Blaser, Martin J; Wagenaar, Jaap A

2014-09-01

A polyphasic study was undertaken to determine the taxonomic position of 13 Campylobacter fetus-like strains from humans (n = 8) and reptiles (n = 5). The results of matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MS and genomic data from sap analysis, 16S rRNA gene and hsp60 sequence comparison, pulsed-field gel electrophoresis, amplified fragment length polymorphism analysis, DNA-DNA hybridization and whole genome sequencing demonstrated that these strains are closely related to C. fetus but clearly differentiated from recognized subspecies of C. fetus. Therefore, this unique cluster of 13 strains represents a novel subspecies within the species C. fetus, for which the name Campylobacter fetus subsp. testudinum subsp. nov. is proposed, with strain 03-427(T) ( = ATCC BAA-2539(T) = LMG 27499(T)) as the type strain. Although this novel taxon could not be differentiated from C. fetus subsp. fetus and C. fetus subsp. venerealis using conventional phenotypic tests, MALDI-TOF MS revealed the presence of multiple phenotypic biomarkers which distinguish Campylobacter fetus subsp. testudinum subsp. nov. from recognized subspecies of C. fetus.

Staphylococcus petrasii subsp. pragensis subsp. nov., occurring in human clinical material.

PubMed

Švec, Pavel; De Bel, Annelies; Sedláček, Ivo; Petráš, Petr; Gelbíčová, Tereza; Černohlávková, Jitka; Mašlanˇová, Ivana; Cnockaert, Margo; Varbanovová, Ivana; Echahidi, Fedoua; Vandamme, Peter; Pantuček, Roman

2015-07-01

Seven coagulase-negative, oxidase-negative and novobiocin-susceptible staphylococci assigned tentatively as Staphylococcus petrasii were investigated in this study in order to elucidate their taxonomic position. All strains were initially shown to form a genetically homogeneous group separated from remaining species of the genus Staphylococcus by using a repetitive sequence-based PCR fingerprinting with the (GTG)5 primer. Phylogenetic analysis based on 16S rRNA gene, hsp60, rpoB, dnaJ, gap and tuf sequences showed that the group is closely related to Staphylococcus petrasii but separated from the three hitherto known subspecies, S. petrasii subsp. petrasii, S. petrasii subsp. croceilyticus and S. petrasii subsp. jettensis. Further investigation using automated ribotyping, MALDI-TOF mass spectrometry, fatty acid methyl ester analysis, DNA-DNA hybridization and extensive biotyping confirmed that the analysed group represents a novel subspecies within S. petrasii, for which the name Staphylococcus petrasii subsp. pragensis subsp. nov. is proposed. The type strain is NRL/St 12/356(T) ( = CCM 8529(T) = LMG 28327(T)).

Species diversity and metabolic impact of the microbiota are low in spontaneously acidified Belgian sausages with an added starter culture of Staphylococcus carnosus.

PubMed

Janssens, M; Myter, N; De Vuyst, L; Leroy, F

2012-04-01

Quality of fermented sausages is affected by acidifying lactic acid bacteria (LAB) and colour- and flavour-promoting coagulase-negative staphylococci (CNS), whether or not used as starter culture. Artisan fermented sausages are often perceived as superior to industrial variants, partially because of the specific microbiota due to spontaneous acidification, which may be considered as an artisan characteristic. Therefore, two kinds of spontaneously acidified Belgian sausages were prepared (Belgian-type salami and Boulogne sausage), but with addition of a Staphylococcus carnosus culture. The Belgian-type salami was made from pork and beef, whereas the Boulogne sausage contained pork and horse meat. In all cases, Lactobacillus sakei was the dominant LAB species present on the raw materials and during fermentation, whereas enterococci remained present in the background. Enterobacteriaceae vanished after fermentation. The CNS species diversity on the raw materials was large and differed between the pork, beef, and horse meat. Nevertheless, this species diversity was annihilated during fermentation by the added S. carnosus culture. The volatiles fraction was mainly composed of aldehydes that originated from lipid oxidation and spices-derived compounds. Aromatic compounds that are typically associated to CNS activity, such as end-products from the metabolism of branched-chain amino acids, were not present in the Belgian-type salami and only marginally present in the Boulogne sausage. In conclusion, spontaneous acidification of Belgian-type fermented sausages leads to dominance of L. sakei and is no guarantee for bacterial contribution to the aroma profile when S. carnosus is added as a starter culture. Copyright © 2011 Elsevier Ltd. All rights reserved.

A peptidomic approach of meat protein degradation in a low-sodium fermented sausage model using autochthonous starter cultures.

PubMed

de Almeida, Marcio Aurélio; Saldaña, Erick; da Silva Pinto, Jair Sebastião; Palacios, Jorge; Contreras-Castillo, Carmen J; Sentandreu, Miguel Angel; Fadda, Silvina Graciela

2018-07-01

Fermented sausage technology is currently compromised in decreasing the addition of NaCl. Use of starter cultures with peptidogenic potential could be a valuable strategy that can mask or hide off flavors produced by the use of NaCl substituents. In the present work, the peptidogenic potential of four lactic acid bacteria species was evaluated in a low-sodium beaker sausage (BS) model. Using a peptidomic approach, a total of 86 low molecular weight (LMW) peptides were accurately identified, mostly derived from myofibrillar proteins, especially actin, which generated 53 peptides. The BS inoculated with L. curvatus CRL705 generated 56 LMW peptides, followed by Enterococcus (E.) mundtii CRL35 with 43 peptides. In addition, BS inoculated with Lactobacillus (L.) plantarum and with L. sakei produced higher amino acid amounts over time as compared to the rest of BS models, highlighting the importance of both, time and sample effect on the overall free amino acid generation. The presence of each LAB strain in BS models generated a unique profile of small peptides and amino acids that could serve as a distinctive biochemical trait to differentiate specific fermented products. According to these results, E. mundtii and L. sakei, which are compatible between them, are proposed as the most efficiently adapted to low-sodium conditions. The use of selected strains during the processing of low-sodium fermented sausages could have a positive effect on the production of small peptides and free amino acids. Copyright © 2018 Elsevier Ltd. All rights reserved.

Combination of culture-dependent and culture-independent molecular methods for the determination of lactic microbiota in sucuk.

PubMed

Kesmen, Z; Yetiman, A E; Gulluce, A; Kacmaz, N; Sagdic, O; Cetin, B; Adiguzel, A; Sahin, F; Yetim, H

2012-02-15

In this study, the culture-dependent and culture-independent molecular methods were used for the identification of lactic acid bacteria (LAB) in sucuk a Turkish fermented dry sausage. On the one hand, the PCR-DGGE method targetting the V1 and V3 regions of 16S DNA was applied to DNA that was directly extracted from sucuk samples. On the other hand, rep-PCR fingerprinting was performed for the primary differentiation and grouping of the isolates, and the results were confirmed by sequencing of the 16S rDNA and 16S-23S rDNA intergenic spacer region. As a result of the PCR-DGGE analysis of all the samples, total 8 different lactic acid bacteria were identified, and Lactobacillus sakei, Lactobacillus curvatus and Weissella viridescens were the dominant microbiota among these bacteria. The culture-dependent approach indicated that the majority of the strains belonged to the Lactobacillus genera including Lb. sakei, Lactobacillus plantarum, Lb. curvatus, Lactobacillus brevis, Lactobacillus farciminis and Lactobacillus alimentarius. However, Leuconostoc and Weisella were also detected as minor genera. Again, Lactococcus piscium, Weissella halotolerans, Staphylococcus succinus and the comigrated Staphylococcus piscifermentans/Staphylococcus condimenti/Staphylococcus carnosus group were detected only with the culture-independent method while Lb. plantarum, Leuconostoc mesenteroides and Leuconostoc citreum were identified only by using the culture-dependent method. In the results, it was concluded that the combination of culture-dependent and culture-independent methods was necessary for reliable and detailed investigation of LAB communities in fermented food products. Copyright © 2011 Elsevier B.V. All rights reserved.

In vitro testing of commercial and potential probiotic lactic acid bacteria.

PubMed

Jensen, Hanne; Grimmer, Stine; Naterstad, Kristine; Axelsson, Lars

2012-02-01

Probiotics are defined as live microorganisms which when administered in adequate amounts confer a health benefit on the host. The objective of this study was to investigate the diversity of selected commercial and potential probiotic lactic acid bacteria using common in vitro screening assays such as transit tolerance in the upper human gastrointestinal tract, adhesion capacity to human intestinal cell lines and effect on epithelial barrier function. The selected bacteria include strains of Lactobacillus plantarum, Lactobacillus pentosus, Lactobacillus farciminis, Lactobacillus sakei, Lactobacillus gasseri, Lactobacillus rhamnosus, Lactobacillus reuteri and Pediococcus pentosaceus. Viable counts after simulated gastric transit tolerance showed that L. reuteri strains and P. pentosaceus tolerate gastric juice well, with no reduction of viability, whereas L. pentosus, L. farciminis and L. sakei strains lost viability over 180min. All strains tested tolerate the simulated small intestinal juice well. The bacterial adhesion capacity to human intestinal cells revealed major species and strain differences. Overall, L. plantarum MF1298 and three L. reuteri strains had a significant higher adhesion capacity compared to the other strains tested. All strains, both living and UV-inactivated, had little effect on the epithelial barrier function. However, living L. reuteri strains revealed a tendency to increase the transepithelial electrical resistance (TER) from 6 to 24h. This work demonstrates the diversity of 18 potential probiotic bacteria, with major species and strain specific effects in the in vitro screening assays applied. Overall, L. reuteri strains reveal some interesting characteristics compared to the other strains investigated. Copyright © 2011 Elsevier B.V. All rights reserved.

Molecular Characterization of Copper Resistance Genes from Xanthomonas citri subsp. citri and Xanthomonas alfalfae subsp. citrumelonis▿

PubMed Central

Behlau, Franklin; Canteros, Blanca I.; Minsavage, Gerald V.; Jones, Jeffrey B.; Graham, James H.

2011-01-01

Copper sprays have been widely used for control of endemic citrus canker caused by Xanthomonas citri subsp. citri in citrus-growing areas for more than 2 decades. Xanthomonas alfalfae subsp. citrumelonis populations were also exposed to frequent sprays of copper for several years as a protective measure against citrus bacterial spot (CBS) in Florida citrus nurseries. Long-term use of these bactericides has led to the development of copper-resistant (Cur) strains in both X. citri subsp. citri and X. alfalfae subsp. citrumelonis, resulting in a reduction of disease control. The objectives of this study were to characterize for the first time the genetics of copper resistance in X. citri subsp. citri and X. alfalfae subsp. citrumelonis and to compare these organisms to other Cur bacteria. Copper resistance determinants from X. citri subsp. citri strain A44(pXccCu2) from Argentina and X. alfalfae subsp. citrumelonis strain 1381(pXacCu2) from Florida were cloned and sequenced. Open reading frames (ORFs) related to the genes copL, copA, copB, copM, copG, copC, copD, and copF were identified in X. citri subsp. citri A44. The same ORFs, except copC and copD, were also present in X. alfalfae subsp. citrumelonis 1381. Transposon mutagenesis of the cloned copper resistance determinants in pXccCu2 revealed that copper resistance in X. citri subsp. citri strain A44 is mostly due to copL, copA, and copB, which are the genes in the cloned cluster with the highest nucleotide homology (≥92%) among different Cur bacteria. PMID:21515725

The characterization of lactic acid bacteria isolated during the traditional production of Užička sausage

NASA Astrophysics Data System (ADS)

Borović, B.; Velebit, B.; Vesković, S.; Lakićević, B.; Baltić, T.

2017-09-01

Užička sausage is a traditionally fermented dry sausage that is produced in western Serbia. It is made of beef and pork with the addition of solid fat and natural spices. The whole manufacturing process lasted for 21 days. The goal of this study was to create a collection of lactic acid bacteria isolated during the ripening and identify them using molecular methods. A total of 50 isolates from different stages of ripening (fermentation and drying) were identified by molecular methods. Leuconococcus mesenteroides, Lactobacillus brevis, and Lactobacillus sakei were the predominant microorganisms in Užička sausage.

[Distiller Yeasts Producing Antibacterial Peptides].

PubMed

Klyachko, E V; Morozkina, E V; Zaitchik, B Ts; Benevolensky, S V

2015-01-01

A new method of controlling lactic acid bacteria contamination was developed with the use of recombinant Saccharomyces cerevisiae strains producing antibacterial peptides. Genes encoding the antibacterial peptides pediocin and plantaricin with codons preferable for S. cerevisiae were synthesized, and a system was constructed for their secretory expression. Recombinant S. cerevisiae strains producing antibacterial peptides effectively inhibit the growth of Lactobacillus sakei, Pediacoccus pentasaceus, Pediacoccus acidilactici, etc. The application of distiller yeasts producing antibacterial peptides enhances the ethanol yield in cases of bacterial contamination. Recombinant yeasts producing the antibacterial peptides pediocin and plantaricin can successfully substitute the available industrial yeast strains upon ethanol production.

Microbial Ecology Dynamics during Rye and Wheat Sourdough Preparation

PubMed Central

Ercolini, Danilo; Pontonio, Erica; De Filippis, Francesca; Minervini, Fabio; La Storia, Antonietta; Gobbetti, Marco

2013-01-01

The bacterial ecology during rye and wheat sourdough preparation was described by 16S rRNA gene pyrosequencing. Viable plate counts of presumptive lactic acid bacteria, the ratio between lactic acid bacteria and yeasts, the rate of acidification, a permutation analysis based on biochemical and microbial features, the number of operational taxonomic units (OTUs), and diversity indices all together demonstrated the maturity of the sourdoughs during 5 to 7 days of propagation. Flours were mainly contaminated by metabolically active genera (Acinetobacter, Pantoea, Pseudomonas, Comamonas, Enterobacter, Erwinia, and Sphingomonas) belonging to the phylum Proteobacteria or Bacteroidetes (genus Chryseobacterium). Their relative abundances varied with the flour. Soon after 1 day of propagation, this population was almost completely inhibited except for the Enterobacteriaceae. Although members of the phylum Firmicutes were present at very low or intermediate relative abundances in the flours, they became dominant soon after 1 day of propagation. Lactic acid bacteria were almost exclusively representative of the Firmicutes by this time. Weissella spp. were already dominant in rye flour and stably persisted, though they were later flanked by the Lactobacillus sakei group. There was a succession of species during 10 days of propagation of wheat sourdoughs. The fluctuation between dominating and subdominating populations of L. sakei group, Leuconostoc spp., Weissella spp., and Lactococcus lactis was demonstrated. Other subdominant species such as Lactobacillus plantarum were detectable throughout propagation. As shown by PCR-denaturing gradient gel electrophoresis (PCR-DGGE) analysis, Saccharomyces cerevisiae dominated throughout the sourdough propagation. Notwithstanding variations due to environmental and technology determinants, the results of this study represent a clear example of how the microbial ecology evolves during sourdough preparation. PMID:24096427

Comparison of vaginal microbial community structure in healthy and endometritis dairy cows by PCR-DGGE and real-time PCR.

PubMed

Wang, Jun; Sun, Chengtao; Liu, Chang; Yang, Yujiang; Lu, Wenfa

2016-04-01

The normal vaginal microflora provides protection against infections of the reproductive tract. Previous studies have focused on the isolation and screening of probiotic strains from the vagina of cows; however, the vaginal microflora of postpartum cows is poorly characterized. The present study was conducted to evaluate and characterize the vaginal microflora of healthy postpartum cows in relation to postpartum cows with endometritis by using PCR followed by denaturing gradient gel electrophoresis (PCR-DGGE) and Real-time PCR. The study population comprised 5 healthy cows and 5 cows with endometritis. The results indicated that the vaginal bacterial microflora of healthy postpartum cows was dominated by Lactobacillus sakei subsp. and Weissella koreensis, while there were no dominant bacterial species in the vaginal microflora of postpartum cows with endometritis. Common microorganisms such as Bacteroides spp., Fusobacterium spp., Enterococcus spp., Prevotella spp., Clostridium perfringens strains, and Escherichia coli were detected in both groups of cows by Real-time PCR. The bacterial diversity in the vagina of cows with endometritis was significantly higher than that in healthy cows. The results indicated that the vaginal microflora of cows with endometritis was more diverse and lacked dominant bacterial species as compared to that of the healthy cows, suggesting that disruption of the normal vaginal microflora may contribute to the onset of endometritis. This microbial community analysis provided information that might be used to develop probiotics to treat endometritis in cows; however, further investigation is needed. Copyright © 2015 Elsevier Ltd. All rights reserved.

Diversity of lactic acid bacteria in sian-sianzih (fermented clams), a traditional fermented food in Taiwan.

PubMed

Chen, Yi-sheng; Wu, Hui-chung; Li, Ya-han; Leong, Kun-hon; Pua, Xiao-hui; Weng, Ming-kai; Yanagida, Fujitoshi

2012-01-30

Sian-sianzih (fermented clams) is a popular traditional fermented food in Taiwan. The lactic acid bacteria (LAB) microflora in sian-sianzih have not been studied in detail. In this study, LAB from sian-sianzih were isolated, characterized and identified. A total of 186 cultures of LAB were isolated from seven sian-sianzih samples and 29 cultures were isolated from its main raw substrate: clams. The identification results revealed up to 11 distinct bacterial species belonging to five genera in sian-sianzih, and three species belonging to two genera in clams. The most common bacterial genera in sian-sianzih were Lactobacillus and Weissella, followed by Leuconostoc, Pediococcus and Lactococcus. A regional similarity in LAB, with differences in diversity, was observed in the current study. On the other hand, Lactococcus lactis subsp. lactis was the most common species found in raw clam samples. The results also suggested that greater LAB diversity could be observed in wild clams than in cultured ones. Furthermore, antibacterial activities of the isolates were determined, and one Weisella hellenica strain showed inhibitory activity against the indicator strain Lactobacilluas sakei JCM 1157(T) . A sensory assessment of seven sian-sianzih samples was also performed and the results indicated that diversity of LAB has a great effect on its aroma and taste formation. The results demonstrate that various LAB species are distributed in sian-sianzih and have a great effect on the flavor of sian-sianzih. Copyright © 2011 Society of Chemical Industry.

Lactococcus lactis subsp. tructae subsp. nov. isolated from the intestinal mucus of brown trout (Salmo trutta) and rainbow trout (Oncorhynchus mykiss).

PubMed

Pérez, Tania; Balcázar, José Luis; Peix, Alvaro; Valverde, Angel; Velázquez, Encarna; de Blas, Ignacio; Ruiz-Zarzuela, Imanol

2011-08-01

The species Lactococcus lactis currently includes three subspecies; L. lactis subsp. lactis and L. lactis subsp. cremoris, isolated from milk sources, and L. lactis subsp. hordniae, isolated from the leafhopper Hordnia circellata. In this study, three strains, designated L105(T), I3 and L101, were isolated from the intestinal mucus of brown trout (Salmo trutta) and rainbow trout (Oncorhynchus mykiss). These strains were closely related to members of the species Lactococcus lactis. Strain L105(T) showed 99.4 % 16S rRNA gene sequence similarity to that of the type strains L. lactis subsp. lactis NCDO 604(T) and L. lactis subsp. hordniae NCDO 2181(T) and showed 99.9 % similarity to the type strain Lactococcus lactis subsp. cremoris NCDO 607(T). Analysis of two housekeeping genes, rpoB and recA, confirmed the close relationship between the novel strains and L. lactis subsp. cremoris with similarities of 99.3 and 99.7 %, respectively. The three strains could, however, be differentiated from their closest relatives on the basis of several phenotypic characteristics, as was the case for L. lactis subsp. lactis and L. lactis subsp. hordniae, which were also closely related on the basis of 16S rRNA, rpoB and recA gene sequence similarities. The strains isolated in this study represent a new subspecies, for which the name Lactococcus lactis subsp. tructae subsp. nov. is proposed. The type strain is L105(T) ( = LMG 24662(T)  = DSM 21502(T)).

[Resistance of Lactobacillus casei subsp. casei SY13 and Lactobacillus delbrueckii subsp. bulgaricus LJJ to reactive oxygen species].

PubMed

Zhang, Shuwen; Lv, Jiaping; Menghe, Bilige; Zhang, Heping; Zhang, Liyu; Song, Jinhui; Wang, Zhifei

2009-02-01

We evaluated antioxidative effect of two antioxidative strains, isolated from the traditional fermented dairy products. Both intact cells and cell-free extract of Lactobacillus casei subsp. casei SY13 and Lactobacillus delbrueckii subsp. bulgaricus LJJ were used to study the inhibited effect of linoleic acid peroxidation, the ability of scavenging 1,1-diphenyl-2-picrylhydrazyl radical, hydroxyl radical, superoxide anion radical,the ability of tolerancing hydrogen peroxide and the chelating capacity of ferrous ion and reducting activity. Lactobacillus casei subsp. casei SY13 and Lactobacillus delbrueckii subsp. bulgaricus LJJ demonstrated highest inhibition on linoleic acid peroxidation by 62.95% and 66.16%, respectively. The cell-free extract showed excellent scavenging superoxide anion and hydroxyl radicals activity. However, the intact cells of Lactobacillus delbrueckii subsp. bulgaricus LJJ scavenging superoxide and hydroxyl radicals capacity were not detected. The intact cells of Lactobacillus casei subsp. casei SY13 and Lactobacillus delbrueckii subsp. bulgaricus LJJ on 1,1-diphenyl-2-picrylhydrazyl radical scavenging ability and chelating ferrous ion capacity were superior to cell-free extract. The highest reduced activety was equivalent to 305 micromol/L and 294 micromol/L L-cysteine. Two latobacilli strains had good antioxidant capacity. As potential probiotics, it can be used in future.

Use of PCR-Based Methods for Rapid Differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis

PubMed Central

Torriani, Sandra; Zapparoli, Giacomo; Dellaglio, Franco

1999-01-01

Two PCR-based methods, specific PCR and randomly amplified polymorphic DNA PCR (RAPD-PCR), were used for rapid and reliable differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis. PCR with a single combination of primers which targeted the proline iminopeptidase (pepIP) gene of L. delbrueckii subsp. bulgaricus allowed amplification of genomic fragments specific for the two subspecies when either DNA from a single colony or cells extracted from dairy products were used. A numerical analysis of the RAPD-PCR patterns obtained with primer M13 gave results that were consistent with the results of specific PCR for all strains except L. delbrueckii subsp. delbrueckii LMG 6412T, which clustered with L. delbrueckii subsp. lactis strains. In addition, RAPD-PCR performed with primer 1254 provided highly polymorphic profiles and thus was superior for distinguishing individual L. delbrueckii strains. PMID:10508059

Use of PCR-based methods for rapid differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis.

PubMed

Torriani, S; Zapparoli, G; Dellaglio, F

1999-10-01

Two PCR-based methods, specific PCR and randomly amplified polymorphic DNA PCR (RAPD-PCR), were used for rapid and reliable differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis. PCR with a single combination of primers which targeted the proline iminopeptidase (pepIP) gene of L. delbrueckii subsp. bulgaricus allowed amplification of genomic fragments specific for the two subspecies when either DNA from a single colony or cells extracted from dairy products were used. A numerical analysis of the RAPD-PCR patterns obtained with primer M13 gave results that were consistent with the results of specific PCR for all strains except L. delbrueckii subsp. delbrueckii LMG 6412(T), which clustered with L. delbrueckii subsp. lactis strains. In addition, RAPD-PCR performed with primer 1254 provided highly polymorphic profiles and thus was superior for distinguishing individual L. delbrueckii strains.

Safety improvement and preservation of typical sensory qualities of traditional dry fermented sausages using autochthonous starter cultures.

PubMed

Talon, Régine; Leroy, Sabine; Lebert, Isabelle; Giammarinaro, Philippe; Chacornac, Jean-Paul; Latorre-Moratalla, Mariluz; Vidal-Carou, Carmen; Zanardi, Emanuela; Conter, Mauro; Lebecque, Annick

2008-08-15

Traditional dry fermented sausages are manufactured without addition of starter cultures in small-scale processing units, their fermentation relying on indigenous microflora. Characterisation and control of these specific bacteria are essential for the sensory quality and the safety of the sausages. The aim of this study was to develop an autochthonous starter culture that improves safety while preserving the typical sensory characteristics of traditional sausages. An autochthonous starter composed of Lactobacillus sakei, Staphylococcus equorum and Staphylococcus succinus isolated from a traditional fermented sausage was developed. These strains were tested for their susceptibility to antibiotics and their production of biogenic amines. This starter was evaluated in situ at the French traditional processing unit where the strains had been isolated. Effects of the autochthonous starter were assessed by analysing the microbial, physico-chemical, biochemical and sensory characteristics of the sausages. Inoculation with the chosen species was confirmed using known species-specific PCR assays for L. sakei and S. equorum and a species-specific PCR assay developed in this study for S. succinus. Strains were monitored by pulse-field gel electrophoresis typing. Addition of autochthonous microbial starter cultures improved safety compared with the traditional natural fermentation of sausages, by inhibiting the pathogen Listeria monocytogenes, decreasing the level of biogenic amines and by limiting fatty acid and cholesterol oxidation. Moreover, autochthonous starter did not affect the typical sensory quality of the traditional sausages. This is the first time to our knowledge that selection, development and validation in situ of autochthonous starter cultures have been carried out, and also the first time that S. equorum together with S. succinus have been used as starter cultures for meat fermentation. Use of autochthonous starter cultures is an effective tool for limiting

Antibacterial activity of selected standard strains of lactic acid bacteria producing bacteriocins--pilot study.

PubMed

Bodaszewska-Lubas, Malgorzata; Brzychczy-Wloch, Monika; Gosiewski, Tomasz; Heczko, Piotr B

2012-10-25

In this paper, an attempt was made to evaluate the antibacterial potential of standard strains of lactic acid bacteria (LAB) producing bacteriocins of various classes, thus demonstrating various mechanisms of cell membrane damages against the Streptococcus agalactiae strains (Group B Streptococcus, GBS), depending on surface polysaccharides and surface alpha-like protein genes. Antimicrobial property of the strains of L. plantarum C 11, L. sakei DSMZ 6333, and L. lactis ATCC 11454 producing bacteriocins: JK and EF plantaricins, sakacin and nisin, respectively, against the GBS strains was evaluated. The chosen to the study GBS strains were represented by serotypes Ia, Ib, II, III, V and they had bca, epsilon, rib, alp2 or alp3 alpha-like protein genes. The experiment was conducted by means of suspension culture and the bacteria count was determined using the serial dilution method. A great ability of L. plantarum C 11 strain was proven to inhibit the GBS growth. The strain of L. sakei DSMZ 6333 did not demonstrate any ability to inhibit the growth of GBS, whereas L. lactis ATCC 11454 inhibited the growth of S. agalactiae indicator strains to a minor extent. Statistically significant differences were demonstrated between the GBS strains representing various serotypes against the antimicrobial activity of model LAB strains. The least sensitive to the activity of bacteriocins were the strains representing serotypes Ib and III, whereas the strains representing serotype II were the most sensitive. The sensitivity of the GBS strains to the antimicrobial activity of LAB was not dependent on alpha-like protein genes. Among the LAB standard strains producing bacteriocins, the strongest antimicrobial property was observed in the strain of L. plantarum C 11. Because of the generally known and verified strong antagonistic property of the strains of L. plantarum species against indicator bacteria, it is necessary to further pursue the research presented in this paper.

Identification of Lactobacillus alimentarius and Lactobacillus farciminis with 16S-23S rDNA intergenic spacer region polymorphism and PCR amplification using species-specific oligonucleotide.

PubMed

Rachman, C N; Kabadjova, P; Prévost, H; Dousset, X

2003-01-01

The restriction fragment length polymorphism (RFLP) method was used to differentiate Lactobacillus species having closely related identities in the 16S-23S rDNA intergenic spacer region (ISR). Species-specific primers for Lact. farciminis and Lact. alimentarius were designed and allowed rapid identification of these species. The 16S-23S rDNA spacer region was amplified by primers tAla and 23S/p10, then digested by HinfI and TaqI enzymes and analysed by electrophoresis. Digestion by HinfI was not sufficient to differentiate Lact. sakei, Lact. curvatus, Lact. farciminis, Lact. alimentarius, Lact. plantarum and Lact. paraplantarum. In contrast, digestion carried out by TaqI revealed five different patterns allowing these species to be distinguished, except for Lact. plantarum from Lact. paraplantarum. The 16S-23S rDNA spacer region of Lact. farciminis and Lact. alimentarius were amplified and then cloned into vector pCR(R)2.1 and sequenced. The DNA sequences obtained were analysed and species-specific primers were designed from these sequences. The specificity of these primers was positively demonstrated as no response was obtained for 14 other species tested. The species-specific primers for Lact. farciminis and Lact. alimentarius were shown to be useful for identifying these species among other lactobacilli. The RFLP profile obtained upon digestion with HinfI and TaqI enzymes can be used to discriminate Lact. farciminis, Lact. alimentarius, Lact. sakei, Lact. curvatus and Lact. plantarum. In this paper, we have established the first species-specific primer for PCR identification of Lact. farciminis and Lact. alimentarius. Both species-specific primer and RFLP, could be used as tools for rapid identification of lactobacilli up to species level.

Microbial ecology involved in the ripening of naturally fermented llama meat sausages. A focus on lactobacilli diversity.

PubMed

Fontana, Cecilia; Bassi, Daniela; López, Constanza; Pisacane, Vincenza; Otero, Maria Claudia; Puglisi, Edoardo; Rebecchi, Annalisa; Cocconcelli, Pier Sandro; Vignolo, Graciela

2016-11-07

Llama represents for the Andean regions a valid alternative to bovine and pork meat and thanks to the high proteins and low fat content; it can constitute a good product for the novel food market. In this study, culture-dependent and independent methods were applied to investigate the microbial ecology of naturally fermented llama sausages produced in Northwest Argentina. Two different production technologies of llama sausage were investigated: a pilot-plant scale (P) and an artisanal one (A). Results obtained by High-Throughput Sequencing (HTS) of 16S rRNA amplicons showed that the production technologies influenced the development of microbial communities with a different composition throughout the entire fermentation process. Both sequencing and microbiological counts demonstrated that Lactic Acid Bacteria (LAB) contributed largely to the dominant microbiota. When a total of 230 isolates were approached by RAPD-PCR, presumptive LAB strains from P production exhibited an initial variability in RAPD fingerprints switching to a single profile at the final of ripening, while A production revealed a more heterogeneous RAPD pattern during the whole fermentation process. The constant presence of Lactobacillus sakei along the fermentation in both productions was revealed by HTS and confirmed by species-specific PCR from isolated strains. The technological characterization of Lb. sakei isolates evidenced their ability to grow at 15°C, pH4.5 and 5% NaCl (95%). Most strains hydrolyzed myofibrillar and sarcoplasmic proteins. Bacteriocins encoding genes and antimicrobial resistance were found in 35% and 42.5% of the strains, respectively. An appropriate choice of a combination of autochthonous strains in a starter formulation is fundamental to improve and standardize llama sausages safety and quality. Copyright © 2016 Elsevier B.V. All rights reserved.

Emendation of Propionibacterium acnes subsp. acnes (Deiko et al. 2015) and proposal of Propionibacterium acnes type II as Propionibacterium acnes subsp. defendens subsp. nov.

PubMed

McDowell, Andrew; Barnard, Emma; Liu, Jared; Li, Huiying; Patrick, Sheila

2016-12-01

Recently, it has been proposed that strains of Propionibacterium acnes from the type III genetic division should be classified as P. acnessubsp. elongatum subsp. nov., with strains from the type I and II divisions collectively classified as P. acnessubsp. acnes subsp. nov. Under such a taxonomic re-appraisal, we believe that types I and II should also have their own separate rank of subspecies. In support of this, we describe a polyphasic taxonomic study based on the analysis of publicly available multilocus and whole-genome sequence datasets, alongside a systematic review of previously published phylogenetic, genomic, phenotypic and clinical data. Strains of types I and II form highly distinct clades on the basis of multilocus sequence analysis (MLSA) and whole-genome phylogenetic reconstructions. In silico or digital DNA-DNA similarity values also fall within the 70-80 % boundary recommended for bacterial subspecies. Furthermore, we see important differences in genome content, including the presence of an active CRISPR/Cas system in type II strains, but not type I, and evidence for increasing linkage equilibrium within the separate divisions. Key biochemical differences include positive test results for β-haemolytic, neuraminidase and sorbitol fermentation activities with type I strains, but not type II. We now propose that type I strains should be classified as P. acnessubsp. acnes subsp. nov., and type II as P. acnessubsp. defendens subsp. nov. The type strain of P. acnessubsp. acnes subsp. nov. is NCTC 737T (=ATCC 6919T=JCM 6425T=DSM 1897T=CCUG 1794T), while the type strain of P. acnessubsp. defendens subsp. nov. is ATCC 11828 (=JCM 6473=CCUG 6369).

Leuconostoc gasicomitatum is the dominating lactic acid bacterium in retail modified-atmosphere-packaged marinated broiler meat strips on sell-by-day.

PubMed

Susiluoto, Tuija; Korkeala, Hannu; Björkroth, K Johanna

2003-01-15

Lactic acid bacteria (LAB) in retail, modified-atmosphere-packaged (MAP), marinated broiler meat strips on sell-by-day were mainly identified as Leuconostoc gasicomitatum. A total of 32 packages, three to five packages of seven differently marinated broiler meat products, were studied at the end of the producer-defined shelf life (at 6 degrees C, 7-9 days depending on the manufacturer). Prior to the microbiological analyses, appearance and smell of the product was checked and pH measured. Bacteria were cultured on MRS and Tomato Juice Agar (TJA), Rogosa SL agar (SLA), Plate Count Agar (PCA) and Streptomycin Thallium Acetate Agar (STAA) for the enumeration of LAB, lactobacilli, total bacterial count and Brochothrix thermosphacta, respectively. The average CFU/g of the 32 packages was 2.3 x 10(8) on PCA. The highest bacterial average, 3.1 x 10(8), was recovered on TJA, the corresponding CFU/g averages on MRS and SLA being 2.3 x 10(8) and 1.3 x 10(8), respectively. Despite the high LAB numbers detected, radical spoilage changes such as unpleasant odor, slime production and formation of gas were not seen. B. thermosphacta did not form a significant part of the bacterial population since none of the levels exceeded the spoilage threshold level of 10(5) CFU/g reported in previous studies for this organism. In order to characterize the dominating LAB population, as many as 85, 85 and 88 colonies from MRS, TJA and SLA, respectively, were randomly picked and cultured pure. LAB were identified to species level using a 16 and 23S rDNA HindIiI RFLP (ribotyping) database. Fifty-six of the 170 isolates picked from the non-selective LAB media (MRS and TJA) were identified as L. gasicomitatum, followed by Carnobacterium divergens (41 isolates), Lactobacillus sakei and Lactobacillus curvatus subsp. melibiosus (31 isolates) and L. curvatus subsp. curvatus (20 isolates) species. SLA proved not to be completely selective for lactobacilli because the growth of Leuconostoc spp. was not

Identification of Mycobacterium avium subsp. hominissuis Isolated From Drinking Water

EPA Science Inventory

Mycobacterium avium (MA) is divided into four subspecies based primarily on host-range and consists of MA subsp. avium (birds), MA subsp. silvaticum (wood pigeons), MA subsp. paratuberculosis (broad, poorly-defined host range), and the recently described MA subsp. hominissuis (hu...

Genome sequencing identifies Listeria fleischmannii subsp. coloradonensis subsp. nov., isolated from a ranch.

PubMed

den Bakker, Henk C; Manuel, Clyde S; Fortes, Esther D; Wiedmann, Martin; Nightingale, Kendra K

2013-09-01

Twenty Listeria-like isolates were obtained from environmental samples collected on a cattle ranch in northern Colorado; all of these isolates were found to share an identical partial sigB sequence, suggesting close relatedness. The isolates were similar to members of the genus Listeria in that they were Gram-stain-positive, short rods, oxidase-negative and catalase-positive; the isolates were similar to Listeria fleischmannii because they were non-motile at 25 °C. 16S rRNA gene sequencing for representative isolates and whole genome sequencing for one isolate was performed. The genome of the type strain of Listeria fleischmannii (strain LU2006-1(T)) was also sequenced. The draft genomes were very similar in size and the average MUMmer nucleotide identity across 91% of the genomes was 95.16%. Genome sequence data were used to design primers for a six-gene multi-locus sequence analysis (MLSA) scheme. Phylogenies based on (i) the near-complete 16S rRNA gene, (ii) 31 core genes and (iii) six housekeeping genes illustrated the close relationship of these Listeria-like isolates to Listeria fleischmannii LU2006-1(T). Sufficient genetic divergence of the Listeria-like isolates from the type strain of Listeria fleischmannii and differing phenotypic characteristics warrant these isolates to be classified as members of a distinct infraspecific taxon, for which the name Listeria fleischmannii subsp. coloradonensis subsp. nov. is proposed. The type strain is TTU M1-001(T) ( =BAA-2414(T) =DSM 25391(T)). The isolates of Listeria fleischmannii subsp. coloradonensis subsp. nov. differ from the nominate subspecies by the inability to utilize melezitose, turanose and sucrose, and the ability to utilize inositol. The results also demonstrate the utility of whole genome sequencing to facilitate identification of novel taxa within a well-described genus. The genomes of both subspecies of Listeria fleischmannii contained putative enhancin genes; the Listeria fleischmannii subsp

Natural control of bacteria affecting meat quality by a neem (Azadirachta indica A. Juss) cake extract.

PubMed

Del Serrone, P; Failla, S; Nicoletti, M

2015-01-01

The antibacterial activity of an ethylacetate neem cake extract (NCE) against bacteria that affect meat quality, namely Campylobacter jejuni, Carnobacterium spp., Lactobacillus curvatus, Lactobacillus sakei and Leuconostoc sp., is reported. The antibacterial activity was detected using standardised disc diffusion and macrodilution methods. The bacterial growth inhibition zone ranged from 11.33 ± 0.58 to 22.67 ± 0.58 mm (100 μL NCE). There is significant difference between the growth inhibition zone of NCE and the control (ciprofloxacin 100 μg). The percent of bacterial growth reduction range was 79.75 ± 1.53 to 90.73 ± 1.53 (100 μg NCE) as compared with control (without NCE). NCE in different amounts counteracted the growth of all tested bacteria.

Description of Mycobacterium chelonae subsp. bovis subsp. nov., isolated from cattle (Bos taurus coreanae), emended description of Mycobacterium chelonae and creation of Mycobacterium chelonae subsp. chelonae subsp. nov.

PubMed

Kim, Byoung-Jun; Kim, Ga-Na; Kim, Bo-Ram; Jeon, Che Ok; Jeong, Joseph; Lee, Seon Ho; Lim, Ji-Hun; Lee, Seung-Heon; Kim, Chang Ki; Kook, Yoon-Hoh; Kim, Bum-Joon

2017-10-01

Three rapidly growing mycobacterial strains, QIA-37 T , QIA-40 and QIA-41, were isolated from the lymph nodes of three separate Korean native cattle, Hanwoo (Bos taurus coreanae). These strains were previously shown to be phylogenetically distinct but closely related to Mycobacterium chelonae ATCC 35752 T by taxonomic approaches targeting three genes (16S rRNA, hsp6 and rpoB) and were further characterized using a polyphasic approach in this study. The 16S rRNA gene sequences of all three strains showed 99.7 % sequence similarity with that of the M. chelonae type strain. A multilocus sequence typing analysis targeting 10 housekeeping genes, including hsp65 and rpoB, revealed a phylogenetic cluster of these strains with M. chelonae. DNA-DNA hybridization values of 78.2 % between QIA-37 T and M. chelonae indicated that it belongs to M. chelonae but is a novel subspecies distinct from M. chelonae. Phylogenetic analysis based on whole-genome sequences revealed a 95.44±0.06 % average nucleotide identity (ANI) value with M. chelonae, slightly higher than the 95.0 % ANI criterion for determining a novel species. In addition, distinct phenotypic characteristics such as positive growth at 37 °C, at which temperature M. chelonae does not grow, further support the taxonomic status of these strains as representatives of a novel subspecies of M. chelonae. Therefore, we propose an emended description of Mycobacterium chelonae, and descriptions of M. chelonae subsp. chelonae subsp. nov. and M. chelonae subsp. bovis subsp. nov. are presented; strains ATCC 35752 T (=CCUG 47445 T =CIP 104535 T =DSM 43804 T =JCM 6388 T =NCTC 946 T ) and QIA-37 T (=KCTC 39630 T =JCM 30986 T ) are the type strains of the two novel subspecies.

Clavibacter michiganensis subsp. phaseoli subsp. nov., pathogenic in bean.

PubMed

González, Ana J; Trapiello, Estefanía

2014-05-01

A yellow Gram-reaction-positive bacterium isolated from bean seeds (Phaseolus vulgaris L.) was identified as Clavibacter michiganensis by 16S rRNA gene sequencing. Molecular methods were employed in order to identify the subspecies. Such methods included the amplification of specific sequences by PCR, 16S amplified rDNA restriction analysis (ARDRA), RFLP and multilocus sequence analysis as well as the analysis of biochemical and phenotypic traits including API 50CH and API ZYM results. The results showed that strain LPPA 982T did not represent any known subspecies of C. michiganensis. Pathogenicity tests revealed that the strain is a bean pathogen causing a newly identified bacterial disease that we name bacterial bean leaf yellowing. On the basis of these results, strain LPPA 982T is regarded as representing a novel subspecies for which the name Clavibacter michiganensis subsp. phaseoli subsp. nov. is proposed. The type strain is LPPA 982T (=CECT 8144T=LMG 27667T).

Process-driven bacterial community dynamics are key to cured meat colour formation by coagulase-negative staphylococci via nitrate reductase or nitric oxide synthase activities.

PubMed

Sánchez Mainar, María; Leroy, Frédéric

2015-11-06

The cured colour of European raw fermented meats is usually achieved by nitrate-into-nitrite reduction by coagulase-negative staphylococci (CNS), subsequently generating nitric oxide to form the relatively stable nitrosomyoglobin pigment. The present study aimed at comparing this classical curing procedure, based on nitrate reductase activity, with a potential alternative colour formation mechanism, based on nitric oxide synthase (NOS) activity, under different acidification profiles. To this end, meat models with and without added nitrate were fermented with cultures of an acidifying strain (Lactobacillus sakei CTC 494) and either a nitrate-reducing Staphylococcus carnosus strain or a rare NOS-positive CNS strain (Staphylococcus haemolyticus G110), or by relying on the background microbiota. Satisfactory colour was obtained in the models prepared with added nitrate and S. carnosus. In the presence of nitrate but absence of added CNS, however, cured colour was only obtained when L. sakei CTC 494 was also omitted. This was ascribed to the pH dependency of the emerging CNS background microbiota, selecting for nitrate-reducing Staphylococcus equorum strains at mild acidification conditions but for Staphylococcus saprophyticus strains with poor colour formation capability when the pH decrease was more rapid. This reliance of colour formation on the composition of the background microbiota was further explored by a side experiment, demonstrating the heterogeneity in nitrate reduction of a set of 88 CNS strains from different species. Finally, in all batches prepared with S. haemolyticus G110, colour generation failed as the strain was systematically outcompeted by the background microbiota, even when imposing milder acidification profiles. Thus, when aiming at colour formation through CNS metabolism, technological processing can severely interfere with the composition and functionality of the meat-associated CNS communities, for both nitrate reductase and NOS activities

Evaluation of an autochthonous starter culture on the production of a traditional dry fermented sausage from Chaco (Argentina) at a small-scale facility.

PubMed

Palavecino Prpich, Noelia Z; Garro, Oscar A; Romero, Mara; Judis, María A; Cayré, María E; Castro, Marcela P

2016-05-01

The performance of a mixed starter culture, SAS-1, comprised of the autochthonous strains Lactobacillus sakei ACU-2 and Staphylococcus vitulinus ACU-10, was evaluated into the production process of a traditional dry sausage. Microbiological, physicochemical and sensory analyses were carried out to accomplish this goal. Results showed an improvement in performance through the introduction of SAS-1; adding mixed starter culture rapidly decreased pH, inhibited the growth of contaminant microorganisms and enhanced the beneficial ones, diminished TBARS, and highlighted color and aroma attributes. However, most influential organoleptic descriptors among consumer acceptance were not affected by the addition of the starter. This starter culture would represent a valuable tool to improve the homogeneity of artisanal manufacture of this traditional food. Copyright © 2016 Elsevier Ltd. All rights reserved.

Tulipa cinnabarina subsp. toprakii (Liliaceae), a new subspecies from southwestern Anatolia.

PubMed

Eker, İsmail; Yıldırım, Hasan; Altıoğlu, Yusuf

2016-01-01

A new subpecies, Tulipa cinnabarina subsp. toprakii subsp. nov. (Liliaceae) from Turkey is described. Diagnostic characters, descriptions, detailed illustrations, geographical distribution, conservation status and ecological observations on the new taxon are provided. It is also compared with the closely related Tulipa cinnabarina subsp. cinnabarina.

Tulipa cinnabarina subsp. toprakii (Liliaceae), a new subspecies from southwestern Anatolia

PubMed Central

Eker, İsmail; Yıldırım, Hasan; Altıoğlu, Yusuf

2016-01-01

Abstract A new subpecies, Tulipa cinnabarina subsp. toprakii subsp. nov. (Liliaceae) from Turkey is described. Diagnostic characters, descriptions, detailed illustrations, geographical distribution, conservation status and ecological observations on the new taxon are provided. It is also compared with the closely related Tulipa cinnabarina subsp. cinnabarina. PMID:27698585

Proposal to rename Carnobacterium inhibens as Carnobacterium inhibens subsp. inhibens subsp. nov. and description of Carnobacterium inhibens subsp. gilichinskyi subsp. nov., a psychrotolerant bacterium isolated from Siberian permafrost.

PubMed

Nicholson, Wayne L; Zhalnina, Kateryna; de Oliveira, Rafael R; Triplett, Eric W

2015-02-01

A novel, psychrotolerant facultative anaerobe, strain WN1359(T), was isolated from a permafrost borehole sample collected at the right bank of the Kolyma River in Siberia, Russia. Gram-positive-staining, non-motile, rod-shaped cells were observed with sizes of 1-2 µm long and 0.4-0.5 µm wide. Growth occurred in the range of pH 5.8-9.0 with optimal growth at pH 7.8-8.6 (pH optimum 8.2). The novel isolate grew at temperatures from 0-37 °C and optimal growth occurred at 25 °C. The novel isolate does not require NaCl; growth was observed between 0 and 8.8 % (1.5 M) NaCl with optimal growth at 0.5 % (w/v) NaCl. The isolate was a catalase-negative, facultatively anaerobic chemo-organoheterotroph that used sugars but not several single amino acids or dipeptides as substrates. The major metabolic end-product was lactic acid in the ratio of 86 % l-lactate : 14 % d-lactate. Strain WN1359(T) was sensitive to ampicillin, chloramphenicol, fusidic acid, lincomycin, monocycline, rifampicin, rifamycin SV, spectinomycin, streptomycin, troleandomycin and vancomycin, and resistant to nalidixic acid and aztreonam. The fatty acid content was predominantly unsaturated (70.2 %), branched-chain unsaturated (11.7 %) and saturated (12.5 %). The DNA G+C content was 35.3 mol% by whole genome sequence analysis. 16S rRNA gene sequence analysis showed 98.7 % sequence identity between strain WN1359(T) and Carnobacterium inhibens. Genome relatedness was computed using both Genome-to-Genome Distance Analysis (GGDA) and Average Nucleotide Identity (ANI), which both strongly supported strain WN1359(T) belonging to the species C. inhibens. On the basis of these results, the permafrost isolate WN1359(T) represents a novel subspecies of C. inhibens, for which the name Carnobacterium inhibens subsp. gilichinskyi subsp. nov. is proposed. The type strain is WN1359(T) ( = ATCC BAA-2557(T) = DSM 27470(T)). The subspecies Carnobacterium inhibens subsp. inhibens subsp. nov. is created automatically. An

The first closed genome sequence of Campylobacter fetus subsp. venerealis biovar intermedius

USDA-ARS?s Scientific Manuscript database

Campylobacter fetus venerealis biovar intermedius is a variant of Campylobacter fetus subsp. venerealis, the causative agent of Bovine Genital Campylobacteriosis. In contrast to Campylobacter fetus subsp. venerealis which is restricted to the genital tract of cattle, Campylobacter fetus subsp. vener...

Cell surface characteristics of Lactobacillus casei subsp. casei, Lactobacillus paracasei subsp. paracasei, and Lactobacillus rhamnosus strains.

PubMed Central

Pelletier, C; Bouley, C; Cayuela, C; Bouttier, S; Bourlioux, P; Bellon-Fontaine, M N

1997-01-01

Hydrophilic and electrostatic cell surface properties of eight Lactobacillus strains were characterized by using the microbial adhesion to solvents method and microelectrophoresis, respectively. All strains appeared relatively hydrophilic. The strong microbial adhesion to chloroform, an acidic solvent, in comparison with microbial adhesion to hexadecane, an apolar n-alkane, demonstrated the particularity of lactobacilli to have an important electron donor and basic character and consequently their potential ability to generate Lewis acid-base interactions with a support. Regardless of their electrophoretic mobility (EM), strains were in general slightly negatively charged at alkaline pH. A pH-dependent behavior concerning cell surface charges was observed. The EM decreased progressively with more acidic pHs for the L. casei subsp. casei and L. paracasei subsp. paracasei strains until the isoelectric point (IEP), i.e., the pH value for which the EM is zero. On the other hand, the EM for the L. rhamnosus strains was stable from pH 8 to pH 3 to 4, at which point there was a shift near the IEP. Both L. casei subsp. casei and L. paracasei subsp. paracasei strains were characterized by an IEP of around 4, whereas L. rhamnosus strains possessed a markedly lower IEP of 2. The present study showed that the cell surface physicochemical properties of lactobacilli seem to be, at least in part and under certain experimental conditions, particular to the bacterial species. Such differences detected between species are likely to be accompanied by some particular changes in cell wall chemical composition. PMID:9143109

Lactobacillus delbrueckii subsp. sunkii subsp. nov., isolated from sunki, a traditional Japanese pickle.

PubMed

Kudo, Yuko; Oki, Kaihei; Watanabe, Koichi

2012-11-01

Although four strains of bacteria isolated from sunki, a traditional Japanese, non-salted pickle, were initially identified as Lactobacillus delbrueckii, the molecular and phenotypic characteristics of the strains did not match those of any of the four recognized subspecies of L. delbrueckii. Together, the results of phenotypic characterization, DNA-DNA hybridizations (in which the relatedness values between the novel strains and type strains of the recognized subspecies of L. delbrueckii were all >88.7%) and 16S rRNA gene sequence, amplified fragment length polymorphism (AFLP) and whole-cell MALDI-TOF/MS spectral pattern analyses indicated that the four novel strains represented a single, novel subspecies, for which the name Lactobacillus delbrueckii subsp. sunkii subsp. nov. is proposed. The type strain is YIT 11221(T) (=JCM 17838(T) =DSM 24966(T)).

Regulation of the production of extracellular pectinase, cellulase, and protease in the soft rot bacterium Erwinia carotovora subsp. carotovora: evidence that aepH of E. carotovora subsp. carotovora 71 activates gene expression in E. carotovora subsp. carotovora, E. carotovora subsp. atroseptica, and Escherichia coli.

PubMed Central

Murata, H; Chatterjee, A; Liu, Y; Chatterjee, A K

1994-01-01

The production of pectolytic enzymes (pectate lyase [Pel] and polygalacturonase [Peh]), cellulase (Cel), and protease (Prt) is activated in the soft rot bacterium Erwinia carotovora subsp. carotovora by aepA (activator of extracellular protein production) and celery extract (Y. Liu, H. Murata, A. Chatterjee, and A. K. Chatterjee, Mol. Plant-Microbe Interact. 6:299-308, 1993). We recently isolated a new class of mutants of strain E. carotovora subsp. carotovora 71 which overproduces Pel, Peh, Cel, and Prt. From the overproducing strain AC5034, we identified an activator locus, designated aepH*, which stimulated Pel, Peh, Cel, and Prt production in E. carotovora subsp. carotovora 71 or its derivatives. The nucleotide sequence of the aepH* DNA segment revealed an open reading frame of 141 bp that could encode a small (5.45-kDa) highly basic (pI 11.7) protein of 47 amino acid residues. Analyses of deletions and MudI insertions indicated that the activator function required the 508-bp DNA segment which contains this open reading frame. The wild-type locus, aepH+, is localized within a DNA segment upstream of aepA. An AepH- strain constructed by exchanging aepH+ with aepH*::MudI was deficient in Pel, Peh, Cel, and Prt production; exoenzyme production was restored upon the introduction of a plasmid carrying aepH+ or aepH*. Plasmids carrying either aepH+ or aepH* activated the production of Pel-1, Peh-1, and Cel in Escherichia coli HB101 carrying the cognate genes. The aepH effect in E. coli was due to the activation of transcription, as indicated by assays of pel-1 and peh-1 mRNAs. The aepH+ and aepH* plasmids also stimulated Pel, Peh, Cel, and Prt production in other wild-type E. carotovora subsp. carotovora strains as well as in E. carotovora subsp. atroseptica. Although the stimulatory effect was generally more pronounced with aepH* than with aepH+, the extent of activation in the wild-type strains depended upon the bacterial strain and the growth medium. Southern blot

Biodegradation of chlorpyrifos by lactic acid bacteria during kimchi fermentation.

PubMed

Cho, Kye Man; Math, Reukaradhya K; Islam, Shah Md Asraful; Lim, Woo Jin; Hong, Su Young; Kim, Jong Min; Yun, Myoung Geun; Cho, Ji Joong; Yun, Han Dae

2009-03-11

We examined the role of microorganisms in the degradation of the organophosphorus (OP) insecticide chlorpyrifos (CP) during kimchi fermentation. During the fermentation of kimchi, 30 mg L(-1) of CP was added and its stability assayed during fermentation. CP was degraded rapidly until day 3 (83.3%) and degraded completely by day 9. Four CP-degrading lactic acid bacteria (LAB) were isolated from kimchi fermentation in the presence of 200 mg L(-1) CP and were identified as Leuconostoc mesenteroides WCP907, Lactobacillus brevis WCP902, Lactobacillus plantarum WCP931, and Lactobacillus sakei WCP904. CP could be utilized by these four strains as the sole source of carbon and phosphorus. Coumaphos (CM), diazinon (DZ), parathion (PT), and methylparathion (MPT) were also degraded by WCP907, WCP902, WCP931, and WCP904 when provided as sole sources of carbon and phosphorus.

Lactobacillus hammesii sp. nov., isolated from French sourdough.

PubMed

Valcheva, Rosica; Korakli, Maher; Onno, Bernard; Prévost, Hervé; Ivanova, Iskra; Ehrmann, Matthias A; Dousset, Xavier; Gänzle, Michael G; Vogel, Rudi F

2005-03-01

Twenty morphologically different strains were chosen from French wheat sourdough isolates. Cells were Gram-positive, non-spore-forming, non-motile rods. The isolates were identified using amplified-fragment length polymorphism, randomly amplified polymorphic DNA and 16S rRNA gene sequence analysis. All isolates were members of the genus Lactobacillus. They were identified as representing Lactobacillus plantarum, Lactobacillus paralimentarius, Lactobacillus sanfranciscensis, Lactobacillus spicheri and Lactobacillus sakei. However, two isolates (LP38(T) and LP39) could be clearly discriminated from recognized Lactobacillus species on the basis of genotyping methods. 16S rRNA gene sequence similarity and DNA-DNA relatedness data indicate that the two strains belong to a novel Lactobacillus species, for which the name Lactobacillus hammesii is proposed. The type strain is LP38(T) (=DSM 16381(T)=CIP 108387(T)=TMW 1.1236(T)).

Lactobacillus delbrueckii subsp. jakobsenii subsp. nov., isolated from dolo wort, an alcoholic fermented beverage in Burkina Faso.

PubMed

Adimpong, David B; Nielsen, Dennis S; Sørensen, Kim I; Vogensen, Finn K; Sawadogo-Lingani, Hagrétou; Derkx, Patrick M F; Jespersen, Lene

2013-10-01

Lactobacillus delbrueckii is divided into five subspecies based on phenotypic and genotypic differences. A novel isolate, designated ZN7a-9(T), was isolated from malted sorghum wort used for making an alcoholic beverage (dolo) in Burkina Faso. The results of 16S rRNA gene sequencing, DNA-DNA hybridization and peptidoglycan cell-wall structure type analyses indicated that it belongs to the species L. delbrueckii. The genome sequence of isolate ZN7a-9(T) was determined by Illumina-based sequencing. Multilocus sequence typing (MLST) and split-decomposition analyses were performed on seven concatenated housekeeping genes obtained from the genome sequence of strain ZN7a-9(T) together with 41 additional L. delbrueckii strains. The results of the MLST and split-decomposition analyses could not establish the exact subspecies of L. delbrueckii represented by strain ZN7a-9(T) as it clustered with L. delbrueckii strains unassigned to any of the recognized subspecies of L. delbrueckii. Strain ZN7a-9(T) additionally differed from the recognized type strains of the subspecies of L. delbrueckii with respect to its carbohydrate fermentation profile. In conclusion, the cumulative results indicate that strain ZN7a-9(T) represents a novel subspecies of L. delbrueckii closely related to Lactobacillus delbrueckii subsp. lactis and Lactobacillus delbrueckii subsp. delbrueckii for which the name Lactobacillus delbrueckii subsp. jakobsenii subsp. nov. is proposed. The type strain is ZN7a-9(T) = DSM 26046(T) = LMG 27067(T).

Assessing the inactivation of Mycobacterium avium subsp. paratuberculosis during composting of livestock carcasses.

PubMed

Tkachuk, Victoria L; Krause, Denis O; McAllister, Tim A; Buckley, Katherine E; Reuter, Tim; Hendrick, Steve; Ominski, Kim H

2013-05-01

Mycobacterium avium subsp. paratuberculosis causes Johne's disease (JD) in ruminants, with substantial economic impacts on the cattle industry. Johne's disease is known for its long latency period, and difficulties in diagnosis are due to insensitivities of current detection methods. Eradication is challenging as M. avium subsp. paratuberculosis can survive for extended periods within the environment, resulting in new infections in naïve animals (W. Xu et al., J. Environ. Qual. 38:437-450, 2009). This study explored the use of a biosecure, static composting structure to inactivate M. avium subsp. paratuberculosis. Mycobacterium smegmatis was also assessed as a surrogate for M. avium subsp. paratuberculosis. Two structures were constructed to hold three cattle carcasses each. Naturally infected tissues and ground beef inoculated with laboratory-cultured M. avium subsp. paratuberculosis and M. smegmatis were placed in nylon and plastic bags to determine effects of temperature and compost environment on viability over 250 days. After removal, samples were cultured and growth of both organisms was assessed after 12 weeks. After 250 days, M. avium subsp. paratuberculosis was still detectable by PCR, while M. smegmatis was not detected after 67 days of composting. Furthermore, M. avium subsp. paratuberculosis remained viable in both implanted nylon and plastic bags over the composting period. As the compost never reached a homogenous thermophilic (55 to 65°C) state throughout each structure, an in vitro experiment was conducted to examine viability of M. avium subsp. paratuberculosis after exposure to 80°C for 90 days. Naturally infected lymph tissues were mixed with and without compost. After 90 days, M. avium subsp. paratuberculosis remained viable despite exposure to temperatures typically higher than that achieved in compost. In conclusion, it is unlikely composting can be used as a means of inactivating M. avium subsp. paratuberculosis associated with cattle

Assessing the Inactivation of Mycobacterium avium subsp. paratuberculosis during Composting of Livestock Carcasses

PubMed Central

Tkachuk, Victoria L.; Krause, Denis O.; McAllister, Tim A.; Buckley, Katherine E.; Reuter, Tim; Hendrick, Steve

2013-01-01

Mycobacterium avium subsp. paratuberculosis causes Johne's disease (JD) in ruminants, with substantial economic impacts on the cattle industry. Johne's disease is known for its long latency period, and difficulties in diagnosis are due to insensitivities of current detection methods. Eradication is challenging as M. avium subsp. paratuberculosis can survive for extended periods within the environment, resulting in new infections in naïve animals (W. Xu et al., J. Environ. Qual. 38:437-450, 2009). This study explored the use of a biosecure, static composting structure to inactivate M. avium subsp. paratuberculosis. Mycobacterium smegmatis was also assessed as a surrogate for M. avium subsp. paratuberculosis. Two structures were constructed to hold three cattle carcasses each. Naturally infected tissues and ground beef inoculated with laboratory-cultured M. avium subsp. paratuberculosis and M. smegmatis were placed in nylon and plastic bags to determine effects of temperature and compost environment on viability over 250 days. After removal, samples were cultured and growth of both organisms was assessed after 12 weeks. After 250 days, M. avium subsp. paratuberculosis was still detectable by PCR, while M. smegmatis was not detected after 67 days of composting. Furthermore, M. avium subsp. paratuberculosis remained viable in both implanted nylon and plastic bags over the composting period. As the compost never reached a homogenous thermophilic (55 to 65°C) state throughout each structure, an in vitro experiment was conducted to examine viability of M. avium subsp. paratuberculosis after exposure to 80°C for 90 days. Naturally infected lymph tissues were mixed with and without compost. After 90 days, M. avium subsp. paratuberculosis remained viable despite exposure to temperatures typically higher than that achieved in compost. In conclusion, it is unlikely composting can be used as a means of inactivating M. avium subsp. paratuberculosis associated with cattle

Environmental Mycobacterium avium subsp. paratuberculosis hosted by free-living amoebae

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis is responsible for paratuberculosis in animals. This disease, leading to an inflammation of the gastrointestinal tract, has a high impact on animal health and an important economic burden. The environmental life cycle of Mycobacterium avium subsp. paratube...

Relationship between presence of cows with milk positive for Mycobacterium avium subsp. paratuberculosis-specific antibody by enzyme-linked immunosorbent assay and viable M. avium subsp. paratuberculosis in dust in cattle barns.

PubMed

Eisenberg, Susanne W F; Chuchaisangrat, Ruj; Nielen, Mirjam; Koets, Ad P

2013-09-01

Paratuberculosis, or Johne's disease, in cattle is caused by Mycobacterium avium subsp. paratuberculosis, which has recently been suspected to be transmitted through dust. This longitudinal study on eight commercial M. avium subsp. paratuberculosis-positive dairy farms studied the relationship between the number of cows with M. avium subsp. paratuberculosis antibody-positive milk and the presence of viable M. avium subsp. paratuberculosis in settled-dust samples, including their temporal relationship. Milk and dust samples were collected in parallel monthly for 2 years. M. avium subsp. paratuberculosis antibodies in milk were measured by enzyme-linked immunosorbent assay (ELISA) and used as a proxy for M. avium subsp. paratuberculosis shedding. Settled-dust samples were collected by using electrostatic dust collectors (EDCs) at six locations in housing for dairy cattle and young stock. The presence of viable M. avium subsp. paratuberculosis was identified by liquid culture and PCR. The results showed a positive relationship (odds ratio [OR], 1.2) between the number of cows with ELISA-positive milk and the odds of having positive EDCs in the same airspace as the adult dairy cattle. Moreover, the total number of lactating cows also showed an OR slightly above 1. This relationship remained the same for settled-dust samples collected up to 2 months before or after the time of milk sampling. The results suggest that removal of adult cows with milk positive for M. avium subsp. paratuberculosis-specific antibody by ELISA might result in a decrease in the presence of viable M. avium subsp. paratuberculosis in dust and therefore in the environment. However, this decrease is likely delayed by several weeks at least. In addition, the data support the notion that M. avium subsp. paratuberculosis exposure of young stock is reduced by separate housing.

Relationship between Presence of Cows with Milk Positive for Mycobacterium avium subsp. paratuberculosis-Specific Antibody by Enzyme-Linked Immunosorbent Assay and Viable M. avium subsp. paratuberculosis in Dust in Cattle Barns

PubMed Central

Chuchaisangrat, Ruj; Nielen, Mirjam; Koets, Ad P.

2013-01-01

Paratuberculosis, or Johne's disease, in cattle is caused by Mycobacterium avium subsp. paratuberculosis, which has recently been suspected to be transmitted through dust. This longitudinal study on eight commercial M. avium subsp. paratuberculosis-positive dairy farms studied the relationship between the number of cows with M. avium subsp. paratuberculosis antibody-positive milk and the presence of viable M. avium subsp. paratuberculosis in settled-dust samples, including their temporal relationship. Milk and dust samples were collected in parallel monthly for 2 years. M. avium subsp. paratuberculosis antibodies in milk were measured by enzyme-linked immunosorbent assay (ELISA) and used as a proxy for M. avium subsp. paratuberculosis shedding. Settled-dust samples were collected by using electrostatic dust collectors (EDCs) at six locations in housing for dairy cattle and young stock. The presence of viable M. avium subsp. paratuberculosis was identified by liquid culture and PCR. The results showed a positive relationship (odds ratio [OR], 1.2) between the number of cows with ELISA-positive milk and the odds of having positive EDCs in the same airspace as the adult dairy cattle. Moreover, the total number of lactating cows also showed an OR slightly above 1. This relationship remained the same for settled-dust samples collected up to 2 months before or after the time of milk sampling. The results suggest that removal of adult cows with milk positive for M. avium subsp. paratuberculosis-specific antibody by ELISA might result in a decrease in the presence of viable M. avium subsp. paratuberculosis in dust and therefore in the environment. However, this decrease is likely delayed by several weeks at least. In addition, the data support the notion that M. avium subsp. paratuberculosis exposure of young stock is reduced by separate housing. PMID:23793639

Thermal Inactivation of Mycobacterium avium subsp. paratuberculosis in Artificially Contaminated Milk by Direct Steam Injection

PubMed Central

Butot, Sophie; Jagadeesan, Balamurugan; Bakker, Douwe; Donaghy, John

2016-01-01

ABSTRACT The efficiency of direct steam injection (DSI) at 105°C for 3 s to inactivate Mycobacterium avium subsp. paratuberculosis in milk at a pilot-plant scale was investigated. Milk samples were artificially contaminated with M. avium subsp. paratuberculosis and also with cow fecal material naturally infected with M. avium subsp. paratuberculosis. We also tested milk artificially contaminated with Mycobacterium smegmatis as a candidate surrogate to compare thermal inactivation between M. smegmatis and M. avium subsp. paratuberculosis. Following the DSI process, no viable M. avium subsp. paratuberculosis or M. smegmatis was recovered using culture methods for both strains. For pure M. avium subsp. paratuberculosis cultures, a minimum reduction of 5.6 log10 was achieved with DSI, and a minimum reduction of 5.7 log10 was found with M. smegmatis. The minimum log10 reduction for wild-type M. avium subsp. paratuberculosis naturally present in feces was 3.3. In addition, 44 dairy and nondairy powdered infant formula (PIF) ingredients used during the manufacturing process of PIF were tested for an alternate source for M. avium subsp. paratuberculosis and were found to be negative by quantitative PCR (qPCR). In conclusion, the results obtained from this study indicate that a >7-fold-log10 reduction of M. avium subsp. paratuberculosis in milk can be achieved with the applied DSI process. IMPORTANCE M. avium subsp. paratuberculosis is widespread in dairy herds in many countries. M. avium subsp. paratuberculosis is the causative agent of Johne's disease in cattle, and infected animals can directly or indirectly (i.e., fecal contamination) contaminate milk. Despite much research and debate, there is no conclusive evidence that M. avium subsp. paratuberculosis is a zoonotic bacterium, i.e., one that causes disease in humans. The presence of M. avium subsp. paratuberculosis or its DNA has been reported in dairy products, including pasteurized milk, cheese, and infant formula

[Chalcones from Bauhinia glauca subsp. pernervosa].

PubMed

Wu, Zengbao; Wang, Bin; Zhao, Yuying; Yang, Xiuwei; Liang, Hong

2009-07-01

To study the chemical constituents of Bauhinia glauca subsp. pernervosa. The coulis of B. glauca subsp. pernervosa were extracted with 95% EtOH at room temperature. The compounds were isolated and separated by chromatographic techniques, and structures were identified by spectroscopic methods. Seven chalcones were isolated and identified: butein-4-methyl ether (1), isoliquiritigenin (2), butein (3), isoliquiritigenin-2'-methyl ether (4), 2',4'-dihydroxychalcone (5), isoliquiritigenin-4-methyl ether (6), 4-hydroxy-2',4'-dimethoxychalcone (7). Compounds 1, 3, and 7 were isolated from the genus Bauhinia for the first time, the other compounds were obtained from this plant for the first time.

Antioxidant activity profiling by spectrophotometric methods of aqueous methanolic extracts of Helichrysum stoechas subsp. rupestre and Phagnalon saxatile subsp. saxatile.

PubMed

Haddouchi, Farah; Chaouche, Tarik Mohammed; Ksouri, Riadh; Medini, Faten; Sekkal, Fatima Zohra; Benmansour, Abdelhafid

2014-06-01

The aqueous methanolic extracts of two plants from Algeria, Helichrysum stoechas subsp. rupestre and Phagnalon saxatile subsp. saxatile, were investigated for their antioxidant activity. Total phenolics, flavonoids, and tannins were determined by spectrophotometric techniques. In vitro antioxidant and radical scavenging profiling was determined by spectrophotometric methods, through: Total antioxidant capacity, and radical scavenging effects by the DPPH and ABTS methods, reducing and chelating power, and blanching inhibition of the β-carotene. All of the extracts showed interesting antioxidant and radical scavenging activity. The highest contents in phenolics, tannins, and the highest total antioxidant capacity as gallic acid equivalents of 97.5 ± 0.33 mg GAE/g DW was obtained for the flowers of H. stoechas subsp. rupestre extract in the phosphomolybdenum assay. An extract of the leafy stems of P. saxatile subsp. saxatile revealed the highest content of flavonoids, and the highest antioxidant activity by the radical scavenging and β-carotene assays when compared with standards. The best activity was by the scavenging radical DPPH with an IC50 value of 5.65 ± 0.10 μg·mL(-1). The studied medicinal plants could provide scientific evidence for some traditional uses in the treatment of diseases related to the production of reactive oxygen species (ROS) and oxidative stress. Copyright © 2014 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

Bioluminescence imaging of Clavibacter michiganensis subsp. michiganensis infection of tomato seeds and plants.

PubMed

Xu, Xiulan; Miller, Sally A; Baysal-Gurel, Fulya; Gartemann, Karl-Heinz; Eichenlaub, Rudolf; Rajashekara, Gireesh

2010-06-01

Clavibacter michiganensis subsp. michiganensis is a Gram-positive bacterium that causes wilting and cankers, leading to severe economic losses in commercial tomato production worldwide. The disease is transmitted from infected seeds to seedlings and mechanically from plant to plant during seedling production, grafting, pruning, and harvesting. Because of the lack of tools for genetic manipulation, very little is known regarding the mechanisms of seed and seedling infection and movement of C. michiganensis subsp. michiganensis in grafted plants, two focal points for application of bacterial canker control measures in tomato. To facilitate studies on the C. michiganensis subsp. michiganensis movement in tomato seed and grafted plants, we isolated a bioluminescent C. michiganensis subsp. michiganensis strain using the modified Tn1409 containing a promoterless lux reporter. A total of 19 bioluminescent C. michiganensis subsp. michiganensis mutants were obtained. All mutants tested induced a hypersensitive response in Mirabilis jalapa and caused wilting of tomato plants. Real-time colonization studies of germinating seeds using a virulent, stable, constitutively bioluminescent strain, BL-Cmm17, showed that C. michiganensis subsp. michiganensis aggregated on hypocotyls and cotyledons at an early stage of germination. In grafted seedlings in which either the rootstock or scion was exposed to BL-Cmm17 via a contaminated grafting knife, bacteria were translocated in both directions from the graft union at higher inoculum doses. These results emphasize the use of bioluminescent C. michiganensis subsp. michiganensis to help better elucidate the C. michiganensis subsp. michiganensis-tomato plant interactions. Further, we demonstrated the broader applicability of this tool by successful transformation of C. michiganensis subsp. nebraskensis with Tn1409::lux. Thus, our approach would be highly useful to understand the pathogenesis of diseases caused by other subspecies of the

Specific 16S ribosomal RNA targeted oligonucleotide probe against Clavibacter michiganensis subsp. sepedonicus.

PubMed

Mirza, M S; Rademaker, J L; Janse, J D; Akkermans, A D

1993-11-01

In this article we report on the polymerase chain reaction amplification of a partial 16S rRNA gene from the plant pathogenic bacterium Clavibacter michiganensis subsp. sepedonicus. A partial sequence (about 400 base pairs) of the gene was determined that covered two variable regions important for oligonucleotide probe development. A specific 24mer oligonucleotide probe targeted against the V6 region of 16S rRNA was designed. Specificity of the probe was determined using dot blot hybridization. Under stringent conditions (60 degrees C), the probe hybridized with all 16 Cl. michiganensis subsp. sepedonicus strains tested. Hybridization did not occur with 32 plant pathogenic and saprophytic bacteria used as controls under the same conditions. Under less stringent conditions (55 degrees C) the related Clavibacter michiganensis subsp. insidiosus, Clavibacter michiganensis subsp. nebraskensis, and Clavibacter michiganensis subsp. tesselarius also showed hybridization. At even lower stringency (40 degrees C), all Cl. michiganensis subspecies tested including Clavibacter michiganensis subsp. michiganensis showed hybridization signal, suggesting that under these conditions the probe may be used as a species-specific probe for Cl. michiganensis.

RAPD-PCR characterization of lactobacilli isolated from artisanal meat plants and traditional fermented sausages of Veneto region (Italy).

PubMed

Andrighetto, C; Zampese, L; Lombardi, A

2001-07-01

The study was carried out to evaluate the use of randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) as a method for the identification of lactobacilli isolated from meat products. RAPD-PCR with primers M13 and D8635 was applied to the identification and intraspecific differentiation of 53 lactobacilli isolates originating from traditional fermented sausages and artisanal meat plants of the Veneto region (Italy). Most of the isolates were assigned to the species Lactobacillus sakei and Lact. curvatus; differentiation of groups of strains within the species was also possible. RAPD-PCR could be applied to the identification of lactobacilli species most commonly found in meat products. The method, which is easy and rapid to perform, could be useful for the study of the lactobacilli populations present in fermented sausages, and could help in the selection of candidate strains to use as starter cultures in meat fermentation.

The in vitro effect of six antimicrobials against Mycoplasma putrefaciens, Mycoplasma mycoides subsp. mycoides LC and Mycoplasma capricolum subsp. capricolum isolated from sheep and goats in Jordan.

PubMed

Al-Momani, W; Nicholas, R A J; Janakat, S; Abu-Basha, E; Ayling, R D

2006-01-01

Respiratory disease in sheep and goats is a major problem in Jordan and is often associated with Mycoplasma species. Without effective vaccines, control is mainly by chemotherapy, but the uncontrolled use of antimicrobials has led to concerns about the potential development of antimicrobial resistance. The in vitro effect of chloramphenicol, florfenicol, enrofloxacin, tylosin, erythromycin and oxytetracycline was determined against 32 isolates of Mycoplasma species-M. mycoides subsp. mycoides LC (6), M. capricolum subsp. capricolum (8) and M. putrefaciens (18), all isolated from either nasal swabs or milk, from sheep and goats in different regions of Jordan. The antimicrobial susceptibility showed some Mycoplasma species-specific differences, with M. capricolum subsp. capricolum being more susceptible to tylosin and erythromycin. Chloramphenicol and florfenicol were the least effective for all three Mycoplasma species. No trends or significant differences in antimicrobial susceptibilities were observed between sheep and goat isolates, between milk or nasal swab isolates, or between isolates from different regions of Jordan. Some isolates of M. capricolum subsp. capricolum and M. putrefaciens showed higher MIC levels with oxytetracycline, as did two isolates of M. mycoides subsp. mycoides LC with tylosin, possibly indicating signs of development of antimicrobial resistance.

Bioluminescence Imaging of Clavibacter michiganensis subsp. michiganensis Infection of Tomato Seeds and Plants ▿

PubMed Central

Xu, Xiulan; Miller, Sally A.; Baysal-Gurel, Fulya; Gartemann, Karl-Heinz; Eichenlaub, Rudolf; Rajashekara, Gireesh

2010-01-01

Clavibacter michiganensis subsp. michiganensis is a Gram-positive bacterium that causes wilting and cankers, leading to severe economic losses in commercial tomato production worldwide. The disease is transmitted from infected seeds to seedlings and mechanically from plant to plant during seedling production, grafting, pruning, and harvesting. Because of the lack of tools for genetic manipulation, very little is known regarding the mechanisms of seed and seedling infection and movement of C. michiganensis subsp. michiganensis in grafted plants, two focal points for application of bacterial canker control measures in tomato. To facilitate studies on the C. michiganensis subsp. michiganensis movement in tomato seed and grafted plants, we isolated a bioluminescent C. michiganensis subsp. michiganensis strain using the modified Tn1409 containing a promoterless lux reporter. A total of 19 bioluminescent C. michiganensis subsp. michiganensis mutants were obtained. All mutants tested induced a hypersensitive response in Mirabilis jalapa and caused wilting of tomato plants. Real-time colonization studies of germinating seeds using a virulent, stable, constitutively bioluminescent strain, BL-Cmm17, showed that C. michiganensis subsp. michiganensis aggregated on hypocotyls and cotyledons at an early stage of germination. In grafted seedlings in which either the rootstock or scion was exposed to BL-Cmm17 via a contaminated grafting knife, bacteria were translocated in both directions from the graft union at higher inoculum doses. These results emphasize the use of bioluminescent C. michiganensis subsp. michiganensis to help better elucidate the C. michiganensis subsp. michiganensis-tomato plant interactions. Further, we demonstrated the broader applicability of this tool by successful transformation of C. michiganensis subsp. nebraskensis with Tn1409::lux. Thus, our approach would be highly useful to understand the pathogenesis of diseases caused by other subspecies of the

A Proteomic Study of Clavibacter Michiganensis Subsp. Michiganensis Culture Supernatants

PubMed Central

Hiery, Eva; Poetsch, Ansgar; Moosbauer, Tanja; Amin, Bushra; Hofmann, Jörg; Burkovski, Andreas

2015-01-01

Clavibacter michiganensis, subsp. michiganensis is a Gram-positive plant pathogen infecting tomato (Solanum lycopersicum). Despite a considerable economic importance due to significant losses of infected plants and fruits, knowledge about virulence factors of C. michiganensis subsp. michiganensis and host-pathogen interactions on a molecular level are rather limited. In the study presented here, the proteome of culture supernatants from C. michiganensis subsp. michiganensis NCPPB382 was analyzed. In total, 1872 proteins were identified in M9 and 1766 proteins in xylem mimicking medium. Filtration of supernatants before protein precipitation reduced these to 1276 proteins in M9 and 976 proteins in the xylem mimicking medium culture filtrate. The results obtained indicate that C. michiganensis subsp. michiganensis reacts to a sucrose- and glucose-depleted medium similar to the xylem sap by utilizing amino acids and host cell polymers as well as their degradation products, mainly peptides, amino acids and various C5 and C6 sugars. Interestingly, the bacterium expresses the previously described virulence factors Pat-1 and CelA not exclusively after host cell contact in planta but already in M9 minimal and xylem mimicking medium. PMID:28248277

Quantification of the Sensitivity of Mycobacterium avium subsp paratuberculosis and Salmonella enterica subsp enterica to Low pH and High Organic Acids using Propidium Monoazide and Quantitative PCR

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp paratuberculosis (Map) and Salmonella enterica subsp enterica (S. enterica) are two pathogens that are a concern to food and animal safety due to their ability to withstand harsh conditions encountered in the natural environment and within the host during pathogenesis. Acid...

Disparate host immunity to Mycobacterium avium subsp. paratuberculosis antigens in calves inoculated with M. avium subsp. paratuberculosis, M. avium subsp. avium, M. kansasii and M. bovis

USDA-ARS?s Scientific Manuscript database

Cross-reactivity of mycobacterial antigens in immune-based diagnostic assays has been a major concern and criticism of current tests for the detection of paratuberculosis. In the present study, host immune responses to antigen preparations of Mycobacterium avium subsp. paratuberculosis (MAP), consis...

Nested PCR for ultrasensitive detection of the potato ring rot bacterium, Clavibacter michiganensis subsp. sepedonicus.

PubMed

Lee, I M; Bartoszyk, I M; Gundersen, D E; Mogen, B; Davis, R E

1997-07-01

Oligonucleotide primers derived from sequences of the 16S rRNA gene (CMR16F1, CMR16R1, CMR16F2, and CMR16R2) and insertion element IS1121 of Clavibacter michiganensis subsp. sepedonicus (CMSIF1, CMSIR1, CMSIF2, and CMISR2) were used in nested PCR to detect the potato ring rot bacterium C. michiganensis subsp. sepedonicus. Nested PCR with primer pair CMSIF1-CMSIR1 followed by primer pair CMSIF2-CMSIR2 specifically detected C. michiganensis subsp. sepedonicus, while nested PCR with CMR16F1-CMR16R1 followed by CMR16F2-CMR16R2 detected C. michiganensis subsp. sepedonicus and the other C. michiganensis subspecies. In the latter case, C. michiganensis subsp. sepedonicus can be differentiated from the other subspecies by restriction fragment length polymorphism (RFLP) analyses of the nested PCR products (16S rDNA sequences). The nested PCR assays developed in this work allow ultrasensitive detection of very low titers of C. michiganensis subsp. sepedonicus which may be present in symptomiess potato plants or tubers and which cannot be readily detected by direct PCR (single PCR amplification). RFLP analysis of PCR products provides for an unambiguous confirmation of the identify of C. michiganensis subsp. sepedonicus.

In Vitro Antiproliferative Activity of Extracts of Carlina acaulis subsp. caulescens and Carlina acanthifolia subsp. utzka

PubMed Central

Strzemski, Maciej; Wojnicki, Kamil; Sowa, Ireneusz; Wojas-Krawczyk, Kamila; Krawczyk, Paweł; Kocjan, Ryszard; Such, Justyna; Latalski, Michał; Wnorowski, Artur; Wójciak-Kosior, Magdalena

2017-01-01

Various species of the Carlina genus have been used in traditional medicine in many countries to treat numerous skin disorders, including cancer. The objective of this work was to assess the anticancer properties of root and leaf extracts from Carlina acaulis subsp. caulescens and C. acanthifolia subsp. utzka. Anti-tumor properties of the extracts were explored using a tetrazolium-based cell viability assay and flow cytometric apoptosis analysis, followed by immunodetection of phosphoactive ERK1/2 in UACC-903, C32, and UACC-647 human melanoma cell lines. Normal human fibroblasts were used as a control. Leaf extracts inhibited the viability of all tested melanoma cell lines in a dose-dependent fashion while the fibroblasts were less sensitive to such extract. The root extracts inhibited the proliferation of UACC-903 and UACC-647 cells only at the highest doses (300 μg/mL). However, the C32 and fibroblast cells exhibited an increase in the cellular proliferation rate and no caspase activity was observed in response to the root extracts (100 μg/mL). An increase in caspase activity was observed in melanoma cells treated with the leaf extracts of both Carlina species. Leaf extracts from C. acaulis subsp. caulescens (100 μg/mL) inhibited proliferatory ERK1/2 in UACC-903 and C32 cells, as demonstrated by the decrease in ERK1/2 phosphorylation. No reduction in phospho-ERK1/2 was observed in the tested cell lines treated with the root extracts, apart from UACC-647 after incubation with the C. acanthifolia subsp. utzka root extract (100 μg/mL). There was no change in ERK1/2 phosphorylation in the fibroblasts. The extracts from the leaves and roots were analyzed by HPLC and the analysis showed the presence of triterpenes and phenolic acids as the main extract components. The research demonstrated that the extracts from the leaves of the plants were cytotoxic against the human melanoma line and induced apoptosis of the cells. The triterpene fraction present in the tested

Lactobacillus plantarum subsp. argentoratensis subsp. nov., isolated from vegetable matrices.

PubMed

Bringel, Françoise; Castioni, Anna; Olukoya, Daniel K; Felis, Giovanna E; Torriani, Sandra; Dellaglio, Franco

2005-07-01

Fourteen strains isolated from vegetable sources and identified as belonging to Lactobacillus plantarum presented an atypical pattern of amplification with a species-specific multiplex-PCR assay. Phylogenetic analysis of two protein-encoding genes, recA (encoding the recombinase A protein) and cpn60 (encoding the GroEL chaperonin), as well as phenotypic and genomic traits revealed a homogeneous group of very closely related strains for which subspecies status is proposed, with the name Lactobacillus plantarum subsp. argentoratensis. The type strain is DKO 22(T) (=CIP 108320(T)=DSM 16365(T)).

Clavibacter michiganensis subsp. capsici subsp. nov., causing bacterial canker disease in pepper.

PubMed

Oh, Eom-Ji; Bae, Chungyun; Lee, Han-Beoyl; Hwang, In Sun; Lee, Hyok-In; Yea, Mi Chi; Yim, Kyu-Ock; Lee, Seungdon; Heu, Sunggi; Cha, Jae-Soon; Oh, Chang-Sik

2016-10-01

Clavibacter michiganensis is a Gram-stain-positive bacterium with eight subspecies. One of these subspecies is C. michiganensis subsp. michiganensis, which causes bacterial canker disease in tomato. Bacterial strains showing very similar canker disease symptoms to those of a strain originally classified as C. michiganensis have been isolated from pepper. In this paper, we reclassified strains isolated from pepper. On the basis of phylogenetic analysis with 16S rRNA gene sequences, the strains isolated from pepper were grouped in a separate clade from other subspecies of C. michiganensis. Biochemical, physiological and genetic characteristics of strain PF008T, which is the representative strain of the isolates from pepper, were examined in this study. Based on multi-locus sequence typing and other biochemical and physiological features including colony color, utilization of carbon sources and enzyme activities, strain PF008T was categorically differentiated from eight subspecies of C. michiganensis. Moreover, genome analysis showed that the DNA G+C content of strain PF008T is 73.2 %. These results indicate that PF008T is distinct from other known subspecies of C. michiganensis. Therefore, we propose a novel subspecies, C. michiganensis subsp. capsici, causing bacterial canker disease in pepper, with a type strain of PF008T (=KACC 18448T=LMG 29047T).

[Identification and phylogenetic analysis of one strain of Lactobacillus delbrueckii subsp. bulgaricus separated from yoghourt].

PubMed

Wang, Chuan; Zhang, Chaowu; Pei, Xiaofang; Liu, Hengchuan

2007-11-01

For being further applied and studied, one strain of Lactobacillus delbrueckii subsp. bulgaricus (wch9901) separated from yoghourt which had been identified by phenotype characteristic analysis was identified by 16S rDNA and phylogenetic analyzed. The 16S rDNA of wch9901 was amplified with the genomic DNA of wch9901 as template, and the conservative sequences of the 16S rDNA as primers. Inserted 16S rDNA amplified into clonal vector pGEM-T under the function of T4 DNA ligase to construct recombined plasmid pGEM-wch9901 16S rDNA. The recombined plasmid was identified by restriction enzyme digestion, and the eligible plasmid was presented to sequencing company for DNA sequencing. Nucleic acid sequence was blast in GenBank and phylogenetic tree was constructed using neighbor-joining method of distance methods by Mega3.1 soft. Results of blastn showed that the homology of 16S rDNA of wch9901 with the 16S rDNA of Lactobacillus delbrueckii subsp. bulgaricus strains was higher than 96%. On the phylogenetic tree, wch9901 formed a separate branch and located between Lactobacillus delbrueckii subsp. bulgaricus LGM2 evolution branch and another evolution branch which was composed of Lactobacillus delbrueckii subsp. bulgaricus DL2 evolution cluster and Lactobacillus delbrueckii subsp. bulgaricus JSQ evolution cluster. The distance between wch9901 evolution branch and Lactobacillus delbrueckii subsp. bulgaricus LGM2 evolution branch was the closest. wch9901 belonged to Lactobacillus delbrueckii subsp. bulgaricus. wch9901 showed the closest evolution relationship to Lactobacillus delbrueckii subsp. bulgaricus LGM2.

High pressure inactivation of relevant target microorganisms in poultry meat products and the evaluation of pressure-induced protein denaturation of marinated poultry under different high pressure treatments

NASA Astrophysics Data System (ADS)

Schmidgall, Johanna; Hertel, Christian; Bindrich, Ute; Heinz, Volker; Toepfl, Stefan

2011-03-01

In this study, the possibility of extending shelf life of marinated poultry meat products by high pressure processing was evaluated. Relevant spoilage and pathogenic strains were selected and used as target microorganisms (MOs) for challenge experiments. Meat and brine were inoculated with MOs and treated at 450 MPa, 4 °C for 3 min. The results of inactivation show a decreasing pressure tolerance in the series Lactobacillus > Arcobacter > Carnobacterium > Bacillus cereus > Brochothrix thermosphacta > Listeria monocytogenes. Leuconostoc gelidum exhibited the highest pressure tolerance in meat. A protective effect of poultry meat was found for L. sakei and L. gelidum. In parallel, the influence of different marinade formulations (pH, carbonates, citrates) on protein structure changes during a pressure treatment was investigated. Addition of sodium carbonate shows a protection against denaturation of myofibrillar proteins and provides a maximum water-holding capacity. Caustic marinades allowed a higher retention of product characteristics than low-pH marinades.

Degradation of PCBs in dry fermented sausages during drying/ripening.

PubMed

Lušnic Polak, M; Zlatić, E; Demšar, L; Žlender, B; Polak, T

2016-12-15

The effects of several commercial meat starter cultures on degradation of polychlorinated biphenyls (PCBs) in dry fermented sausages over 28days of drying/ripening were investigated. The sausage batter was prepared according to a classic recipe and spiked with a standard solution of a PCB congener mixture. With addition of different commercial meat starter cultures, five experimental groups were prepared: no further addition; and separate addition of each of four starter cultures: Texel DCM-1, Texel LM-30, Biostar Sprint, and SM-181. Samples were taken at the beginning of fermentation (zero time), and after 4, 7, 14, 21 and 28days. PCB residues were extracted with hexane. The PCB contents were determined using gas chromatography-mass spectrometry. The PCB levels were reduced in all of the experimental groups tested, where addition of starter culture Biostar Sprint (Lactobacillus sakei, Staphylococcus carnosus, Staphylococcus xylosus) showed the highest PCB degradation rates. Copyright © 2016 Elsevier Ltd. All rights reserved.

Tracing microbiota changes in yamahai-moto, the traditional Japanese sake starter.

PubMed

Koyanagi, Takashi; Nakagawa, Akira; Kiyohara, Masashi; Matsui, Hiroshi; Tsuji, Atsushi; Barla, Florin; Take, Harumi; Katsuyama, Yoko; Tokuda, Koji; Nakamura, Shizuo; Minami, Hiromichi; Enomoto, Toshiki; Katayama, Takane; Kumagai, Hidehiko

2016-01-01

Sake is made from steamed rice, malted rice, and water. Sake production begins with the preparation of a small-scale starter (moto); the quality of moto significantly influences the flavor and richness of sake. In the traditional starter, yamahai-moto, the growth of naturally occurring lactic acid bacteria represses the putrefactive micro-organisms, whereas in the modern starter, sokujo-moto, this is achieved by adding lactic acid. In this study, the successive change in bacterial flora of yamahai-moto was analyzed by pyrosequencing 16S ribosomal RNA genes. Lactobacillus was dominant throughout the process (93-98%). Nitrate-reducing bacteria that have been generally assumed to be the first colonizers of yamahai-moto were scarcely found in the early stage, but Lactobacillus acidipiscis dominated. Lactobacillus sakei drastically increased in the middle stage. This is the first report, though one case study, to show how the early stage microbiota in Japanese yamahai-moto is varyingly controlled without nitrate-reducing bacteria using next-generation sequencing.

Characterisation of lab in typical Salento Pecorino cheese.

PubMed

Cappello, M S; Laddomada, B; Poltronieri, P; Zacheo, G

2001-01-01

Twenty-nine strains of Lactic Acid Bacteria isolated from the typical Pecorino cheese of the Salento area of Italy, were identified and grouped according to their genetic similarity. A preliminary characterisation of the strains was conducted by means of morphological and biochemical analysis, but molecular approaches were necessary for the clear identification of the species. For the species detection, the amplification and sequencing of the 16S rDNA gene was employed In addition, restriction analysis of amplified rDNA (ARDRA) and PCR and AFLP fingerprinting enabled inter- and intra-specific variation to be estimated UPGMA cluster analysis was used to divide the strains into distinct clusters which corresponded with the species delineation obtained by molecular identification. The data obtained show that the community of lactobacilli responsible for the fermentation and aging of Pecorino cheese is composed of a limited number of species. The main identified strains were Lactobacillus brevis, L. plantarum, L. casei, L. sakei, L. pentosus, L. farciminis and Leuconostoc mesenteroides.

Role of jeotgal, a Korean traditional fermented fish sauce, in microbial dynamics and metabolite profiles during kimchi fermentation.

PubMed

Jung, Min Young; Kim, Tae-Woon; Lee, Changsu; Kim, Joon Yong; Song, Hye Seon; Kim, Yeon Bee; Ahn, Seung Woo; Kim, Ju Seok; Roh, Seong Woon; Lee, Se Hee

2018-11-01

We investigated the effects of jeotgal (fermented fish sauce) on kimchi fermentation, with or without saeu-jeot and myeolchi-jeot. Bacterial community analysis showed that Leuconostoc, Weissella, Lactobacillus, and Tetragenococcus were the dominant genera; however, their succession depended on the presence of jeotgal. Leuconostoc gasicomitatum was the dominant species in kimchi without jeotgal, whereas Weissella koreensis and Lactobacillus sakei were the dominant species in kimchi with myeolchi-jeot and saeu-jeot, respectively. Metabolite analysis, using 1 H NMR, showed that the amounts of amino acids and gamma-aminobutyric acid (GABA) were higher in kimchi with jeotgal. Increases in acetate, lactate, and mannitol contents depended on fructose consumption and were more rapid in kimchi with jeotgal. Moreover, the consumption of various amino acids affected the increase in kimchi LAB. Thus, the role of jeotgal in kimchi fermentation was related to enhancement of taste, the amino acid source, and the increases in levels of functional metabolites. Copyright © 2018. Published by Elsevier Ltd.

Isolation of Campylobacter fetus subsp. jejuni from migratory waterfowl.

PubMed Central

Luechtefeld, N A; Blaser, M J; Reller, L B; Wang, W L

1980-01-01

Since the sources from which humans acquire Campylobacter enteritis are only partially known, we studied the frequency of carriage of Campylobacter fetus subsp. jejuni in migratory waterfowl. Cecal contents of various species of wild ducks were cultured on selective media that contained antibiotics to inhibit normal flora. Thirty-five percent of the 445 ducks cultured harbored C. fetus subsp. jejuni. Migratory waterfowl are yet another reservoir for this enteric pathogen and may be of public health importance for humans in the contamination of water or when used as food. PMID:7217334

Tomato fruit and seed colonization by Clavibacter michiganensis subsp. michiganensis through external and internal routes.

PubMed

Tancos, Matthew A; Chalupowicz, Laura; Barash, Isaac; Manulis-Sasson, Shulamit; Smart, Christine D

2013-11-01

The Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis, causal agent of bacterial wilt and canker of tomato, is an economically devastating pathogen that inflicts considerable damage throughout all major tomato-producing regions. Annual outbreaks continue to occur in New York, where C. michiganensis subsp. michiganensis spreads via infected transplants, trellising stakes, tools, and/or soil. Globally, new outbreaks can be accompanied by the introduction of contaminated seed stock; however, the route of seed infection, especially the role of fruit lesions, remains undefined. In order to investigate the modes of seed infection, New York C. michiganensis subsp. michiganensis field strains were stably transformed with a gene encoding enhanced green fluorescent protein (eGFP). A constitutively eGFP-expressing virulent C. michiganensis subsp. michiganensis isolate, GCMM-22, was used to demonstrate that C. michiganensis subsp. michiganensis could not only access seeds systemically through the xylem but also externally through tomato fruit lesions, which harbored high intra- and intercellular populations. Active movement and expansion of bacteria into the fruit mesocarp and nearby xylem vessels followed, once the fruits began to ripen. These results highlight the ability of C. michiganensis subsp. michiganensis to invade tomato fruits and seeds through multiple entry routes.

Loop-mediated amplification of the Clavibacter michiganensis subsp. michiganensis micA gene is highly specific.

PubMed

Yasuhara-Bell, Jarred; Kubota, Ryo; Jenkins, Daniel M; Alvarez, Anne M

2013-12-01

Loop-mediated amplification (LAMP) was used to specifically identify Clavibacter michiganensis subsp. michiganensis, causal agent of bacterial canker of tomato. LAMP primers were developed to detect micA, a chromosomally stable gene that encodes a type II lantibiotic, michiganin A, which inhibits growth of other C. michiganensis subspecies. In all, 409 bacterial strains (351 C. michiganensis subsp. michiganensis and 58 non-C. michiganensis subsp. michiganensis) from a worldwide collection were tested with LAMP to determine its specificity. LAMP results were compared with genetic profiles established using polymerase chain reaction (PCR) amplification of seven genes (dnaA, ppaJ, pat-1, chpC, tomA, ppaA, and ppaC). C. michiganensis subsp. michiganensis strains produced eight distinct profiles. The LAMP reaction identified all C. michiganensis subsp. michiganensis strains and discriminated them from other C. michiganensis subspecies and non-Clavibacter bacteria. LAMP has advantages over immunodiagnostic and other molecular detection methods because of its specificity and isothermal nature, which allows for easy field application. The LAMP reaction is also not affected by as many inhibitors as PCR. This diagnostic tool has potential to provide an easy, one-step test for rapid identification of C. michiganensis subsp. michiganensis.

Genetic variation in Mediterranean Helichrysum italicum (Asteraceae; Gnaphalieae): do disjunct populations of subsp. microphyllum have a common origin?

PubMed

Galbany-Casals, M; Blanco-Moreno, J M; Garcia-Jacas, N; Breitwieser, I; Smissen, R D

2011-07-01

The yellow-flowered everlasting daisy Helichrysum italicum (Asteraceae, Gnaphalieae) is widely distributed in the Mediterranean basin, where it grows in continuous and widespread populations in diverse open habitats. Helichrysum italicum subsp. microphyllum has a disjunct distribution in the Balearic Islands (Majorca and Dragonera), Corsica, Sardinia, Crete and Cyprus. Numerous morphological intermediates between subsp. italicum and subsp. microphyllum are known from Corsica, where the two subspecies co-occur. The aims of the study were to investigate if subsp. microphyllum has a common origin, constituting an independent gene pool from subsp. italicum, or if the morphological differences between subsp. microphyllum and subsp. italicum have arisen independently in different locations from a common wider gene pool. Our analyses of AFLP, cpDNA sequences and morphological characters show that there is geographic structure to the genetic variation within H. italicum, with eastern and western Mediterranean groups, which do not correspond with the division into subsp. microphyllum and subsp. italicum as currently circumscribed. Local selection on quantitative trait loci provides sufficient explanation for the morphological divergence observed and is consistent with genetic data. Within the western Mediterranean group of the species we found considerable polymorphism in chloroplast DNA sequences among and within some populations. Comparison with chloroplast DNA sequences from other Helichrysum species showed that some chloroplast haplotypes are shared across species. © 2010 German Botanical Society and The Royal Botanical Society of the Netherlands.

Pork Meat as a Potential Source of Salmonella enterica subsp. arizonae Infection in Humans

PubMed Central

Kritas, Spyridon; Govaris, Alexander; Burriel, Angeliki R.

2014-01-01

Salmonella enterica subsp. arizonae was isolated from 13 of 123 slaughtered pigs in central Greece. The samples cultured were feces, ileum tissue, mesenteric lymph nodes, and gallbladder swabs. A total of 74 isolates from 492 samples were identified as Salmonella spp. by use of standard laboratory culture media and two commercial micromethods and by use of a polyvalent slide agglutination test for the detection of O and H antigens. Among them were 19 (25.68%) suspected to be S. enterica subsp. arizonae according to analysis with standard laboratory culture media. Of those, 14 were identified as S. enterica subsp. arizonae by the API 20E (bioMérieux, France) and the Microgen GnA+B-ID (Microgen Bioproducts, Ltd., United Kingdom) identification systems. All the isolates were tested for resistance to 23 antimicrobials. Strains identified as S. enterica subsp. arizonae were resistant to 17 (70.8%) antibiotics. The highest proportions of resistance were observed for sulfamethoxazole-trimethoprim (71.4%), tetracycline (71.4%), ampicillin (64.3%), and amoxicillin (57.1%). Two isolates were resistant to aztreonam (7.1%) and tigecycline (7.1%), used only for the treatment of humans. Thus, pork meat may play a role in the transmission of antibiotic-resistant S. enterica subsp. arizonae to human consumers. This is the first report of S. enterica subsp. arizonae isolation from pigs. PMID:24335956

Pork meat as a potential source of Salmonella enterica subsp. arizonae infection in humans.

PubMed

Evangelopoulou, Grammato; Kritas, Spyridon; Govaris, Alexander; Burriel, Angeliki R

2014-03-01

Salmonella enterica subsp. arizonae was isolated from 13 of 123 slaughtered pigs in central Greece. The samples cultured were feces, ileum tissue, mesenteric lymph nodes, and gallbladder swabs. A total of 74 isolates from 492 samples were identified as Salmonella spp. by use of standard laboratory culture media and two commercial micromethods and by use of a polyvalent slide agglutination test for the detection of O and H antigens. Among them were 19 (25.68%) suspected to be S. enterica subsp. arizonae according to analysis with standard laboratory culture media. Of those, 14 were identified as S. enterica subsp. arizonae by the API 20E (bioMérieux, France) and the Microgen GnA+B-ID (Microgen Bioproducts, Ltd., United Kingdom) identification systems. All the isolates were tested for resistance to 23 antimicrobials. Strains identified as S. enterica subsp. arizonae were resistant to 17 (70.8%) antibiotics. The highest proportions of resistance were observed for sulfamethoxazole-trimethoprim (71.4%), tetracycline (71.4%), ampicillin (64.3%), and amoxicillin (57.1%). Two isolates were resistant to aztreonam (7.1%) and tigecycline (7.1%), used only for the treatment of humans. Thus, pork meat may play a role in the transmission of antibiotic-resistant S. enterica subsp. arizonae to human consumers. This is the first report of S. enterica subsp. arizonae isolation from pigs.

Complete genome sequence of Mycobacterium avium subsp. paratuberculosis, isolated from human breast milk

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp paratuberculosis is the etiologic agent of Johne’s disease. We report the draft genome sequences of six M. avium subsp paratuberculosis isolates obtained from diverse hosts including bison, cattle and sheep. These sequences will deepen our understanding of host association ...

Improvement of DNA transfer frequency and transposon mutagenesis of Erwinia carotovora subsp. betavasculorum.

PubMed Central

Rella, M; Axelrood, P E; Weinhold, A R; Schroth, M N

1989-01-01

The production of antibiotics and their role in microbial competition under natural conditions can be readily studied by the use of transposon mutants. Several antibiotic-producing strains of Erwinia carotovora subsp. betavasculorum were unable to accept foreign DNA. A plasmid delivery system was developed, using ethyl methanesulfonate mutagenesis, which entailed isolating E. carotovora subsp. betavasculorum mutants able to accept foreign DNA and transfer it to other strains. This enabled transposon mutagenesis of a wild-type antibiotic-producing strain of E. carotovora subsp. betavasculorum. Twelve antibiotic-negative mutants were isolated, and one of these showed a reduction in antibiotic production in vitro. Many of these mutants also showed a reduction in their ability to macerate potato tissue. The mutants were classified into four genetic groups on the basis of their genetic and phenotypic characteristics, indicating that several genes are involved in antibiotic biosynthesis by E. carotovora subsp. betavasculorum. PMID:2543291

Seed-associated subspecies of the genus Clavibacter are clearly distinguishable from Clavibacter michiganensis subsp. michiganensis.

PubMed

Yasuhara-Bell, Jarred; Alvarez, Anne M

2015-03-01

The genus Clavibacter contains one recognized species, Clavibacter michiganensis. Clavibacter michiganensis is subdivided into subspecies based on host specificity and bacteriological characteristics, with Clavibacter michiganensis subsp. michiganensis causing bacterial canker of tomato. Clavibacter michiganensis subsp. michiganensis is often spread through contaminated seed leading to outbreaks of bacterial canker in tomato production areas worldwide. The frequent occurrence of non-pathogenic Clavibacter michiganensis subsp. michiganensis-like bacteria (CMB) is a concern for seed producers because Clavibacter michiganensis subsp. michiganensis is a quarantine organism and detection of a non-pathogenic variant may result in destruction of an otherwise healthy seed lot. A thorough biological and genetic characterization of these seed-associated CMB strains was performed using standard biochemical tests, cell wall analyses, metabolic profiling using Biolog, and single-gene and multilocus sequence analyses. Combined, these tests revealed two distinct populations of seed-associated members of the genus Clavibacter that differed from each other, as well as from all other described subspecies of Clavibacter michiganensis. DNA-DNA hybridization values are 70 % or higher, justifying placement into the single recognized species, C. michiganensis, but other analyses justify separate subspecies designations. Additionally, strains belonging to the genus Clavibacter isolated from pepper also represent a distinct population and warrant separate subspecies designation. On the basis of these data we propose subspecies designations for separate non-pathogenic subpopulations of Clavibacter michiganensis: Clavibacter michiganensis subsp. californiensis subsp. nov. and Clavibacter michiganensis subsp. chilensis subsp. nov. for seed-associated strains represented by C55(T) ( = ATCC BAA-2691(T) = CFBP 8216(T)) and ZUM3936(T) ( = ATCC BAA-2690(T) = CFBP 8217(T

Effects of six substances on the growth and freeze-drying of Lactobacillus delbrueckii subsp. bulgaricus.

PubMed

Chen, He; Huang, Jie; Shi, Xiaoyu; Li, Yichao; Liu, Yu

2017-01-01

The efficacy of Lactobacillus delbrueckii subsp. bulgaricus as starter cultures for the dairy industry depends largely on the number of viable and active cells. Freeze-drying is the most convenient and successful method to preserve the bacterial cells. However, not all strains survived during freeze-drying. The effects of six substances including NaCl, sorbitol, mannitol, mannose, sodium glutamate, betaine added to the MRS medium on the growth and freeze-drying survival rate and viable counts of Lb. delbrueckii subsp. bulgaricus were studied through a single-factor test and Plackett-Burman design. Subsequently, the optimum freeze-drying conditions of Lb. delbrueckii subsp. bulgaricus were determined. Lb. delbrueckii subsp. bulgaricus survival rates were up to the maximum of 42.7%, 45.4%, 23.6%, while the concentrations of NaCl, sorbitol, sodium glutamate were 0.6%, 0.15%, 0.09%, respectively. In the optimum concentration, the viable counts in broth is 6.1, 6.9, 5.13 (×108 CFU/mL), respectively; the viable counts in freeze-drying power are 3.09, 5.2, 2.7 (×1010 CFU/g), respectively. Three antifreeze factors including NaCl, sorbitol, sodium glutamate have a positive effect on the growth and freeze-drying of Lb. delbrueckii subsp. bulgaricus. The results are beneficial for developing Lb. delbrueckii subsp. bulgaricus.

Tomato Fruit and Seed Colonization by Clavibacter michiganensis subsp. michiganensis through External and Internal Routes

PubMed Central

Tancos, Matthew A.; Chalupowicz, Laura; Barash, Isaac; Manulis-Sasson, Shulamit

2013-01-01

The Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis, causal agent of bacterial wilt and canker of tomato, is an economically devastating pathogen that inflicts considerable damage throughout all major tomato-producing regions. Annual outbreaks continue to occur in New York, where C. michiganensis subsp. michiganensis spreads via infected transplants, trellising stakes, tools, and/or soil. Globally, new outbreaks can be accompanied by the introduction of contaminated seed stock; however, the route of seed infection, especially the role of fruit lesions, remains undefined. In order to investigate the modes of seed infection, New York C. michiganensis subsp. michiganensis field strains were stably transformed with a gene encoding enhanced green fluorescent protein (eGFP). A constitutively eGFP-expressing virulent C. michiganensis subsp. michiganensis isolate, GCMM-22, was used to demonstrate that C. michiganensis subsp. michiganensis could not only access seeds systemically through the xylem but also externally through tomato fruit lesions, which harbored high intra- and intercellular populations. Active movement and expansion of bacteria into the fruit mesocarp and nearby xylem vessels followed, once the fruits began to ripen. These results highlight the ability of C. michiganensis subsp. michiganensis to invade tomato fruits and seeds through multiple entry routes. PMID:24014525

Effects of Pistacia atlantica subsp. kurdica on Growth and Aflatoxin Production by Aspergillus parasiticus

PubMed Central

Khodavaisy, Sadegh; Rezaie, Sassan; Noorbakhsh, Fatemeh; Baghdadi, Elham; Sharifynia, Somayeh; Aala, Farzad

2016-01-01

Background Aflatoxins are highly toxic secondary metabolites mainly produced by Aspergillus parasiticus. This species can contaminate a wide range of agricultural commodities, including cereals, peanuts, and crops in the field. In recent years, research on medicinal herbs, such as Pistacia atlantica subsp. kurdica, have led to reduced microbial growth, and these herbs also have a particular effect on the production of aflatoxins as carcinogenic compounds. Objectives In this study, we to examine P. atlantica subsp. kurdica as a natural compound used to inhibit the growth of A. parasiticus and to act as an anti-mycotoxin. Materials and Methods In vitro antifungal susceptibility testing of P. atlantica subsp. kurdica for A. parasiticus was performed according to CLSI document M38-A2. The rate of aflatoxin production was determined using the HPLC technique after exposure to different concentrations (62.5 - 125 mg/mL) of the gum. The changes in expression levels of the aflR gene were analyzed with a quantitative real-time PCR assay. Results The results showed that P. atlantica subsp. kurdica can inhibit A. parasiticus growth at a concentration of 125 mg/mL. HPLC results revealed a significant decrease in aflatoxin production with 125 mg/mL of P. atlantica subsp. kurdica, and AFL-B1 production was entirely inhibited. Based on quantitative real-time PCR results, the rate of aflR gene expression was significantly decreased after treatment with P. atlantica subsp. kurdica. Conclusions Pistacia atlantica subsp. kurdica has anti-toxic properties in addition to an inhibitory effect on A. parasiticus growth, and is able to decrease aflatoxin production effectively in a dose-dependent manner. Therefore, this herbal extract maybe considered a potential anti-mycotoxin agent in medicine or industrial agriculture. PMID:27800127

Bioaccessible Antioxidants in Milk Fermented by Bifidobacterium longum subsp. longum Strains

PubMed Central

Gagnon, Mérilie; Savard, Patricia; Rivière, Audrey; LaPointe, Gisèle

2015-01-01

Bifidobacterium longum subsp. longum is among the dominant species of the human gastrointestinal microbiota and could thus have potential as probiotics. New targets such as antioxidant properties have interest for beneficial effects on health. The objective of this study was to evaluate the bioaccessibility of antioxidants in milk fermented by selected B. longum subsp. longum strains during in vitro dynamic digestion. The antioxidant capacity of cell extracts from 38 strains, of which 32 belong to B. longum subsp. longum, was evaluated with the ORAC (oxygen radical absorbance capacity) method. On the basis of screening and gene sequence typing by multilocus locus sequence analysis (MLSA), five strains were chosen for fermenting reconstituted skim milk. Antioxidant capacity varied among the strains tested (P = 0.0009). Two strains of B. longum subsp. longum (CUETM 172 and 171) showed significantly higher ORAC values than the other bifidobacteria strains. However, there does not appear to be a relationship between gene sequence types and antioxidant capacity. The milk fermented by each of the five strains selected (CUETM 268, 172, 245, 247, or PRO 16-10) did not have higher initial ORAC values compared to the nonfermented milk samples. However, higher bioaccessibility of antioxidants in fermented milk (175–358%) was observed during digestion. PMID:25802836

Use of Potential Probiotic Lactic Acid Bacteria (LAB) Biofilms for the Control of Listeria monocytogenes, Salmonella Typhimurium, and Escherichia coli O157:H7 Biofilms Formation

PubMed Central

Gómez, Natacha C.; Ramiro, Juan M. P.; Quecan, Beatriz X. V.; de Melo Franco, Bernadette D. G.

2016-01-01

Use of probiotic biofilms can be an alternative approach for reducing the formation of pathogenic biofilms in food industries. The aims of this study were (i) to evaluate the probiotic properties of bacteriocinogenic (Lactococcus lactis VB69, L. lactis VB94, Lactobacillus sakei MBSa1, and Lactobacillus curvatus MBSa3) and non-bacteriocinogenic (L. lactis 368, Lactobacillus helveticus 354, Lactobacillus casei 40, and Weissela viridescens 113) lactic acid bacteria (LAB) isolated from Brazilian’s foods and (ii) to develop protective biofilms with these strains and test them for exclusion of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium. LAB were tested for survival in acid and bile salt conditions, surface properties, biosurfactant production, β-galactosidase and gelatinase activity, antibiotic resistance and presence of virulence genes. Most strains survived exposure to pH 2 and 4% bile salts. The highest percentages of auto-aggregation were obtained after 24 h of incubation. Sixty-seven percentage auto-aggregation value was observed in W. viridescens 113 and Lactobacillus curvatus MBSa3 exhibited the highest co-aggregation (69% with Listeria monocytogenes and 74.6% with E. coli O157:H7), while the lowest co-aggregation was exhibited by W. viridescens 113 (53.4% with Listeria monocytogenes and 38% with E. coli O157:H7). Tests for hemolytic activity, bacterial cell adherence with xylene, and drop collapse confirmed the biosurfactant-producing ability of most strains. Only one strain (L. lactis 368) produced β-galactosidase. All strains were negative for virulence genes cob, ccf, cylLL, cylLs, cyllM, cylB, cylA and efaAfs and gelatinase production. The antibiotic susceptibility tests indicated that the MIC for ciprofloxacin, clindamycin, gentamicin, kanamycin, and streptomycin did not exceed the epidemiological cut-off suggested by the European Food Safety Authority. Some strains were resistant to one or more antibiotics and

Applicability of a microbial Time Temperature Indicator (TTI) for monitoring spoilage of modified atmosphere packed minced meat.

PubMed

Vaikousi, Hariklia; Biliaderis, Costas G; Koutsoumanis, Konstantinos P

2009-08-15

The applicability of a microbial Time Temperature Indicator (TTI) prototype, based on the growth and metabolic activity of a Lactobacillus sakei strain developed in a previous study, in monitoring quality of modified atmosphere packed (MAP) minced beef was evaluated at conditions simulating the chill chain. At all storage temperatures examined (0, 5, 10, 15 degrees C), the results showed that lactic acid bacteria (LAB) were the dominant bacteria and can be used as a good spoilage index of MAP minced beef. The end of product's shelf life as revealed by the sensory evaluation coincided with a LAB population level of 7 log(10) CFU/g. For all temperatures tested, the growth of L. sakei in the TTI resembled closely the growth of LAB in the meat product, with similar temperature dependence of the micro(max) and thus similar activation energy values calculated as 111.90 and 106.90 kJ/mol, for the two systems, respectively. In addition, the end point of TTI colour change coincided with the time of sensory rejection point of the beef product during its storage under isothermal chilled temperature conditions. The estimated activation energy, E(alpha), values obtained for parameters related to the response of DeltaE (total colour change of the TTI) describing the kinetics of colour change of the TTI during isothermal storage (i.e. the maximum specific rate of DeltaEpsilon evolution curve, micro(DeltaEpsilon), and also the reciprocal of t(i), time at which half of the maximum DeltaEpsilon is reached), were 112.77 and 127.28 kJ/mol, respectively. Finally, the application of the microbial TTI in monitoring the quality deterioration of MAP minced beef due to spoilage was further evaluated under dynamic conditions of storage, using two separate low temperature periodic changing scenarios, resembling the actual conditions occurring in the distribution chill chain. The results showed that the end point of TTI, after storage at those fluctuating temperature conditions, was noted very

Isolation of Vibrio tapetis from two native fish species (Genypterus chilensis and Paralichthys adspersus) reared in Chile and description of Vibrio tapetis subsp. quintayensis subsp. nov.

PubMed

Levican, Arturo; Lasa, Aide; Irgang, Rute; Romalde, Jesús L; Poblete-Morales, Matías; Avendaño-Herrera, Ruben

2017-04-01

A group of seven Chilean isolates presumptively belonging to Vibrio tapetis was isolated from diseased fine flounders (Paralichthys adspersus) and red conger eel (Genypterus chilensis) experimentally reared in Quintay (Chile). All isolates were confirmed as members of V. tapetis on the basis of matrix-assisted laser desorption ionization time-of-flight MS, 16S rRNA gene sequencing, DNA-DNA hybridization values and G+C content. The ERIC-PCR and REP-PCR patterns were homogeneous among those isolates recovered from the same host (red conger or fine flounders), but distinct from the type strains V. tapetis subsp. tapetis CECT 4600T and V. tapetis subsp. britannicus CECT 8161T. On the basis of atpA, rpoA, rpoD, recA and pyrH gene sequence similarities (99.7-100 %) and clustering in the phylogenetic trees, the red conger isolates (Q20, Q047, Q48 and Q50) were confirmed as representing V. tapetis subsp. tapetis. However, they differed from V. tapetis subsp. tapetis CECT 4600T in their lipase, alpha quimiotripsin and non-acid phosphatase production. On the other hand, the fine flounder isolates (QL-9T, QL-35 and QL-41) showed rpoD, recA and pyrH gene sequence similarities ranging from 91.6 to 97.7 % with the type strains of the two V. tapetis subspecies (CECT 4600T and CECT 8161T) and consistently clustered together as an independent phylogenetic line within V. tapetis. Moreover, they could be differentiated phenotypically from strains CECT 4600T and CECT 8161T by nine and three different biochemical tests, respectively. In conclusion, the presence of V. tapetis in diseased red conger eel and fine flounder was demonstrated, extending the known host range and geographical location for this pathogen. Furthermore, this study demonstrates that the three isolates from fine flounder represent a novel subdivision within V. tapetis, for which the name V. tapetis subsp. quintayensis subsp. nov. is proposed and with QL-9T (=CECT 8851T=LMG 28759T) as the type strain. Although QL

Aeromonas hydrophila subsp. dhakensis Isolated from Feces, Water and Fish in Mediterranean Spain

PubMed Central

Esteve, Consuelo; Alcaide, Elena; Blasco, María Dolores

2012-01-01

Eight Aeromonas hydrophila-like arabinose-negative isolates from diverse sources (i.e., river freshwater, cooling-system water pond, diseased wild European eels, and human stools) sampled in Valencia (Spain) during 2004–2005, were characterized by 16S rRNA gene sequencing and extensive biochemical testing along with reference strains of most Aeromonas species. These isolates and all reference strains of A. hydrophila subsp. dhakensis and A. aquariorum showed a 16S rRNA sequence similarity of 99.8–100%, and they all shared an identical phenotype. This matched exactly with that of A. hydrophila subsp. dhakensis since all strains displayed positive responses to the Voges-Prokauer test and to the use of dl-lactate. This is the first report of A. hydrophila subsp. dhakensis recovered from environmental samples, and further, from its original isolation in India during 1993–1994. This was accurately identified and segregated from other clinical aeromonads (A. hydrophila subsp. hydrophila, A. caviae, A. veronii biovars veronii and sobria, A. trota, A. schubertii and A. jandaei) by using biochemical key tests. The API 20 E profile for all strains included in A. hydrophila subsp. dhakensis was 7047125. The prevalence of this species in Spanish sources was higher for water (9.4%) than for feces (6%) or eels (1.3%). Isolates recovered as pure cultures from diseased eels were moderately virulent (LD50 of 3.3×106 CFU fish−1) to challenged eels in experimental trials. They were all resistant to ticarcillin, amoxicillin-clavuranic acid, cefoxitin, and imipenem, regardless of its source. Our data point to A. hydrophila subsp. dhakensis as an emerging pathogen for humans and fish in temperate countries. PMID:22472298

PCR-Mediated Detection and Quantification of the Goss's Wilt Pathogen Clavibacter michiganensis subsp. nebraskensis Via a Novel Gene Target.

PubMed

McNally, R Ryan; Ishimaru, Carol A; Malvick, Dean K

2016-12-01

Goss's leaf blight and wilt of maize (corn) is a significant and reemerging disease caused by the bacterium Clavibacter michiganensis subsp. nebraskensis. Despite its importance, molecular tools for diagnosing and studying this disease remain limited. We report the identification of CMN_01184 as a novel gene target and its use in conventional PCR (cPCR) and SYBR green-based quantitative PCR (qPCR) assays for specific detection and quantification of C. michiganensis subsp. nebraskensis. The cPCR and qPCR assays based on primers targeting CMN_01184 specifically amplified only C. michiganensis subsp. nebraskensis among a diverse collection of 129 bacterial and fungal isolates, including multiple maize bacterial and fungal pathogens, environmental organisms from agricultural fields, and all known subspecies of C. michiganensis. Specificity of the assays for detection of only C. michiganensis subsp. nebraskensis was also validated with field samples of C. michiganensis subsp. nebraskensis-infected and uninfected maize leaves and C. michiganensis subsp. nebraskensis-infested and uninfested soil. Detection limits were determined at 30 and 3 ng of pure C. michiganensis subsp. nebraskensis DNA, and 100 and 10 CFU of C. michiganensis subsp. nebraskensis for the cPCR and qPCR assays, respectively. Infection of maize leaves by C. michiganensis subsp. nebraskensis was quantified from infected field samples and was standardized using an internal maize DNA control. These novel, specific, and sensitive PCR assays based on CMN_01184 are effective for diagnosis of Goss's wilt and for studies of the epidemiology and host-pathogen interactions of C. michiganensis subsp. nebraskensis.

A Novel Enzyme-Linked Immunosorbent Assay for Diagnosis of Mycobacterium avium subsp. paratuberculosis Infections (Johne's Disease) in Cattle

PubMed Central

Speer, C. A.; Scott, M. Cathy; Bannantine, John P.; Waters, W. Ray; Mori, Yasuyuki; Whitlock, Robert H.; Eda, Shigetoshi

2006-01-01

Enzyme-linked immunosorbent assays (ELISAs) for the diagnosis of Johne's disease (JD), caused by Mycobacterium avium subsp. paratuberculosis, were developed using whole bacilli treated with formaldehyde (called WELISA) or surface antigens obtained by treatment of M. avium subsp. paratuberculosis bacilli with formaldehyde and then brief sonication (called SELISA). ELISA plates were coated with either whole bacilli or sonicated antigens and tested for reactivity against serum obtained from JD-positive and JD-negative cattle or from calves experimentally inoculated with M. avium subsp. paratuberculosis, Mycobacterium avium subsp. avium, or Mycobacterium bovis. Because the initial results obtained from the WELISA and SELISA were similar, most of the subsequent experiments reported herein were performed using the SELISA method. To optimize the SELISA test, various concentrations (3.7 to 37%) of formaldehyde and intervals of sonication (2 to 300 s) were tested. With an increase in formaldehyde concentration and a decreased interval of sonication, there was a concomitant decrease in nonspecific binding by the SELISA. SELISAs prepared by treating M. avium subsp. paratuberculosis with 37% formaldehyde and then a 2-s burst of sonication produced the greatest difference (7×) between M. avium subsp. paratuberculosis-negative and M. avium subsp. paratuberculosis-positive serum samples. The diagnostic sensitivity and specificity for JD by the SELISA were greater than 95%. The SELISA showed subspecies-specific detection of M. avium subsp. paratuberculosis infections in calves experimentally inoculated with M. avium subsp. paratuberculosis or other mycobacteria. Based on diagnostic sensitivity and specificity, the SELISA appears superior to the commercial ELISAs routinely used for the diagnosis of JD. PMID:16682472

Laminaria japonica Extract, an Inhibitor of Clavibater michiganense Subsp. Sepedonicum

PubMed Central

Cai, Jin; Feng, Jia; Xie, Shulian; Wang, Feipeng; Xu, Qiufeng

2014-01-01

Bacterial ring rot of potato is one of the most serious potato plant and tuber diseases. Laminaria japonica extract was investigated for its antimicrobial activity against Clavibater michiganense subsp. sepedonicum (Spieckermann & Kotthoff) Davis et al., the causative agent of bacterial ring rot of potato. The results showed that the optimum extraction conditions of antimicrobial substances from L. japonica were an extraction temperature of 80°C, an extraction time of 12 h, and a solid to liquid ratio of 1∶25. Active compounds of L. japonica were isolated by solvent partition, thin layer chromatography (TLC) and column chromatography. All nineteen fractionations had antimicrobial activities against C. michiganense subsp. sepedonicum, while Fractionation three (Fr.3) had the highest (P<0.05) antimicrobial activity. Chemical composition analysis identified a total of 26 components in Fr.3. The main constituents of Fr.3 were alkanes (80.97%), esters (5.24%), acids (4.87%) and alcohols (2.21%). Antimicrobial activity of Fr.3 against C. michiganense subsp. sepedonicum could be attributed to its ability to damage the cell wall and cell membrane, induce the production of reactive oxygen species (ROS), increase cytosolic Ca2+ concentration, inhibit the glycolytic pathway (EMP) and tricarboxylic acid (TCA) cycle, inhibit protein and nucleic acid synthesis, and disrupt the normal cycle of DNA replication. These findings indicate that L. japonica extracts have potential for inhibiting C. michiganense subsp. sepedonicum. PMID:24714388

Bifidobacterium animalis subsp. lactis ATCC 27673 Is a Genomically Unique Strain within Its Conserved Subspecies

PubMed Central

Loquasto, Joseph R.; Barrangou, Rodolphe; Dudley, Edward G.; Stahl, Buffy; Chen, Chun

2013-01-01

Many strains of Bifidobacterium animalis subsp. lactis are considered health-promoting probiotic microorganisms and are commonly formulated into fermented dairy foods. Analyses of previously sequenced genomes of B. animalis subsp. lactis have revealed little genetic diversity, suggesting that it is a monomorphic subspecies. However, during a multilocus sequence typing survey of Bifidobacterium, it was revealed that B. animalis subsp. lactis ATCC 27673 gave a profile distinct from that of the other strains of the subspecies. As part of an ongoing study designed to understand the genetic diversity of this subspecies, the genome of this strain was sequenced and compared to other sequenced genomes of B. animalis subsp. lactis and B. animalis subsp. animalis. The complete genome of ATCC 27673 was 1,963,012 bp, contained 1,616 genes and 4 rRNA operons, and had a G+C content of 61.55%. Comparative analyses revealed that the genome of ATCC 27673 contained six distinct genomic islands encoding 83 open reading frames not found in other strains of the same subspecies. In four islands, either phage or mobile genetic elements were identified. In island 6, a novel clustered regularly interspaced short palindromic repeat (CRISPR) locus which contained 81 unique spacers was identified. This type I-E CRISPR-cas system differs from the type I-C systems previously identified in this subspecies, representing the first identification of a different system in B. animalis subsp. lactis. This study revealed that ATCC 27673 is a strain of B. animalis subsp. lactis with novel genetic content and suggests that the lack of genetic variability observed is likely due to the repeated sequencing of a limited number of widely distributed commercial strains. PMID:23995933

Rapid and sensitive method to identify Mycobacterium avium subsp. paratuberculosis in cow's milk by DNA methylase genotyping.

PubMed

Mundo, Silvia Leonor; Gilardoni, Liliana Rosa; Hoffman, Federico José; Lopez, Osvaldo Jorge

2013-03-01

Paratuberculosis is an infectious, chronic, and incurable disease that affects ruminants, caused by Mycobacterium avium subsp. paratuberculosis. This bacterium is shed primarily through feces of infected cows but can be also excreted in colostrum and milk and might survive pasteurization. Since an association of genomic sequences of M. avium subsp. paratuberculosis in patients with Crohn's disease has been described; it is of interest to rapidly detect M. avium subsp. paratuberculosis in milk for human consumption. IS900 insertion is used as a target for PCR amplification to identify the presence of M. avium subsp. paratuberculosis in biological samples. Two target sequences were selected: IS1 (155 bp) and IS2 (94 bp). These fragments have a 100% identity among all M. avium subsp. paratuberculosis strains sequenced. M. avium subsp. paratuberculosis was specifically concentrated from milk samples by immunomagnetic separation prior to performing PCR. The amplicons were characterized using DNA methylase Genotyping, i.e., the amplicons were methylated with 6-methyl-adenine and digested with restriction enzymes to confirm their identity. The methylated amplicons from 100 CFU of M. avium subsp. paratuberculosis can be visualized in a Western blot format using an anti-6-methyl-adenine monoclonal antibody. The use of DNA methyltransferase genotyping coupled to a scintillation proximity assay allows for the detection of up to 10 CFU of M. avium subsp. paratuberculosis per ml of milk. This test is rapid and sensitive and allows for automation and thus multiple samples can be tested at the same time.

Rapid and Sensitive Method To Identify Mycobacterium avium subsp. paratuberculosis in Cow's Milk by DNA Methylase Genotyping

PubMed Central

Mundo, Silvia Leonor; Gilardoni, Liliana Rosa; Hoffman, Federico José

2013-01-01

Paratuberculosis is an infectious, chronic, and incurable disease that affects ruminants, caused by Mycobacterium avium subsp. paratuberculosis. This bacterium is shed primarily through feces of infected cows but can be also excreted in colostrum and milk and might survive pasteurization. Since an association of genomic sequences of M. avium subsp. paratuberculosis in patients with Crohn's disease has been described; it is of interest to rapidly detect M. avium subsp. paratuberculosis in milk for human consumption. IS900 insertion is used as a target for PCR amplification to identify the presence of M. avium subsp. paratuberculosis in biological samples. Two target sequences were selected: IS1 (155 bp) and IS2 (94 bp). These fragments have a 100% identity among all M. avium subsp. paratuberculosis strains sequenced. M. avium subsp. paratuberculosis was specifically concentrated from milk samples by immunomagnetic separation prior to performing PCR. The amplicons were characterized using DNA methylase Genotyping, i.e., the amplicons were methylated with 6-methyl-adenine and digested with restriction enzymes to confirm their identity. The methylated amplicons from 100 CFU of M. avium subsp. paratuberculosis can be visualized in a Western blot format using an anti-6-methyl-adenine monoclonal antibody. The use of DNA methyltransferase genotyping coupled to a scintillation proximity assay allows for the detection of up to 10 CFU of M. avium subsp. paratuberculosis per ml of milk. This test is rapid and sensitive and allows for automation and thus multiple samples can be tested at the same time. PMID:23275511

Septic pneumonic tularaemia caused by Francisella tularensis subsp. holarctica biovar II.

PubMed

Fritzsch, Joerg; Splettstoesser, Wolf D

2010-09-01

This case of pneumonic tularaemia elucidates two aspects: it is believed to be the first documented case of bacteraemia caused by Francisella tularensis subsp. holarctica biovar II; furthermore, it illustrates the remission of septic pneumonic tularaemia without appropriate anti-infective therapy. A blood culture from a patient with community-acquired pneumonia was found to be positive for F. tularensis subsp. holarctica biovar II after 10 days of cultivation. Meanwhile, the patient had been treated with ceftriaxone, followed by sultamicillin and clindamycin. The patient continued suffering from fever of up to 40.7 degrees C and rising C-reactive protein (CRP) for 4 days before the fever and CRP declined. The isolated strain was later tested and found to be resistant to the antibiotics used. The present case underlines that F. tularensis subsp. holarctica infections may cause severe symptoms but mostly have a favourable outcome.

Toxicity of Food-Grade TiO2 to Commensal Intestinal and Transient Food-Borne Bacteria: New Insights Using Nano-SIMS and Synchrotron UV Fluorescence Imaging.

PubMed

Radziwill-Bienkowska, Joanna M; Talbot, Pauline; Kamphuis, Jasper B J; Robert, Véronique; Cartier, Christel; Fourquaux, Isabelle; Lentzen, Esther; Audinot, Jean-Nicolas; Jamme, Frédéric; Réfrégiers, Matthieu; Bardowski, Jacek K; Langella, Philippe; Kowalczyk, Magdalena; Houdeau, Eric; Thomas, Muriel; Mercier-Bonin, Muriel

2018-01-01

Titanium dioxide (TiO 2 ) is commonly used as a food additive (E171 in the EU) for its whitening and opacifying properties. However, a risk of intestinal barrier disruption, including dysbiosis of the gut microbiota, is increasingly suspected because of the presence of a nano-sized fraction in this additive. We hypothesized that food-grade E171 and Aeroxyde P25 (identical to the NM-105 OECD reference nanomaterial in the European Union Joint Research Centre) interact with both commensal intestinal bacteria and transient food-borne bacteria under non-UV-irradiated conditions. Based on differences in their physicochemical properties, we expect a difference in their respective effects. To test these hypotheses, we chose a panel of eight Gram-positive/Gram-negative bacterial strains, isolated from different biotopes and belonging to the species Escherichia coli , Lactobacillus rhamnosus , Lactococcus lactis (subsp. lactis and cremoris ), Streptococcus thermophilus , and Lactobacillus sakei . Bacterial cells were exposed to food-grade E171 vs. P25 in vitro and the interactions were explored with innovative (nano)imaging methods. The ability of bacteria to trap TiO 2 was demonstrated using synchrotron UV fluorescence imaging with single cell resolution. Subsequent alterations in the growth profiles were shown, notably for the transient food-borne L. lactis and the commensal intestinal E. coli in contact with food-grade TiO 2 . However, for both species, the reduction in cell cultivability remained moderate, and the morphological and ultrastructural damages, observed with electron microscopy, were restricted to a small number of cells. E. coli exposed to food-grade TiO 2 showed some internalization of TiO 2 (7% of cells), observed with high-resolution nano-secondary ion mass spectrometry (Nano-SIMS) chemical imaging. Taken together, these data show that E171 may be trapped by commensal and transient food-borne bacteria within the gut. In return, it may induce some

Toxicity of Food-Grade TiO2 to Commensal Intestinal and Transient Food-Borne Bacteria: New Insights Using Nano-SIMS and Synchrotron UV Fluorescence Imaging

PubMed Central

Radziwill-Bienkowska, Joanna M.; Talbot, Pauline; Kamphuis, Jasper B. J.; Robert, Véronique; Cartier, Christel; Fourquaux, Isabelle; Lentzen, Esther; Audinot, Jean-Nicolas; Jamme, Frédéric; Réfrégiers, Matthieu; Bardowski, Jacek K.; Langella, Philippe; Kowalczyk, Magdalena; Houdeau, Eric; Thomas, Muriel; Mercier-Bonin, Muriel

2018-01-01

Titanium dioxide (TiO2) is commonly used as a food additive (E171 in the EU) for its whitening and opacifying properties. However, a risk of intestinal barrier disruption, including dysbiosis of the gut microbiota, is increasingly suspected because of the presence of a nano-sized fraction in this additive. We hypothesized that food-grade E171 and Aeroxyde P25 (identical to the NM-105 OECD reference nanomaterial in the European Union Joint Research Centre) interact with both commensal intestinal bacteria and transient food-borne bacteria under non-UV-irradiated conditions. Based on differences in their physicochemical properties, we expect a difference in their respective effects. To test these hypotheses, we chose a panel of eight Gram-positive/Gram-negative bacterial strains, isolated from different biotopes and belonging to the species Escherichia coli, Lactobacillus rhamnosus, Lactococcus lactis (subsp. lactis and cremoris), Streptococcus thermophilus, and Lactobacillus sakei. Bacterial cells were exposed to food-grade E171 vs. P25 in vitro and the interactions were explored with innovative (nano)imaging methods. The ability of bacteria to trap TiO2 was demonstrated using synchrotron UV fluorescence imaging with single cell resolution. Subsequent alterations in the growth profiles were shown, notably for the transient food-borne L. lactis and the commensal intestinal E. coli in contact with food-grade TiO2. However, for both species, the reduction in cell cultivability remained moderate, and the morphological and ultrastructural damages, observed with electron microscopy, were restricted to a small number of cells. E. coli exposed to food-grade TiO2 showed some internalization of TiO2 (7% of cells), observed with high-resolution nano-secondary ion mass spectrometry (Nano-SIMS) chemical imaging. Taken together, these data show that E171 may be trapped by commensal and transient food-borne bacteria within the gut. In return, it may induce some physiological

Draft Genome Sequences of Three Pectobacterium Strains Causing Blackleg of Potato: P. carotovorum subsp. brasiliensis ICMP 19477, P. atrosepticum ICMP 1526, and P. carotovorum subsp. carotovorum UGC32

PubMed Central

Fiers, Mark W. E. J.; Lu, Ashley; Armstrong, Karen F.

2015-01-01

Blackleg is a disease caused by several species of Pectobacterium that results in losses to potato crops worldwide. Here, we report the draft genomes of three taxonomically and geographically distinct blackleg-causing strains of Pectobacterium: P. carotovorum subsp. brasiliensis ICMP 19477, P. atrosepticum ICMP 1526, and P. carotovorum subsp. carotovorum UGC32. Comparison of these genomes will support the identification of common traits associated with their capacity to cause blackleg. PMID:26251497

Characterization of a novel bacteriophage, Phda1, infecting the histamine-producing Photobacterium damselae subsp. damselae.

PubMed

Yamaki, S; Kawai, Y; Yamazaki, K

2015-06-01

Photobacterium damselae subsp. damselae is a potent histamine-producing micro-organism. The aim of this study was to isolate and characterize a bacteriophage Phda1 that infected P. damselae subsp. damselae to inhibit its growth and histamine accumulation. Phda1 was isolated from a raw oyster, and the host range, morphology and the bacteriophage genome size were analysed. Phda1 formed a clear plaque only against P. damselae subsp. damselae JCM8969 among five Gram-positive and 32 Gram-negative bacterial strains tested. Phda1 belongs to the family Myoviridae, and its genome size was estimated as 35·2-39·5 kb. According to the one-step growth curve analysis, the latent period, rise period and burst size of Phda1 were 60 min, 50 min and 19 plaque-forming units per infected cell, respectively. Divalent cations, especially Ca(2+) and Mg(2+) , strongly improved Phda1 adsorption to the host cells and its propagation. Phda1 treatment delayed the growth and histamine production of P. damselae subsp. damselae in an in vitro challenge test. The bacteriophage Phda1 might serve as a potential antimicrobial agent to inhibit the histamine poisoning caused by P. damselae subsp. damselae. This is the first description of a bacteriophage specifically infecting P. damselae subsp. damselae and its potential applications. Bacteriophage therapy could prove useful in the prevention of histamine poisoning. © 2015 The Society for Applied Microbiology.

Detection of Goss's Wilt Pathogen Clavibacter michiganensis subsp. nebraskensis in Maize by Loop-Mediated Amplification.

PubMed

Yasuhara-Bell, Jarred; de Silva, Asoka; Heuchelin, Scott A; Chaky, Jennifer L; Alvarez, Anne M

2016-03-01

The Goss's wilt pathogen, Clavibacter michiganensis subsp. nebraskensis, can cause considerable losses in maize (Zea mays) production. Diagnosis of Goss's wilt currently is based on symptomology and identification of C. michiganensis subsp. nebraskensis, following isolation on a semiselective medium and/or serological testing. In an effort to provide a more efficient identification method, a loop-mediated amplification (LAMP) assay was developed to detect the tripartite ATP-independent periplasmic (TRAP)-type C4-dicarboxylate transport system large permease component and tested using strains of C. michiganensis subsp. nebraskensis, all other C. michiganensis subspecies and several genera of nontarget bacteria. Only strains of C. michiganensis subsp. nebraskensis reacted positively with the LAMP assay. The LAMP assay was then used to identify bacterial isolates from diseased maize. 16S rDNA and dnaA sequence analyses were used to confirm the identity of the maize isolates and validate assay specificity. The Cmm ImmunoStrip assay was included as a presumptive identification test of C. michiganensis subsp. nebraskensis at the species level. The Cmn-LAMP assay was further tested using symptomatic leaf tissue. The Cmn-LAMP assay was run in a hand-held real-time monitoring device (SMART-DART) and performed equally to in-lab quantitative polymerase chain reaction equipment. The Cmn-LAMP assay accurately identified C. michiganensis subsp. nebraskensis and has potential as a field test. The targeted sequence also has potential application in other molecular detection platforms.

Bioprocessing of some agro-industrial residues for endoglucanase production by the new subsp.; Streptomyces albogriseolus subsp. cellulolyticus strain NEAE-J

PubMed Central

El-Naggar, Noura El-Ahmady; Abdelwahed, Nayera A.M.; Saber, Wesam I.A.; Mohamed, Asem A.

2014-01-01

The use of low cost agro-industrial residues for the production of industrial enzymes is one of the ways to reduce significantly production costs. Cellulase producing actinomycetes were isolated from soil and decayed agricultural wastes. Among them, a potential culture, strain NEAE-J, was selected and identified on the basis of morphological, cultural, physiological and chemotaxonomic properties, together with 16S rDNA sequence. It is proposed that strain NEAE-J should be included in the species Streptomyces albogriseolus as a representative of a novel sub-species, Streptomyces albogriseolus subsp. cellulolyticus strain NEAE-J and sequencing product was deposited in the GenBank database under accession number JN229412. This organism was tested for its ability to produce endoglucanase and release reducing sugars from agro-industrial residues as substrates. Sugarcane bagasse was the most suitable substrate for endoglucanase production. Effects of process variables, namely incubation time, temperature, initial pH and nitrogen source on production of endoglucanase by submerged fermentation using Streptomyces albogriseolus subsp. cellulolyticus have been studied. Accordingly optimum conditions have been determined. Incubation temperature of 30 °C after 6 days, pH of 6.5, 1% sugarcane bagasse as carbon source and peptone as nitrogen source were found to be the optimum for endoglucanase production. Optimization of the process parameters resulted in about 2.6 fold increase in the endoglucanase activity. Therefore, Streptomyces albogriseolus subsp. cellulolyticus coud be potential microorganism for the intended application. PMID:25242966

Effect of Three Factors in Cheese Production (pH, Salt, and Heat) on Mycobacterium avium subsp. paratuberculosis Viability

PubMed Central

Sung, Nackmoon; Collins, Michael T.

2000-01-01

Low pH and salt are two factors contributing to the inactivation of bacterial pathogens during a 60-day curing period for cheese. The kinetics of inactivation for Mycobacterium avium subsp. paratuberculosis strains ATCC 19698 and Dominic were measured at 20°C under different pH and NaCl conditions commonly used in processing cheese. The corresponding D values (decimal reduction times; the time required to kill 1 log10 concentration of bacteria) were measured. Also measured were the D values for heat-treated and nonheated M. avium subsp. paratuberculosis in 50 mM acetate buffer (pH 5.0, 2% [wt/vol] NaCl) and a soft white Hispanic-style cheese (pH 6.0, 2% [wt/vol] NaCl). Samples were removed at various intervals until no viable cells were detected using the radiometric culture method (BACTEC) for enumeration of M. avium subsp. paratuberculosis. NaCl had little or no effect on the inactivation of M. avium subsp. paratuberculosis, and increasing NaCl concentrations were not associated with decreasing D values (faster killing) in the acetate buffer. Lower pHs, however, were significantly correlated with decreasing D values of M. avium subsp. paratuberculosis in the acetate buffer. The D values for heat-treated M. avium subsp. paratuberculosis ATCC 19698 in the cheese were higher than those predicted by studies done in acetate buffer. The heat-treated M. avium subsp. paratuberculosis strains had lower D values than the nonheated cells (faster killing) both in the acetate buffer (pH 5, 2% [wt/vol] NaCl) and in the soft white cheese. The D value for heat-treated M. avium subsp. paratuberculosis ATCC 19698 in the cheese (36.5 days) suggests that heat treatment of raw milk coupled with a 60-day curing period will inactivate about 103 cells of M. avium subsp. paratuberculosis per ml. PMID:10742208

Anoxybacillus kamchatkensis subsp. asaccharedens subsp. nov., a thermophilic bacterium isolated from a hot spring in Batman.

PubMed

Gul-Guven, Reyhan; Guven, Kemal; Poli, Annarita; Nicolaus, Barbara

2008-12-01

A new thermophilic spore-forming strain KG8(T) was isolated from the mud of Taslidere hot spring in Batman. Strain KG8(T) was aerobe, Gram-positive, rod-shaped, motile, occurring in pairs or filamentous. Growth was observed from 35-65 degrees C (optimum 55 degrees C) and at pH 5.5-9.5 (optimum pH 7.5). It was capable of utilizing starch, growth was observed until 3% NaCl (w/v) and it was positive for nitrate reduction. On the basis of 16S rRNA gene sequence similarity, strain KG8(T) was shown to be related most closely to Anoxybacillus species. Chemotaxonomic data (major isoprenoid quinone-menaquinone-7; major fatty acid-iso-C15:0 and iso-C17:0) supported the affiliation of strain KG8(T) to the genus Anoxybacillus. The results of DNA-DNA hybridization, physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain KG8(T). Based on these results we propose assigning a novel subspecies of Anoxybacillus kamchatkensis, to be named Anoxybacillus kamchatkensis subsp. asaccharedens subsp. nov. with the type strain KG8(T) (DSM 18475(T)=CIP 109280(T)).

Proteomes of Lactobacillus delbrueckii subsp. bulgaricus LBB.B5 Incubated in Milk at Optimal and Low Temperatures

PubMed Central

Yin, Xiaochen; Salemi, Michelle R.; Phinney, Brett S.; Gotcheva, Velitchka; Angelov, Angel

2017-01-01

ABSTRACT We identified the proteins synthesized by Lactobacillus delbrueckii subsp. bulgaricus strain LBB.B5 in laboratory culture medium (MRS) at 37°C and milk at 37 and 4°C. Cell-associated proteins were measured by gel-free, shotgun proteomics using high-performance liquid chromatography coupled with tandem mass spectrophotometry. A total of 635 proteins were recovered from all cultures, among which 72 proteins were milk associated (unique or significantly more abundant in milk). LBB.B5 responded to milk by increasing the production of proteins required for purine biosynthesis, carbohydrate metabolism (LacZ and ManM), energy metabolism (TpiA, PgK, Eno, SdhA, and GapN), amino acid synthesis (MetE, CysK, LBU0412, and AspC) and transport (GlnM and GlnP), and stress response (Trx, MsrA, MecA, and SmpB). The requirement for purines was confirmed by the significantly improved cell yields of L. delbrueckii subsp. bulgaricus when incubated in milk supplemented with adenine and guanine. The L. delbrueckii subsp. bulgaricus-expressed proteome in milk changed upon incubation at 4°C for 5 days and included increased levels of 17 proteins, several of which confer functions in stress tolerance (AddB, UvrC, RecA, and DnaJ). However, even with the activation of stress responses in either milk or MRS, L. delbrueckii subsp. bulgaricus did not survive passage through the murine digestive tract. These findings inform efforts to understand how L. delbrueckii subsp. bulgaricus is adapted to the dairy environment and its implications for its health-benefiting properties in the human digestive tract. IMPORTANCE Lactobacillus delbrueckii subsp. bulgaricus has a long history of use in yogurt production. Although commonly cocultured with Streptococcus salivarius subsp. thermophilus in milk, fundamental knowledge of the adaptive responses of L. delbrueckii subsp. bulgaricus to the dairy environment and the consequences of those responses on the use of L. delbrueckii subsp

Proteomes of Lactobacillus delbrueckii subsp. bulgaricus LBB.B5 Incubated in Milk at Optimal and Low Temperatures.

PubMed

Yin, Xiaochen; Salemi, Michelle R; Phinney, Brett S; Gotcheva, Velitchka; Angelov, Angel; Marco, Maria L

2017-01-01

We identified the proteins synthesized by Lactobacillus delbrueckii subsp. bulgaricus strain LBB.B5 in laboratory culture medium (MRS) at 37°C and milk at 37 and 4°C. Cell-associated proteins were measured by gel-free, shotgun proteomics using high-performance liquid chromatography coupled with tandem mass spectrophotometry. A total of 635 proteins were recovered from all cultures, among which 72 proteins were milk associated (unique or significantly more abundant in milk). LBB.B5 responded to milk by increasing the production of proteins required for purine biosynthesis, carbohydrate metabolism (LacZ and ManM), energy metabolism (TpiA, PgK, Eno, SdhA, and GapN), amino acid synthesis (MetE, CysK, LBU0412, and AspC) and transport (GlnM and GlnP), and stress response (Trx, MsrA, MecA, and SmpB). The requirement for purines was confirmed by the significantly improved cell yields of L. delbrueckii subsp. bulgaricus when incubated in milk supplemented with adenine and guanine. The L. delbrueckii subsp. bulgaricus -expressed proteome in milk changed upon incubation at 4°C for 5 days and included increased levels of 17 proteins, several of which confer functions in stress tolerance (AddB, UvrC, RecA, and DnaJ). However, even with the activation of stress responses in either milk or MRS, L. delbrueckii subsp. bulgaricus did not survive passage through the murine digestive tract. These findings inform efforts to understand how L. delbrueckii subsp. bulgaricus is adapted to the dairy environment and its implications for its health-benefiting properties in the human digestive tract. IMPORTANCE Lactobacillus delbrueckii subsp. bulgaricus has a long history of use in yogurt production. Although commonly cocultured with Streptococcus salivarius subsp. thermophilus in milk, fundamental knowledge of the adaptive responses of L. delbrueckii subsp. bulgaricus to the dairy environment and the consequences of those responses on the use of L. delbrueckii subsp. bulgaricus as

Distribution of Bacillus thuringiensis subsp. israelensis in Soil of a Swiss Wetland reserve after 22 years of mosquito control.

PubMed

Guidi, Valeria; Patocchi, Nicola; Lüthy, Peter; Tonolla, Mauro

2011-06-01

Recurrent treatments with Bacillus thuringiensis subsp. israelensis are required to control the floodwater mosquito Aedes vexans that breeds in large numbers in the wetlands of the Bolle di Magadino Reserve in Canton Ticino, Switzerland. Interventions have been carried out since 1988. In the present study, the spatial distribution of resting B. thuringiensis subsp. israelensis spores in the soil was measured. The B. thuringiensis subsp. israelensis concentration was determined in soil samples collected along six transects covering different elevations within the periodically flooded zones. A total of 258 samples were processed and analyzed by quantitative PCR that targeted an identical fragment of 159 bp for the B. thuringiensis subsp. israelensis cry4Aa and cry4Ba genes. B. thuringiensis subsp. israelensis spores were found to persist in soils of the wetland reserve at concentrations of up to 6.8 log per gram of soil. Continuous accumulation due to regular treatments could be excluded, as the decrease in spores amounted to 95.8% (95% confidence interval, 93.9 to 97.7%). The distribution of spores was correlated to the number of B. thuringiensis subsp. israelensis treatments, the elevation of the sampling point, and the duration of the flooding periods. The number of B. thuringiensis subsp. israelensis treatments was the major factor influencing the distribution of spores in the different topographic zones (P < 0.0001). These findings indicated that B. thuringiensis subsp. israelensis spores are rather immobile after their introduction into the environment.

Xylella fastidiosa Isolates from Both subsp. multiplex and fastidiosa Cause Disease on Southern Highbush Blueberry (Vaccinium sp.) Under Greenhouse Conditions.

PubMed

Oliver, J E; Cobine, P A; De La Fuente, L

2015-07-01

Xylella fastidiosa is a xylem-limited gram-negative plant pathogen that affects numerous crop species, including grape, citrus, peach, pecan, and almond. Recently, X. fastidiosa has also been found to be the cause of bacterial leaf scorch on blueberry in the southeastern United States. Thus far, all X. fastidiosa isolates obtained from infected blueberry have been classified as X. fastidiosa subsp. multiplex; however, X. fastidiosa subsp. fastidiosa isolates are also present in the southeastern United States and commonly cause Pierce's disease of grapevines. In this study, seven southeastern U.S. isolates of X. fastidiosa, including three X. fastidiosa subsp. fastidiosa isolates from grape, one X. fastidiosa subsp. fastidiosa isolate from elderberry, and three X. fastidiosa subsp. multiplex isolates from blueberry, were used to infect the southern highbush blueberry 'Rebel'. Following inoculation, all isolates colonized blueberry, and isolates from both X. fastidiosa subsp. multiplex and X. fastidiosa subsp. fastidiosa caused symptoms, including characteristic stem yellowing and leaf scorch symptoms as well as dieback of the stem tips. Two X. fastidiosa subsp. multiplex isolates from blueberry caused more severe symptoms than the other isolates examined, and infection with these two isolates also had a significant impact on host mineral nutrient content in sap and leaves. These findings have potential implications for understanding X. fastidiosa host adaptation and expansion and the development of emerging diseases caused by this bacterium.

Isolation of Bartonella henselae and Two New Bartonella Subspecies, Bartonella koehlerae Subspecies boulouisii subsp. nov. and Bartonella koehlerae Subspecies bothieri subsp. nov. from Free-Ranging Californian Mountain Lions and Bobcats

PubMed Central

Chomel, Bruno B.; Molia, Sophie; Kasten, Rickie W.; Borgo, Gina M.; Stuckey, Matthew J.; Maruyama, Soichi; Chang, Chao-chin; Haddad, Nadia; Koehler, Jane E.

2016-01-01

Domestic cats are the natural reservoir of Bartonella henselae, B. clarridgeiae and B. koehlerae. To determine the role of wild felids in the epidemiology of Bartonella infections, blood was collected from 14 free-ranging California mountain lions (Puma concolor) and 19 bobcats (Lynx rufus). Bartonella spp. were isolated from four (29%) mountain lions and seven (37%) bobcats. These isolates were characterized using growth characteristics, biochemical reactions, molecular techniques, including PCR-RFLP of selected genes or interspacer region, pulsed-field gel electrophoresis (PFGE), partial sequencing of several genes, and DNA-DNA hybridization. Two isolates were identical to B. henselae genotype II. All other isolates were distinguished from B. henselae and B. koehlerae by PCR-RFLP of the gltA gene using endonucleases HhaI, TaqI and AciI, with the latter two discriminating between the mountain lion and the bobcat isolates. These two novel isolates displayed specific PFGE profiles distinct from B. henselae, B. koehlerae and B. clarridgeiae. Sequences of amplified gene fragments from the three mountain lion and six bobcat isolates were closely related to, but distinct from, B. henselae and B. koehlerae. Finally, DNA-DNA hybridization studies demonstrated that the mountain lion and bobcat strains are most closely related to B. koehlerae. We propose naming the mountain lion isolates B. koehlerae subsp. boulouisii subsp. nov. (type strain: L-42-94), and the bobcat isolates B. koehlerae subsp. bothieri subsp. nov. (type strain: L-17-96), and to emend B. koehlerae as B. koehlerae subsp. koehlerae. The mode of transmission and the zoonotic potential of these new Bartonella subspecies remain to be determined. PMID:26981874

Complete Genome Sequence of the Yogurt Isolate Lactobacillus delbrueckii subsp. bulgaricus ACA-DC 87.

PubMed

Alexandraki, Voula; Kazou, Maria; Pot, Bruno; Tsakalidou, Effie; Papadimitriou, Konstantinos

2017-08-24

Lactobacillus delbrueckii subsp. bulgaricus is widely used in the production of yogurt and cheese. In this study, we present the complete genome sequence of L. delbrueckii subsp. bulgaricus ACA-DC 87 isolated from traditional Greek yogurt. Whole-genome analysis may reveal desirable technological traits of the strain for dairy fermentations. Copyright © 2017 Alexandraki et al.

A Rapid Method for Quantifying Viable Mycobacterium avium subsp. paratuberculosis in Cellular Infection Assays

PubMed Central

Pooley, Hannah B.; de Silva, Kumudika; Purdie, Auriol C.; Begg, Douglas J.; Whittington, Richard J.

2016-01-01

ABSTRACT Determining the viability of bacteria is a key outcome of in vitro cellular infection assays. Currently, this is done by culture, which is problematic for fastidious slow-growing bacteria such as Mycobacterium avium subsp. paratuberculosis, where it can take up to 4 months to confirm growth. This study aimed to identify an assay that can rapidly quantify the number of viable M. avium subsp. paratuberculosis cells in a cellular sample. Three commercially available bacterial viability assays along with a modified liquid culture method coupled with high-throughput quantitative PCR growth detection were assessed. Criteria for assessment included the ability of each assay to differentiate live and dead M. avium subsp. paratuberculosis organisms and their accuracy at low bacterial concentrations. Using the culture-based method, M. avium subsp. paratuberculosis growth was reliably detected and quantified within 2 weeks. There was a strong linear association between the 2-week growth rate and the initial inoculum concentration. The number of viable M. avium subsp. paratuberculosis cells in an unknown sample was quantified based on the growth rate, by using growth standards. In contrast, none of the commercially available viability assays were suitable for use with samples from in vitro cellular infection assays. IMPORTANCE Rapid quantification of the viability of Mycobacterium avium subsp. paratuberculosis in samples from in vitro cellular infection assays is important, as it allows these assays to be carried out on a large scale. In vitro cellular infection assays can function as a preliminary screening tool, for vaccine development or antimicrobial screening, and also to extend findings derived from experimental animal trials. Currently, by using culture, it takes up to 4 months to obtain quantifiable results regarding M. avium subsp. paratuberculosis viability after an in vitro infection assay; however, with the quantitative PCR and liquid culture method

Experimental infection of dogs with Bartonella henselae and Bartonella vinsonii subsp. berkhoffii.

PubMed

Balakrishnan, Nandhakumar; Cherry, Natalie A; Linder, Keith E; Pierce, Eric; Sontakke, Neal; Hegarty, Barbara C; Bradley, Julie M; Maggi, Ricardo G; Breitschwerdt, Edward B

2013-11-15

The lack of a suitable infection model remains an important obstacle for the pathophysiological understanding of Bartonella spp. The following pilot study was designed to determine whether cell culture-grown Bartonella henselae SA2 and Bartonella vinsonii subsp. berkhoffii genotype III would cause persistent bacteremia in dogs. Pre-inoculation screening established that two laboratory-raised Golden retrievers were naturally-infected with Bartonella koehlerae. Despite prior infection, one dog each was inoculated subcutaneously with 5 × 10(4)B. henselae (SA2 strain) or 3 × 10(4)B. vinsonii subsp. berkhoffii genotype III. Dogs were bled weekly for serological testing and culture using Bartonella alpha proteobacteria growth medium (BAPGM) diagnostic platform. Dog 1 seroconverted to B. henselae and Dog 2 seroconverted to B. vinsonii subsp. berkhoffii genotype III. Throughout the study period, Bartonella spp. DNA was neither amplified nor isolated in ante-mortem BAPGM enrichment blood cultures. B. henselae SA2 was isolated from a postmortem bone marrow from Dog 1 and B. koehlerae DNA was amplified from postmortem lung from Dog 2 following BAPGM enrichment culture. Limitations include lack of uninfected controls, a potentially suboptimal B. vinsonii subsp. berkhoffii inoculum and a relatively short duration of study. We conclude that following intradermal infection, sequestration of Bartonella spp. in tissues may limit diagnostic detection of these bacteria in dog blood samples. Copyright © 2013 Elsevier B.V. All rights reserved.

Specific Detection of Clavibacter michiganensis subsp. sepedonicus by Amplification of Three Unique DNA Sequences Isolated by Subtraction Hybridization.

PubMed

Mills, D; Russell, B W; Hanus, J W

1997-08-01

ABSTRACT Three single-copy, unique DNA fragments, designated Cms50, Cms72, and Cms85, were isolated from strain CS3 of Clavibacter michiganensis subsp. sepedonicus by subtraction hybridization using driver DNA from C. michiganensis subsp. insidiosus, C. michiganensis subsp. michiganensis, and Rhodococcus facians. Radio-labeled probes made of these fragments and used in Southern blot analysis revealed each to be absolutely specific to all North American C. michiganensis subsp. sepedonicus strains tested, including plasmidless and nonmucoid strains. The probes have no homology with genomic DNA from related C. michiganensis subspecies insidiosus, michiganensis, and tessellarius, nor with DNA from 11 additional bacterial species and three unidentified strains, some of which have been previously reported to display cross-reactivity with C. michiganensis subsp. sepedonicus-specific antisera. The three fragments shared no homology, and they appeared to be separated from each other by at least 20 kbp in the CS3 genome. Internal primer sets permitted amplification of each fragment by the polymerase chain reaction (PCR) only from C. michiganensis subsp. sepedonicus DNA. In a PCR-based sensitivity assay using a primer set that amplifies Cms85, the lowest level of detection of C. michiganensis subsp. sepedonicus was 100 CFU per milliliter when cells were added to potato core fluid. Erroneous results that may arise from PCR artifacts and mutational events are, therefore, minimized by the redundancy of the primer sets, and the products should be verifiable with unique capture probes in sequence-based detection systems.

Two efficient nitrite-reducing Lactobacillus strains isolated from traditional fermented pork (Nanx Wudl) as competitive starter cultures for Chinese fermented dry sausage.

PubMed

Chen, Xi; Li, Jiapeng; Zhou, Tong; Li, Jinchun; Yang, Junna; Chen, Wenhua; Xiong, Youling L

2016-11-01

Lactic acid bacteria isolated from traditional Dong pork product (Nanx Wudl) were investigated for their potential as starter cultures for Chinese fermented dry sausages. Based on preliminary screening, Lactobacillus plantarum CMRC6 and Lactobacillus sakei CMRC15, both showing excellent nitrite-reducing capacity, were used as single-strain starter cultures. For comparison, a commercial composite starter was also tested. In CMRC6 and CMRC15-inoculated sausages, lactic acid bacteria dominated the microflora and improved the microbiological safety by suppression of Enterobacteriaceae growth. Nitrite content of all inoculated sausages declined rapidly during ripening compared to non-inoculated. Texture profiles analysis showed inoculated sausages had more pronounced textural development during ripening. Sensory evaluation indicated CMRC6 and CMRC15-fermented sausages had comparable or more desirable organoleptic characteristics than sausage made with commercial starters. Therefore, CMRC6 and CMRC15 are promising candidates as multi-functional starter cultures for microbiological safety and residual nitrite control in gourmet Chinese dry sausage production. Copyright © 2016 Elsevier Ltd. All rights reserved.

Lactobacillus and Leuconostoc volatilomes in cheese conditions.

PubMed

Pogačić, Tomislav; Maillard, Marie-Bernadette; Leclerc, Aurélie; Hervé, Christophe; Chuat, Victoria; Valence, Florence; Thierry, Anne

2016-03-01

New strains are desirable to diversify flavour of fermented dairy products. The objective of this study was to evaluate the potential of Leuconostoc spp. and Lactobacillus spp. in the production of aroma compounds by metabolic fingerprints of volatiles. Eighteen strains, including five Lactobacillus species (Lactobacillus fermentum, Lactobacillus helveticus, Lactobacillus paracasei, Lactobacillus rhamnosus, Lactobacillus sakei) and three Leuconostoc species (Leuconostoc citreum, Leuconostoc lactis, and Leuconostoc mesenteroides) were incubated for 5 weeks in a curd-based slurry medium under conditions mimicking cheese ripening. Populations were enumerated and volatile compounds were analysed by headspace trap gas chromatography-mass spectrometry (GC-MS). A metabolomics approach followed by multivariate statistical analysis was applied for data processing and analysis. In total, 12 alcohols, 10 aldehydes, 7 esters, 11 ketones, 5 acids and 2 sulphur compounds were identified. Very large differences in concentration of volatile compounds between the highest producing strains and the control medium were observed in particular for diacetyl, 2-butanol, ethyl acetate, 3-methylbutanol, 3-methylbutanoic acid and 2-methylbutanoic acid. Some of the characterized strains demonstrated an interesting aromatizing potential to be used as adjunct culture.

Valorization of indigenous dairy cattle breed through salami production.

PubMed

Gaglio, Raimondo; Francesca, Nicola; Maniaci, Giuseppe; Corona, Onofrio; Alfonzo, Antonio; Giosuè, Cristina; Di Noto, Annamaria; Cardamone, Cinzia; Sardina, Maria Teresa; Portolano, Baldassare; Alabiso, Marco

2016-04-01

The aim of the research was to produce salami manufactured with meat of three different commercial categories of bovine breed: cow on retirement, beef and young bull. A total of six experimental productions, at small-scale plant, were carried out with and without starter culture inoculums. The evolution of physico-chemical parameters in all trials followed the trend already registered for other fermented meat products. Several LAB species were found during process with different levels of species diversity and frequency of isolation among inoculated (mainly Pediococcus pentosaceus and Staphylococcus xylosus) and uninoculated (mainly Enterococcus devriesei, Lactobacillus curvatus and Lactobacillus sakei) trials. Enterobacteriaceae were found at very low levels during the entire ripening period and no pathogenic bacteria were found in any samples. The multivariate analysis showed that starter inoculums and meat affected significantly the physico-chemical and the microbiological composition of salami. The sensory analysis evidenced the highest overall acceptability was displayed by salami produced with meat from cow on retirement. Copyright © 2015 Elsevier Ltd. All rights reserved.

Purification and characterization of multiple bacteriocins and an inducing peptide produced by Enterococcus faecium NKR-5-3 from Thai fermented fish.

PubMed

Ishibashi, Naoki; Himeno, Kohei; Fujita, Koji; Masuda, Yoshimitsu; Perez, Rodney Honrada; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2012-01-01

Enterocins NKR-5-3A, B, C, and D were purified from the culture supernatant of Enterococcus faecium NKR-5-3 and characterized. Among the four purified peptides, enterocin NKR-5-3A (5242.3 Da) was identical to brochocin A, produced by Brochothrix campestris ATCC 43754, in mature peptides, and its putative synergistic peptide, enterocin NKR-5-3Z, was found to be encoded in ent53Z downstream of ent53A, encoding enterocin NKR-5-3A. Enterocin NKR-5-3B (6316.4 Da) showed a broad antimicrobial spectrum, and enterocin NKR-5-3C (4512.8 Da) showed high activity against Listeria. Enterocin NKR-5-3D (2843.5 Da), showing high homology to an inducing peptide produced by Lactobacillus sakei 5, induced the production of the enterocins. The enterocins showed different antimicrobial spectra and intensities. E. faecium NKR-5-3 concomitantly produced enterocins NKR-5-3A, B, C, and D which probably belong to different classes of bacteriocins. Furthermore, NKR-5-3 production was induced by enterocin NKR-5-3D.

Comparison of the microbial dynamics and biochemistry of laboratory sourdoughs prepared with grape, apple and yogurt.

PubMed

Gordún, Elena; del Valle, Luis J; Ginovart, Marta; Carbó, Rosa

2015-09-01

The microbiological culture-dependent characterization and physicochemical characteristics of laboratory sourdough prepared with grape (GS) were evaluated and compared with apple (AS) and yogurt (YS), which are the usual Spanish sourdough ingredients. Ripe GS took longer than AS and YS to reach the appropriate acidity and achieved lower values of lactic acid. In all sourdoughs, the lactic acid bacteria (LAB) increased during processing and were the dominant microorganisms (>1E+8 CFU/g). GS, as well as AS, had high diversity of LAB species. In ripe YS, Pediococcus pentosaceus was the only species identified; in GS and AS, several Lactobacilli were also found, Lb. plantarum, Lb. brevis, and Lb. sakei; in addition, in GS Weisella cibaria also appeared. Regarding the yeast population, non-Saccharomyces yeasts from GS and AS showed a very high specific population (>1E+7 CFU/g), but this was reduced in ripe sourdough (<1E+4 CFU/g). Finally, the Saccharomyces group dominated in all sourdoughs. Starting ingredients or raw material provided microbiological specificity to sourdoughs, and grape could be considered one of them. © The Author(s) 2014.

Salmonella enterica suppresses Pectobacterium carotovorum subsp. carotovorum population and soft rot progression by acidifying the microaerophilic environment.

PubMed

Kwan, Grace; Charkowski, Amy O; Barak, Jeri D

2013-02-12

Although enteric human pathogens are usually studied in the context of their animal hosts, a significant portion of their life cycle occurs on plants. Plant disease alters the phyllosphere, leading to enhanced growth of human pathogens; however, the impact of human pathogens on phytopathogen biology and plant health is largely unknown. To characterize the interaction between human pathogens and phytobacterial pathogens in the phyllosphere, we examined the interactions between Pectobacterium carotovorum subsp. carotovorum and Salmonella enterica or Escherichia coli O157:H7 with regard to bacterial populations, soft rot progression, and changes in local pH. The presence of P. carotovorum subsp. carotovorum enhanced the growth of both S. enterica and E. coli O157:H7 on leaves. However, in a microaerophilic environment, S. enterica reduced P. carotovorum subsp. carotovorum populations and soft rot progression by moderating local environmental pH. Reduced soft rot was not due to S. enterica proteolytic activity. Limitations on P. carotovorum subsp. carotovorum growth, disease progression, and pH elevation were not observed on leaves coinoculated with E. coli O157:H7 or when leaves were coinoculated with S. enterica in an aerobic environment. S. enterica also severely undermined the relationship between the phytobacterial population and disease progression of a P. carotovorum subsp. carotovorum budB mutant defective in the 2,3-butanediol pathway for acid neutralization. Our results show that S. enterica and E. coli O157:H7 interact differently with the enteric phytobacterial pathogen P. carotovorum subsp. carotovorum. S. enterica inhibition of soft rot progression may conceal a rapidly growing human pathogen population. Whereas soft rotted produce can alert consumers to the possibility of food-borne pathogens, healthy-looking produce may entice consumption of contaminated vegetables. Salmonella enterica and Escherichia coli O157:H7 may use plants to move between animal

Development of a pentaplex PCR assay for the simultaneous detection of Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis, L. helveticus, L. fermentum in whey starter for Grana Padano cheese.

PubMed

Cremonesi, Paola; Vanoni, Laura; Morandi, Stefano; Silvetti, Tiziana; Castiglioni, Bianca; Brasca, Milena

2011-03-30

A pentaplex PCR assay for the rapid, selective and simultaneous detection of Lactobacillus helveticus, L. delbrueckii subsp. lactis, L. delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and L. fermentum, was developed. The target sequences were a group of genes coding for beta-galactosidase production (S. thermophilus and L. delbrueckii subsp. bulgaricus), for cell-enveloped associated proteinase synthesis (L. helveticus), for dipeptide transport system production (L. delbrueckii subsp. lactis) and for arginine-ornithine antiporter protein production (L. fermentum). The analytical specificity of the assay was evaluated with 5 reference strains and 140 lactic acid bacterial strains derived from raw milk cheeses and belonging to the Lactobacillus, Streptococcus, Lactococcus and Enterococcus genera. The identification limit for each target strain was 10(3)CFU/ml. This new molecular assay was used to investigate the LAB population by direct extraction of DNA from the 12 whey cultures for Grana Padano. The pentaplex PCR assay revealed a good correspondence with microbiological analyses and allowed to identify even minor LAB community members which, can be out-competed in vitro by numerically more abundant microbial species. Copyright © 2011 Elsevier B.V. All rights reserved.

Antibacterial activities of naturally occurring compounds against Mycobacterium avium subsp. paratuberculosis.

PubMed

Wong, Stella Y Y; Grant, Irene R; Friedman, Mendel; Elliott, Christopher T; Situ, Chen

2008-10-01

The antibacterial activities of 18 naturally occurring compounds (including essential oils and some of their isolated constituents, apple and green tea polyphenols, and other plant extracts) against three strains of Mycobacterium avium subsp. paratuberculosis (a bovine isolate [NCTC 8578], a raw-milk isolate [806R], and a human isolate [ATCC 43015]) were evaluated using a macrobroth susceptibility testing method. M. avium subsp. paratuberculosis was grown in 4 ml Middlebrook 7H9 broth containing 10% oleic acid-albumin-dextrose-catalase, 0.05% Tween 80 (or 0.2% glycerol), and 2 microg/ml mycobactin J supplemented with five concentrations of each test compound. The changes in the optical densities of the cultures at 600 nm as a measure of CFU were recorded at intervals over an incubation period of 42 days at 37 degrees C. Six of the compounds were found to inhibit the growth of M. avium subsp. paratuberculosis. The most effective compound was trans-cinnamaldehyde, with a MIC of 25.9 microg/ml, followed by cinnamon oil (26.2 microg/ml), oregano oil (68.2 microg/ml), carvacrol (72.2 microg/ml), 2,5-dihydroxybenzaldehyde (74 microg/ml), and 2-hydroxy-5-methoxybenzaldehyde (90.4 microg/ml). With the exception of carvacrol, a phenolic compound, three of the four most active compounds are aldehydes, suggesting that the structure of the phenolic group or the aldehyde group may be important to the antibacterial activity. No difference in compound activity was observed between the three M. avium subsp. paratuberculosis strains studied. Possible mechanisms of the antimicrobial effects are discussed.

Common genomic features of Campylobacter jejuni subsp. doylei strains distinguish them from C. jejuni subsp. jejuni

PubMed Central

Parker, Craig T; Miller, William G; Horn, Sharon T; Lastovica, Albert J

2007-01-01

Background Campylobacter jejuni has been divided into two subspecies: C. jejuni subsp. jejuni (Cjj) and C. jejuni subsp. doylei (Cjd). Nearly all of the C. jejuni strains isolated are Cjj; nevertheless, although Cjd strains are isolated infrequently, they differ from Cjj in two key aspects: they are obtained primarily from human clinical samples and are associated often with bacteremia, in addition to gastroenteritis. In this study, we utilized multilocus sequence typing (MLST) and a DNA microarray-based comparative genomic indexing (CGI) approach to examine the genomic diversity and gene content of Cjd strains. Results A geographically diverse collection of eight Cjd strains was examined by MLST and determined to be phylogenetically distinct from Cjj strains. Microarray-based CGI approach also supported this. We were able to demonstrate that Cjd strains exhibited divergence from Cjj strains NCTC 11168 and RM1221 in many of the intraspecies hypervariable regions. Moreover, multiple metabolic, transport and virulence functions (e.g. cytolethal distending toxin) were shown to be absent in the Cjd strains examined. Conclusion Our data demonstrate that Cjd are phylogenetically distinct from Cjj strains. Using the CGI approach, we identified subsets of absent genes from amongst the C. jejuni genes that provide clues as to the potential evolutionary origin and unusual pathogenicity of Cjd. PMID:17535437

Distribution of Bacillus thuringiensis subsp. israelensis in Soil of a Swiss Wetland Reserve after 22 Years of Mosquito Control▿†

PubMed Central

Guidi, Valeria; Patocchi, Nicola; Lüthy, Peter; Tonolla, Mauro

2011-01-01

Recurrent treatments with Bacillus thuringiensis subsp. israelensis are required to control the floodwater mosquito Aedes vexans that breeds in large numbers in the wetlands of the Bolle di Magadino Reserve in Canton Ticino, Switzerland. Interventions have been carried out since 1988. In the present study, the spatial distribution of resting B. thuringiensis subsp. israelensis spores in the soil was measured. The B. thuringiensis subsp. israelensis concentration was determined in soil samples collected along six transects covering different elevations within the periodically flooded zones. A total of 258 samples were processed and analyzed by quantitative PCR that targeted an identical fragment of 159 bp for the B. thuringiensis subsp. israelensis cry4Aa and cry4Ba genes. B. thuringiensis subsp. israelensis spores were found to persist in soils of the wetland reserve at concentrations of up to 6.8 log per gram of soil. Continuous accumulation due to regular treatments could be excluded, as the decrease in spores amounted to 95.8% (95% confidence interval, 93.9 to 97.7%). The distribution of spores was correlated to the number of B. thuringiensis subsp. israelensis treatments, the elevation of the sampling point, and the duration of the flooding periods. The number of B. thuringiensis subsp. israelensis treatments was the major factor influencing the distribution of spores in the different topographic zones (P < 0.0001). These findings indicated that B. thuringiensis subsp. israelensis spores are rather immobile after their introduction into the environment. PMID:21498758

Detection and Verification of Mycobacterium avium subsp. paratuberculosis in Fresh Ileocolonic Mucosal Biopsy Specimens from Individuals with and without Crohn's Disease

PubMed Central

Bull, Tim J.; McMinn, Elizabeth J.; Sidi-Boumedine, Karim; Skull, Angela; Durkin, Damien; Neild, Penny; Rhodes, Glenn; Pickup, Roger; Hermon-Taylor, John

2003-01-01

Mycobacterium avium subsp. paratuberculosis is a robust and phenotypically versatile pathogen which causes chronic inflammation of the intestine in many species, including primates. M. avium subsp. paratuberculosis infection is widespread in domestic livestock and is present in retail pasteurized cows' milk in the United Kingdom and, potentially, elsewhere. Water supplies are also at risk. The involvement of M. avium subsp. paratuberculosis in Crohn's disease (CD) in humans has been uncertain because of the substantial difficulties in detecting this pathogen. In its Ziehl-Neelsen staining-negative form, M. avium subsp. paratuberculosis is highly resistant to chemical and enzymatic lysis. The present study describes the development of optimized sample processing and DNA extraction procedures with fresh human intestinal mucosal biopsy specimens which ensure access to M. avium subsp. paratuberculosis DNA and maximize detection of these low-abundance pathogens. Also described are two nested PCR methodologies targeted at IS900, designated IS900[L/AV] and IS900[TJ1-4], which are uniquely specific for IS900. Detection of M. avium subsp. paratuberculosis in mucosal biopsy specimens was also evaluated by using mycobacterial growth indicator tube (MGIT) cultures (Becton Dickinson). IS900[L/AV] PCR detected M. avium subsp. paratuberculosis in 34 of 37 (92%) patients with CD and in 9 of 34 (26%) controls without CD (noninflammatory bowel disease [nIBD] controls) (P = 0.0002; odds ratio = 3.47). M. avium subsp. paratuberculosis was detected by IS900[L/AV] PCR in MGIT cultures after 14 to 88 weeks of incubation in 14 of 33 (42%) CD patients and 3 of 33 (9%) nIBD controls (P = 0.0019; odds ratio = 4.66). Nine of 15 (60%) MGIT cultures of specimens from CD patients incubated for more than 38 weeks were positive for M. avium subsp. paratuberculosis. In each case the identity of IS900 from M. avium subsp. paratuberculosis was verified by amplicon sequencing. The rate of detection of

Electrotransformation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis with Various Plasmids

PubMed Central

Serror, Pascale; Sasaki, Takashi; Ehrlich, S. Dusko; Maguin, Emmanuelle

2002-01-01

We describe, for the first time, a detailed electroporation procedure for Lactobacillus delbrueckii. Three L. delbrueckii strains were successfully transformed. Under optimal conditions, the transformation efficiency was 104 transformants per μg of DNA. Using this procedure, we identified several plasmids able to replicate in L. delbrueckii and integrated an integrative vector based on phage integrative elements into the L. delbrueckii subsp. bulgaricus chromosome. These vectors provide a good basis for developing molecular tools for L. delbrueckii and open the field of genetic studies in L. delbrueckii. PMID:11772607

Fate of Mycobacterium avium subsp. paratuberculosis in Swiss hard and semihard cheese manufactured from raw milk.

PubMed

Spahr, U; Schafroth, K

2001-09-01

Raw milk was artificially contaminated with declumped cells of Mycobacterium avium subsp. paratuberculosis at a concentration of 10(4) to 10(5) CFU/ml and was used to manufacture model hard (Swiss Emmentaler) and semihard (Swiss Tisliter) cheese. Two different strains of M. avium subsp. paratuberculosis were tested, and for each strain, two model hard and semihard cheeses were produced. The survival of M. avium subsp. paratuberculosis cells was monitored over a ripening period of 120 days by plating out homogenized cheese samples onto 7H10-PANTA agar. In both the hard and the semihard cheeses, counts decreased steadily but slowly during cheese ripening. Nevertheless, viable cells could still be detected in 120-day cheese. D values were calculated at 27.8 days for hard and 45.5 days for semihard cheese. The most important factors responsible for the death of M. avium subsp. paratuberculosis in cheese were the temperatures applied during cheese manufacture and the low pH at the early stages of cheese ripening. Since the ripening period for these raw milk cheeses lasts at least 90 to 120 days, the D values found indicate that 10(3) to 10(4) cells of M. avium subsp. paratuberculosis per g will be inactivated.

Fate of Mycobacterium avium subsp. paratuberculosis in Swiss Hard and Semihard Cheese Manufactured from Raw Milk

PubMed Central

Spahr, U.; Schafroth, K.

2001-01-01

Raw milk was artificially contaminated with declumped cells of Mycobacterium avium subsp. paratuberculosis at a concentration of 104 to 105 CFU/ml and was used to manufacture model hard (Swiss Emmentaler) and semihard (Swiss Tisliter) cheese. Two different strains of M. avium subsp. paratuberculosis were tested, and for each strain, two model hard and semihard cheeses were produced. The survival of M. avium subsp. paratuberculosis cells was monitored over a ripening period of 120 days by plating out homogenized cheese samples onto 7H10-PANTA agar. In both the hard and the semihard cheeses, counts decreased steadily but slowly during cheese ripening. Nevertheless, viable cells could still be detected in 120-day cheese. D values were calculated at 27.8 days for hard and 45.5 days for semihard cheese. The most important factors responsible for the death of M. avium subsp. paratuberculosis in cheese were the temperatures applied during cheese manufacture and the low pH at the early stages of cheese ripening. Since the ripening period for these raw milk cheeses lasts at least 90 to 120 days, the D values found indicate that 103 to 104 cells of M. avium subsp. paratuberculosis per g will be inactivated. PMID:11526024

Culture Phenotypes of Genomically and Geographically Diverse Mycobacterium avium subsp. paratuberculosis Isolates from Different Hosts▿

PubMed Central

Whittington, Richard J.; Marsh, Ian B.; Saunders, Vanessa; Grant, Irene R.; Juste, Ramon; Sevilla, Iker A.; Manning, Elizabeth J. B.; Whitlock, Robert H.

2011-01-01

Mycobacterium avium subsp. paratuberculosis causes paratuberculosis (Johne's disease) in ruminants in most countries. Historical data suggest substantial differences in culturability of M. avium subsp. paratuberculosis isolates from small ruminants and cattle; however, a systematic comparison of culture media and isolates from different countries and hosts has not been undertaken. Here, 35 field isolates from the United States, Spain, Northern Ireland, and Australia were propagated in Bactec 12B medium and Middlebrook 7H10 agar, genomically characterized, and subcultured to Lowenstein-Jensen (LJ), Herrold's egg yolk (HEY), modified Middlebrook 7H10, Middlebrook 7H11, and Watson-Reid (WR) agars, all with and without mycobactin J and some with sodium pyruvate. Fourteen genotypes of M. avium subsp. paratuberculosis were represented as determined by BstEII IS900 and IS1311 restriction fragment length polymorphism analysis. There was no correlation between genotype and overall culturability, although most S strains tended to grow poorly on HEY agar. Pyruvate was inhibitory to some isolates. All strains grew on modified Middlebrook 7H10 agar but more slowly and less prolifically on LJ agar. Mycobactin J was required for growth on all media except 7H11 agar, but growth was improved by the addition of mycobactin J to 7H11 agar. WR agar supported the growth of few isolates. The differences in growth of M. avium subsp. paratuberculosis that have historically been reported in diverse settings have been strongly influenced by the type of culture medium used. When an optimal culture medium, such as modified Middlebrook 7H10 agar, is used, very little difference between the growth phenotypes of diverse strains of M. avium subsp. paratuberculosis was observed. This optimal medium is recommended to remove bias in the isolation and cultivation of M. avium subsp. paratuberculosis. PMID:21430104

Faecal bacterial composition in dairy cows shedding Mycobacterium avium subsp. paratuberculosis in faeces in comparison with nonshedding cows.

PubMed

Kaevska, Marija; Videnska, Petra; Sedlar, Karel; Bartejsova, Iva; Kralova, Alena; Slana, Iva

2016-06-01

The aim of this study was to determine possible differences in the faecal microbiota of dairy cows infected with Mycobacterium avium subsp. paratuberculosis (Johne's disease) in comparison with noninfected cows from the same herds. Faecal samples from cows in 4 herds were tested for M. avium subsp. paratuberculosis by real-time PCR, and faecal bacterial populations were analysed by 454 pyrosequencing of the 16S rRNA gene. The most notable differences between shedding and nonshedding cows were an increase in the genus Psychrobacter and a decrease in the genera Oscillospira, Ruminococcus, and Bifidobacterium in cows infected with M. avium subsp. paratuberculosis. The present study is the first to report the faecal microbial composition in dairy cows infected with M. avium subsp. paratuberculosis.

Microbiota of Minas cheese as influenced by the nisin producer Lactococcus lactis subsp. lactis GLc05.

PubMed

Perin, Luana Martins; Dal Bello, Barbara; Belviso, Simona; Zeppa, Giuseppe; Carvalho, Antônio Fernandes de; Cocolin, Luca; Nero, Luís Augusto

2015-12-02

Minas cheese is a popular dairy product in Brazil that is traditionally produced using raw or pasteurized cow milk. This study proposed an alternative production of Minas cheese using raw goat milk added of a nisin producer Lactococcus lactis subsp. lactis GLc05. An in situ investigation was carried on to evaluate the interactions between the L. lactis subsp. lactis GLc05 and the autochthonous microbiota of a Minas cheese during the ripening; production of biogenic amines (BAs) was assessed as a safety aspect. Minas cheese was produced in two treatments (A, by adding L. lactis subsp. lactis GLc05, and B, without adding this strain), in three independent repetitions (R1, R2, and R3). Culture dependent (direct plating) and independent (rep-PCR and PCR-DGGE) methods were employed to characterize the microbiota and to assess the possible interferences caused by L. lactis subsp. lactis GLc05. BA amounts were measured using HPLC. A significant decrease in coagulase-positive cocci was observed in the cheeses produced by adding L. lactis subsp. lactis GLc05 (cheese A). The rep-PCR and PCR-DGGE highlighted the differences in the microbiota of both cheeses, separating them into two different clusters. Lactococcus sp. was found as the main microorganism in both cheeses, and the microbiota of cheese A presented a higher number of species. High concentrations of tyramine were found in both cheeses and, at specific ripening times, the BA amounts in cheese B were significantly higher than in cheese A (p<0.05). The interaction of nisin producer L. lactis subsp. lactis GLc05 was demonstrated in situ, by demonstration of its influence in the complex microbiota naturally present in a raw goat milk cheese and by controlling the growth of coagulase-positive cocci. L. lactis subsp. lactis GLc05 influenced also the production of BA determining that their amounts in the cheeses were maintained at acceptable levels for human consumption. Copyright © 2015 Elsevier B.V. All rights reserved.

Antibacterial Activities of Naturally Occurring Compounds against Mycobacterium avium subsp. paratuberculosis▿

PubMed Central

Wong, Stella Y. Y.; Grant, Irene R.; Friedman, Mendel; Elliott, Christopher T.; Situ, Chen

2008-01-01

The antibacterial activities of 18 naturally occurring compounds (including essential oils and some of their isolated constituents, apple and green tea polyphenols, and other plant extracts) against three strains of Mycobacterium avium subsp. paratuberculosis (a bovine isolate [NCTC 8578], a raw-milk isolate [806R], and a human isolate [ATCC 43015]) were evaluated using a macrobroth susceptibility testing method. M. avium subsp. paratuberculosis was grown in 4 ml Middlebrook 7H9 broth containing 10% oleic acid-albumin-dextrose-catalase, 0.05% Tween 80 (or 0.2% glycerol), and 2 μg/ml mycobactin J supplemented with five concentrations of each test compound. The changes in the optical densities of the cultures at 600 nm as a measure of CFU were recorded at intervals over an incubation period of 42 days at 37°C. Six of the compounds were found to inhibit the growth of M. avium subsp. paratuberculosis. The most effective compound was trans-cinnamaldehyde, with a MIC of 25.9 μg/ml, followed by cinnamon oil (26.2 μg/ml), oregano oil (68.2 μg/ml), carvacrol (72.2 μg/ml), 2,5-dihydroxybenzaldehyde (74 μg/ml), and 2-hydroxy-5-methoxybenzaldehyde (90.4 μg/ml). With the exception of carvacrol, a phenolic compound, three of the four most active compounds are aldehydes, suggesting that the structure of the phenolic group or the aldehyde group may be important to the antibacterial activity. No difference in compound activity was observed between the three M. avium subsp. paratuberculosis strains studied. Possible mechanisms of the antimicrobial effects are discussed. PMID:18676709

Effect of Soil Slope on the Appearance of Mycobacterium avium subsp. paratuberculosis in Water Running off Grassland Soil after Application of Contaminated Slurry

PubMed Central

Alfaro, M.; Salazar, F.; Troncoso, E.; Mitchell, R. M.; Ramirez, L.; Naguil, A.; Zamorano, P.; Collins, M. T.

2013-01-01

The study assessed the effect of soil slope on Mycobacterium avium subsp. paratuberculosis transport into rainwater runoff from agricultural soil after application of M. avium subsp. paratuberculosis-contaminated slurry. Under field conditions, 24 plots of undisturbed loamy soil 1 by 2 m2 were placed on platforms. Twelve plots were used for water runoff: 6 plots at a 3% slope and 6 plots at a 15% slope. Half of the plots of each slope were treated with M. avium subsp. paratuberculosis-contaminated slurry, and half were not treated. Using the same experimental design, 12 plots were established for soil sampling on a monthly basis using the same spiked slurry application and soil slopes. Runoff following natural rainfall was collected and analyzed for M. avium subsp. paratuberculosis, coliforms, and turbidity. M. avium subsp. paratuberculosis was detected in runoff from all plots treated with contaminated slurry and one control plot. A higher slope (15%) increased the likelihood of M. avium subsp. paratuberculosis detection but did not affect the likelihood of finding coliforms. Daily rainfall increased the likelihood that runoff would have coliforms and the coliform concentration, but it decreased the M. avium subsp. paratuberculosis concentration in the runoff. When there was no runoff, rain was associated with increased M. avium subsp. paratuberculosis concentrations. Coliform counts in runoff were related to runoff turbidity. M. avium subsp. paratuberculosis presence/absence, however, was related to turbidity. Study duration decreased bacterial detection and concentration. These findings demonstrate the high likelihood that M. avium subsp. paratuberculosis in slurry spread on pastures will contaminate water runoff, particularly during seasons with high rainfall. M. avium subsp. paratuberculosis contamination of water has potential consequences for both animal and human health. PMID:23542616

Complete genome sequence of Clavibacter michiganensis subsp. insidiosus

USDA-ARS?s Scientific Manuscript database

Clavibacter michiganensis subsp. insidiosus (Cmi) causes bacterial wilt disease of alfalfa (Medicago sativa L.) and can also infect the model legume plant M. truncatula. The virulence mechanisms of Cmi are yet to be identified, hampered by the lack of efficient mutagenesis tools as well as by the la...

Unusual Outbreak of Clinical Mastitis in Dairy Sheep Caused by Streptococcus equi subsp. zooepidemicus

PubMed Central

Las Heras, Alfonso; Vela, Ana I.; Fernández, Elena; Legaz, Emilio; Domínguez, Lucas; Fernández-Garayzábal, Jose F.

2002-01-01

This work describes an outbreak of clinical mastitis affecting 13 of 58 lactating ewes due to Streptococcus equi subsp. zooepidemicus. S. equi subsp. zooepidemicus was isolated in pure culture from all milk samples. All the clinical isolates had identical biochemical profiles and antimicrobial susceptibility patterns and also exhibited indistinguishable macrorestriction patterns by pulsed-field gel electrophoresis, indicating that all cases of mastitis were produced by a single strain. PMID:11880454

Human Treponema pallidum 11q/j isolate belongs to subsp. endemicum but contains two loci with a sequence in TP0548 and TP0488 similar to subsp. pertenue and subsp. pallidum, respectively

PubMed Central

Mikalová, Lenka; Strouhal, Michal; Oppelt, Jan; Grange, Philippe Alain; Janier, Michel; Benhaddou, Nadjet; Dupin, Nicolas; Šmajs, David

2017-01-01

Background Treponema pallidum subsp. endemicum (TEN) is the causative agent of endemic syphilis (bejel). An unusual human TEN 11q/j isolate was obtained from a syphilis-like primary genital lesion from a patient that returned to France from Pakistan. Methodology/Principal findings The TEN 11q/j isolate was characterized using nested PCR followed by Sanger sequencing and/or direct Illumina sequencing. Altogether, 44 chromosomal regions were analyzed. Overall, the 11q/j isolate clustered with TEN strains Bosnia A and Iraq B as expected from previous TEN classification of the 11q/j isolate. However, the 11q/j sequence in a 505 bp-long region at the TP0488 locus was similar to Treponema pallidum subsp. pallidum (TPA) strains, but not to TEN Bosnia A and Iraq B sequences, suggesting a recombination event at this locus. Similarly, the 11q/j sequence in a 613 bp-long region at the TP0548 locus was similar to Treponema pallidum subsp. pertenue (TPE) strains, but not to TEN sequences. Conclusions/Significance A detailed analysis of two recombinant loci found in the 11q/j clinical isolate revealed that the recombination event occurred just once, in the TP0488, with the donor sequence originating from a TPA strain. Since TEN Bosnia A and Iraq B were found to contain TPA-like sequences at the TP0548 locus, the recombination at TP0548 took place in a treponeme that was an ancestor to both TEN Bosnia A and Iraq B. The sequence of 11q/j isolate in TP0548 represents an ancestral TEN sequence that is similar to yaws-causing treponemes. In addition to the importance of the 11q/j isolate for reconstruction of the TEN phylogeny, this case emphasizes the possible role of TEN strains in development of syphilis-like lesions. PMID:28263990

Production of Angiotensin-I-Converting-Enzyme-Inhibitory Peptides in Fermented Milks Started by Lactobacillus delbrueckii subsp. bulgaricus SS1 and Lactococcus lactis subsp. cremoris FT4

PubMed Central

Gobbetti, M.; Ferranti, P.; Smacchi, E.; Goffredi, F.; Addeo, F.

2000-01-01

Two fermented milks containing angiotensin-I-converting-enzyme (ACE)-inhibitory peptides were produced by using selected Lactobacillus delbrueckii subsp. bulgaricus SS1 and L. lactis subsp. cremoris FT4. The pH 4.6-soluble nitrogen fraction of the two fermented milks was fractionated by reversed-phase fast-protein liquid chromatography. The fractions which showed the highest ACE-inhibitory indexes were further purified, and the related peptides were sequenced by tandem fast atom bombardment-mass spectrometry. The most inhibitory fractions of the milk fermented by L. delbrueckii subsp. bulgaricus SS1 contained the sequences of β-casein (β-CN) fragment 6-14 (f6-14), f7-14, f73-82, f74-82, and f75-82. Those from the milk fermented by L. lactis subsp. cremoris FT4 contained the sequences of β-CN f7-14, f47-52, and f169-175 and κ-CN f155-160 and f152-160. Most of these sequences had features in common with other ACE-inhibitory peptides reported in the literature. In particular, the β-CN f47-52 sequence had high homology with that of angiotensin-II. Some of these peptides were chemically synthesized. The 50% inhibitory concentrations (IC50s) of the crude purified fractions containing the peptide mixture were very low (8.0 to 11.2 mg/liter). When the synthesized peptides were used individually, the ACE-inhibitory activity was confirmed but the IC50s increased considerably. A strengthened inhibitory effect of the peptide mixtures with respect to the activity of individual peptides was presumed. Once generated, the inhibitory peptides were resistant to further proteolysis either during dairy processing or by trypsin and chymotrypsin. PMID:10966406

tuf Gene Sequence Variation in Bifidobacterium longum subsp. infantis Detected in the Fecal Microbiota of Chinese Infants.

PubMed

Lawley, Blair; Centanni, Manuela; Watanabe, Jun; Sims, Ian; Carnachan, Susan; Broadbent, Roland; Lee, Pheng Soon; Wong, Khai Hong; Tannock, Gerald W

2018-07-01

Members of the bacterial genus Bifidobacterium generally dominate the fecal microbiota of infants. The species Bifidobacterium longum is prevalent, but the B. longum subsp. longum and B. longum subsp. infantis strains that are known to colonize the infant bowel are not usually differentiated in microbiota investigations. These subspecies differ in their capacities to metabolize human milk oligosaccharides (HMO) and may have different ecological and symbiotic roles in humans. Quantitative PCR provides a quick analytical method by which to accurately ascertain the abundances of target species in microbiotas and microcosms. However, amplification targets in DNA extracted from samples need to be dependably differential. We evaluated the tuf gene sequence as a molecular target for quantitative PCR measurements of the abundances of B. longum subsp. infantis and B. longum subsp. longum in fecal microbiotas. This approach resulted in the detection of a tuf gene variant (operational taxonomic unit 49 [OTU49]) in Chinese infants that has sequence similarities to both B. longum subsp. infantis and B. longum subsp. longum We compared the genome sequence and growth and transcriptional characteristics of an OTU49 isolate cultured in HMO medium to those of other B. longum subsp. infantis cultures. We concluded from these studies that OTU49 belongs to B. longum subsp. infantis , that dependable quantitative PCR (qPCR) differentiation between the B. longum subspecies cannot be achieved by targeting tuf gene sequences, and that functional genes involved in carbohydrate metabolism might be better targets because they delineate ecological functions. IMPORTANCE High-throughput DNA sequencing methods and advanced bioinformatics analysis have revealed the composition and biochemical capacities of microbial communities (microbiota and microbiome), including those that inhabit the gut of human infants. However, the microbiology and function of natural ecosystems have received little

High-Throughput Direct Fecal PCR Assay for Detection of Mycobacterium avium subsp. paratuberculosis in Sheep and Cattle

PubMed Central

Waldron, Anna M.; Galea, Francesca; Whittington, Ann-Michele; Saunders, Vanessa F.; Begg, Douglas J.; de Silva, Kumudika; Purdie, Auriol C.; Whittington, Richard J.

2014-01-01

Johne's disease (JD) is a chronic enteric disease caused by Mycobacterium avium subsp. paratuberculosis that affects ruminants. Transmission occurs by the fecal-oral route. A commonly used antemortem diagnostic test for the detection of M. avium subsp. paratuberculosis in feces is liquid culture; however, a major constraint is the 2- to 3-month incubation period needed for this method. Rapid methods for the detection of M. avium subsp. paratuberculosis based on PCR have been reported, but comprehensive validation data are lacking. We describe here a new test, the high-throughput-Johnes (HT-J), to detect M. avium subsp. paratuberculosis in feces. Its diagnostic accuracy was compared with that of liquid radiometric (Bactec) fecal culture using samples from cattle (1,330 samples from 23 herds) and sheep (596 samples from 16 flocks). The multistage protocol involves the recovery of M. avium subsp. paratuberculosis cells from a fecal suspension, cell rupture by bead beating, extraction of DNA using magnetic beads, and IS900 quantitative PCR. The limit of detection of the assay was 0.0005 pg, and the limit of quantification was 0.005 pg M. avium subsp. paratuberculosis genomic DNA. Only M. avium subsp. paratuberculosis was detected from a panel of 51 mycobacterial isolates, including 10 with IS900-like sequences. Of the 549 culture-negative fecal samples from unexposed herds and flocks, 99% were negative in the HT-J test, while 60% of the bovine- and 84% of the ovine-culture-positive samples were positive in the HT-J test. As similar total numbers of samples from M. avium subsp. paratuberculosis-exposed animals were positive in culture and HT-J tests in both species, and as the results of a McNemar's test were not significant, these methods probably have similar sensitivities, but the true diagnostic sensitivities of these tests are unknown. These validation data meet the consensus-based reporting standards for diagnostic test accuracy studies for paratuberculosis and

Lactobacillus delbrueckii subsp. bulgaricus CRL 454 cleaves allergenic peptides of β-lactoglobulin.

PubMed

Pescuma, Micaela; Hébert, Elvira M; Haertlé, Thomas; Chobert, Jean-Marc; Mozzi, Fernanda; Font de Valdez, Graciela

2015-03-01

Whey, a cheese by-product used as a food additive, is produced worldwide at 40.7 million tons per year. β-Lactoglobulin (BLG), the main whey protein, is poorly digested and is highly allergenic. We aimed to study the contribution of Lactobacillus delbrueckii subsp. bulgaricus CRL 454 to BLG digestion and to analyse its ability to degrade the main allergenic sequences of this protein. Pre-hydrolysis of BLG by L. delbrueckii subsp. bulgaricus CRL 454 increases digestion of BLG assayed by an in vitro simulated gastrointestinal system. Moreover, peptides from hydrolysis of the allergenic sequences V41-K60, Y102-R124, C121-L140 and L149-I162 were found when BLG was hydrolysed by this strain. Interestingly, peptides possessing antioxidant, ACE inhibitory, antimicrobial and immuno-modulating properties were found in BLG degraded by both the Lactobacillus strain and digestive enzymes. To conclude, pre-hydrolysis of BLG by L. delbrueckii subsp. bulgaricus CRL 454 has a positive effect on BLG digestion and could diminish allergenic reactions. Copyright © 2014 Elsevier Ltd. All rights reserved.

A new methodology for rapid detection of Lactobacillus delbrueckii subsp. bulgaricus based on multiplex PCR.

PubMed

Nikolaou, Anastasios; Saxami, Georgia; Kourkoutas, Yiannis; Galanis, Alex

2011-02-01

In this study we present a novel multiplex PCR assay for rapid and efficient detection of Lactobacillus delbrueckii subsp. bulgaricus. The accuracy of our method was confirmed by the successful identification of L. delbrueckii subsp. bulgaricus in commercial yoghurts and food supplements and it may be readily applied to the food industry. Copyright © 2010 Elsevier B.V. All rights reserved.

Chemical decontamination with N-acetyl-L-cysteine-sodium hydroxide improves recovery of viable Mycobacterium avium subsp. paratuberculosis organisms from cultured milk.

PubMed

Bradner, L; Robbe-Austerman, S; Beitz, D C; Stabel, J R

2013-07-01

Mycobacterium avium subsp. paratuberculosis is shed into the milk and feces of cows with advanced Johne's disease, allowing the transmission of M. avium subsp. paratuberculosis between animals. The objective of this study was to formulate an optimized protocol for the isolation of M. avium subsp. paratuberculosis in milk. The parameters investigated included chemical decontamination with N-acetyl-l-cysteine-sodium hydroxide (NALC-NaOH), alone and in combination with antibiotics (vancomycin, amphotericin B, and nalidixic acid), and the efficacy of solid (Herrold's egg yolk medium [HEY]) and liquid (Bactec 12B and para-JEM) culture media. For each experiment, raw milk samples from a known noninfected cow were inoculated with 10(2) to 10(8) CFU/ml of live M. avium subsp. paratuberculosis organisms. The results indicate that an increased length of exposure to NALC-NaOH from 5 to 30 min and an increased concentration of NaOH from 0.5 to 2.0% did not affect the viability of M. avium subsp. paratuberculosis. Additional treatment of milk samples with the antibiotics following NALC-NaOH treatment decreased the recovery of viable M. avium subsp. paratuberculosis cells more than treatment with NALC-NaOH alone. The Bactec 12B medium was the superior medium of the three evaluated for the isolation of M. avium subsp. paratuberculosis from milk, as it achieved the lowest threshold of detection. The optimal conditions for NALC-NaOH decontamination were determined to be exposure to 1.50% NaOH for 15 min followed by culture in Bactec 12B medium. This study demonstrates that chemical decontamination with NALC-NaOH resulted in a greater recovery of viable M. avium subsp. paratuberculosis cells from milk than from samples treated with hexadecylpyridinium chloride (HPC). Therefore, it is important to optimize milk decontamination protocols to ensure that low concentrations of M. avium subsp. paratuberculosis can be detected.

Phenotypic variation in Lactococcus lactis subsp. lactis isolates derived from intestinal tracts of marine and freshwater fish.

PubMed

Itoi, S; Yuasa, K; Washio, S; Abe, T; Ikuno, E; Sugita, H

2009-09-01

We compared phenotypic characteristics of Lactococcus lactis subsp. lactis derived from different sources including the intestinal tract of marine fish and freshwater fish, and cheese starter culture. In the phylogenetic analysis based on partial 16S rRNA gene nucleotide sequences (1371 bp), freshwater fish-, marine fish- and cheese starter culture-derived strains were identical to that of L. lactis subsp. lactis previously reported. Fermentation profiles determined using the API 50 CH system were similar except for fermentation of several sugars including l-arabinose, mannitol, amygdalin, saccharose, trehalose, inulin and gluconate. The strains did have distinct levels of halotolerance: marine fish-derived strains > cheese starter-derived strain > freshwater fish-derived isolate. Lactococcus lactis subsp. lactis showed extensive diversity in phenotypic adaptation to various environments. The phenotypic properties of these strains suggested that L. lactis subsp. lactis strains from fish intestine have additional functions compared with the cheese starter-derived strain that has previously described. The unique phenotypic traits of the fish intestinal tract-derived L. lactis subsp. lactis might make them useful as a probiotics in aquaculture, and contribute to the development of functional foods and novel food additives, since the strains derived from fish intestines might have additional functions such as antibacterial activity.

Persistence of Mycobacterium avium subsp. paratuberculosis at a Farm-Scale Biogas Plant Supplied with Manure from Paratuberculosis-Affected Dairy Cattle▿

PubMed Central

Slana, I.; Pribylova, R.; Kralova, A.; Pavlik, I.

2011-01-01

In this study, products from all steps of anaerobic digestion at a farm-scale biogas plant supplied with manure from paratuberculosis-affected dairy cattle were examined and quantified for the presence of the causal agent of paratuberculosis, Mycobacterium avium subsp. paratuberculosis, using culture and quantitative real-time PCR (qPCR). Viable M. avium subsp. paratuberculosis cells were detected using culture in fermentors for up to 2 months; the presence of M. avium subsp. paratuberculosis DNA (101 cells/g) was demonstrated in all anaerobic fermentors and digestate 16 months after initiation of work at a biogas plant, using IS900 qPCR. F57 qPCR was able to detect M. avium subsp. paratuberculosis DNA (102 cells/g) at up to 12 months. According to these results, a fermentation process that extended beyond 2 months removed all viable M. avium subsp. paratuberculosis cells and therefore rendered its product M. avium subsp. paratuberculosis free. However, M. avium subsp. paratuberculosis DNA was found during all the examined periods (more than 1 year), which could be explained by either residual DNA being released from dead cells or by the presence of viable cells whose amount was under the limit of cultivability. As the latter hypothesis cannot be excluded, the safety of the final products of digestion used for fertilization or animal bedding cannot be defined, and further investigation is necessary to confirm or refute this risk. PMID:21398476

Lactococcus lactis subsp. cremoris strain JFR1 attenuates Salmonella adhesion to human intestinal cells in vitro.

PubMed

Zhang, Justina Su; Guri, Anilda; Corredig, Milena; Morales-Rayas, Rocio; Hassan, Ashraf; Griffiths, Mansel; LaPointe, Gisèle

2016-12-01

Lactococcus lactis subsp. cremoris JFR1 has been studied in reduced fat cheese due to its ability to produce exopolysaccharides (EPS) in situ, contributing to improved textural and organoleptic properties. In this study, the effect of strain JFR1 on virulence gene expression and attachment of Salmonella to HT-29 human colon carcinoma cells was investigated. Overnight cultures of L. lactis subsp. cremoris JFR1 containing EPS, grown in M17 media with 0.5% glucose supplementation, decreased attachment as well as down regulated virulence gene expression in Salmonella enterica subsp. enterica when tested on HT-29 cells. However, EPS isolated from milk fermented with L. lactis subsp. cremoris JFR1 did not affect Salmonella virulence gene expression or attachment to HT-29 cells. These results suggest that EPS does not contribute to the attachment of Salmonella to human intestinal cells. However, the possibility that the isolation process may have affected the structural features of EPS cannot be ruled out. Copyright © 2016 Elsevier Ltd. All rights reserved.

Bifidobacterium animalis subsp. lactis decreases urinary oxalate excretion in a mouse model of primary hyperoxaluria

PubMed Central

Whittamore, Jonathan M.; Hatch, Marguerite

2015-01-01

Hyperoxaluria significantly increases the risk of calcium oxalate kidney stone formation. Since several bacteria have been shown to metabolize oxalate in vitro, including probiotic bifidobacteria, we focused on the efficiency and possible mechanisms by which bifidobacteria can infuence oxalate handling in vivo, especially in the intestines, and compared these results with the reported effects of Oxalobacter formigenes. Bifidobacterium animalis subsp. lactis DSM 10140 and B. adolescentis ATCC 15703 were administered to wild-type (WT) mice and to mice defcient in the hepatic enzyme alanine-glyoxylate aminotransferase (Agxt−/−, a mouse model of Primary Hyperoxaluria) that were fed an oxalate-supplemented diet. The administration of B. animalis subsp. lactis led to a significant decrease in urinary oxalate excretion in WT and Agxt−/− mice when compared to treatment with B. adolescent-is. Detection of B. animalis subsp. lactis in feces revealed that 3 weeks after oral gavage with the bacteria 64 % of WT mice, but only 37 % of Agxt−/− mice were colonized. Examining intestinal oxalate fuxes showed there were no significant changes to net oxalate secretion in colonized animals and were therefore not associated with the changes in urinary oxalate excretion. These results indicate that colonization with B. animalis subsp. lactis decreased urinary oxalate excretion by degrading dietary oxalate thus limiting its absorption across the intestine but it did not promote enteric oxalate excretion as reported for O. formigenes. Preventive or therapeutic administration of B. animalis subsp. lactis appears to have some potential to beneficially infuence dietary hyperoxaluria in mice. PMID:25269440

Efficacy of various pasteurization time-temperature conditions in combination with homogenization on inactivation of Mycobacterium avium subsp. paratuberculosis in milk.

PubMed

Grant, Irene R; Williams, Alan G; Rowe, Michael T; Muir, D Donald

2005-06-01

The effect of various pasteurization time-temperature conditions with and without homogenization on the viability of Mycobacterium avium subsp. paratuberculosis was investigated using a pilot-scale commercial high-temperature, short-time (HTST) pasteurizer and raw milk spiked with 10(1) to 10(5) M. avium subsp. paratuberculosis cells/ml. Viable M. avium subsp. paratuberculosis was cultured from 27 (3.3%) of 816 pasteurized milk samples overall, 5 on Herrold's egg yolk medium and 22 by BACTEC culture. Therefore, in 96.7% of samples, M. avium subsp. paratuberculosis had been completely inactivated by HTST pasteurization, alone or in combination with homogenization. Heat treatments incorporating homogenization at 2,500 lb/in2, applied upstream (as a separate process) or in hold (at the start of a holding section), resulted in significantly fewer culture-positive samples than pasteurization treatments without homogenization (P < 0.001 for those in hold and P < 0.05 for those upstream). Where colony counts were obtained, the number of surviving M. avium subsp. paratuberculosis cells was estimated to be 10 to 20 CFU/150 ml, and the reduction in numbers achieved by HTST pasteurization with or without homogenization was estimated to be 4.0 to 5.2 log10. The impact of homogenization on clump size distribution in M. avium subsp. paratuberculosis broth suspensions was subsequently assessed using a Mastersizer X spectrometer. These experiments demonstrated that large clumps of M. avium subsp. paratuberculosis cells were reduced to single-cell or "miniclump" status by homogenization at 2,500 lb/in2. Consequently, when HTST pasteurization was being applied to homogenized milk, the M. avium subsp. paratuberculosis cells would have been present as predominantly declumped cells, which may possibly explain the greater inactivation achieved by the combination of pasteurization and homogenization.

Efficacy of Various Pasteurization Time-Temperature Conditions in Combination with Homogenization on Inactivation of Mycobacterium avium subsp. paratuberculosis in Milk

PubMed Central

Grant, Irene R.; Williams, Alan G.; Rowe, Michael T.; Muir, D. Donald

2005-01-01

The effect of various pasteurization time-temperature conditions with and without homogenization on the viability of Mycobacterium avium subsp. paratuberculosis was investigated using a pilot-scale commercial high-temperature, short-time (HTST) pasteurizer and raw milk spiked with 101 to 105 M. avium subsp. paratuberculosis cells/ml. Viable M. avium subsp. paratuberculosis was cultured from 27 (3.3%) of 816 pasteurized milk samples overall, 5 on Herrold's egg yolk medium and 22 by BACTEC culture. Therefore, in 96.7% of samples, M. avium subsp. paratuberculosis had been completely inactivated by HTST pasteurization, alone or in combination with homogenization. Heat treatments incorporating homogenization at 2,500 lb/in2, applied upstream (as a separate process) or in hold (at the start of a holding section), resulted in significantly fewer culture-positive samples than pasteurization treatments without homogenization (P < 0.001 for those in hold and P < 0.05 for those upstream). Where colony counts were obtained, the number of surviving M. avium subsp. paratuberculosis cells was estimated to be 10 to 20 CFU/150 ml, and the reduction in numbers achieved by HTST pasteurization with or without homogenization was estimated to be 4.0 to 5.2 log10. The impact of homogenization on clump size distribution in M. avium subsp. paratuberculosis broth suspensions was subsequently assessed using a Mastersizer X spectrometer. These experiments demonstrated that large clumps of M. avium subsp. paratuberculosis cells were reduced to single-cell or “miniclump” status by homogenization at 2,500 lb/in2. Consequently, when HTST pasteurization was being applied to homogenized milk, the M. avium subsp. paratuberculosis cells would have been present as predominantly declumped cells, which may possibly explain the greater inactivation achieved by the combination of pasteurization and homogenization. PMID:15932977

Campylobacter fetus subsp. jejuni in poultry reared under different management systems in Nigeria.

PubMed

Adekeye, J O; Abdu, P A; Bawa, E K

1989-01-01

Cloacal swabs from 487 live birds in 36 flocks and 70 poultry carcasses were cultured for Campylobacter fetus subsp. jejuni. It was isolated from 12.3% of the birds in 19 flocks. Chickens, turkeys, and guinea fowl differed from one another in isolation rates of the organism. Management system affected its occurrence, and only 7.1% of eviscerated carcasses yielded it. It was concluded that bird species, management system, and immersing slaughtered poultry in boiling water before dressing affect recovery of C. fetus subsp. jejuni from live birds and carcasses.

Draft Genome Sequence of Lactobacillus delbrueckii subsp. bulgaricus CRL871, a Folate-Producing Strain Isolated from a Northwestern Argentinian Yogurt.

PubMed

Laiño, Jonathan Emiliano; Hebert, Elvira María; Savoy de Giori, Graciela; LeBlanc, Jean Guy

2015-06-25

Lactobacillus delbrueckii subsp. bulgaricus CRL871 is the first strain of L. delbrueckii subsp. bulgaricus reported as a folate-producing strain. We report the draft genome sequence of L. delbrueckii subsp. bulgaricus CRL871 (2,063,981 bp, G+C content of 49.1%). This strain is of great biotechnological importance to the dairy industry because it constitutes an alternative to folic acid fortification. Copyright © 2015 Laiño et al.

Complete Genome Sequence of Campylobacter fetus subsp. venerealis Biovar Intermedius, Isolated from the Prepuce of a Bull

PubMed Central

Iraola, Gregorio; Pérez, Ruben; Naya, Hugo; Paolicchi, Fernando; Harris, David; Lawley, Trevor D.; Rego, Natalia; Hernández, Martín; Calleros, Lucía; Carretto, Luis; Velilla, Alejandra; Morsella, Claudia; Méndez, Alejandra

2013-01-01

Campylobacter fetus subsp. venerealis is the causative agent of bovine genital campylobacteriosis, a sexually transmitted disease distributed worldwide. Campylobacter fetus subsp. venerealis biovar Intermedius strains differ in their biochemical behavior and are prevalent in some countries. We report the first genome sequence for this biovar, isolated from bull prepuce. PMID:23908278

First identification of Francisella noatunensis subsp. orientalis causing mortality in Mexican tilapia Oreochromis spp.

PubMed

Ortega, Cesar; Mancera, Gerardo; Enríquez, Ricardo; Vargas, Augusto; Martínez, Simón; Fajardo, Raúl; Avendaño-Herrera, Ruben; Navarrete, María José; Romero, Alex

2016-08-09

Francisellosis, an emerging disease in tilapia Oreochromis spp., is caused by the facultative, intracellular bacterium Francisella noatunensis subsp. orientalis, which is present in various countries where tilapia farming is commercially important. We confirmed the presence of francisellosis in Mexican tilapia cultures in association with an outbreak during the second semester of 2012. Broodstock fish presented a mortality rate of approximately 40%, and disease was characterized by histologically classified granulomas, or whitish nodules, in different organs, mainly the spleen and kidney. Through DNA obtained from infected tissue and pure cultures in a cysteine heart medium supplemented with hemoglobin, F. noatunensis subsp. orientalis was initially confirmed through the amplification and analysis of the 16S rRNA gene and the internal transcribed spacer region. Phylogenetic analysis of these genes demonstrated close similarity with previously reported F. noatunensis subsp. orientalis sequences obtained from infected tilapia from various countries. The identification of this subspecies as the causative agent of the outbreak was confirmed using the iglC gene as a target sequence, which showed 99.5% identity to 2 F. noatunensis subsp. orientalis strains (Ethime-1 and Toba04). These findings represent the first documented occurrence of francisellosis in Mexican tilapia cultures, which highlights the importance of establishing preventative measures to minimize the spread of this disease within the Mexican aquaculture industry.

Quick detection of Leifsonia xyli subsp. xyli by PCR and necleotide sequence analysis of PCR amplicons from Chinese Leifsonia xyli subsp. xyli isolates

USDA-ARS?s Scientific Manuscript database

A quick polymerase chain reaction (PCR) assay was developed for the detection of Leifsonia xyli subsp. xyli (Lxx), the bacterial causal agent of ratoon stunting disease (RSD) of sugarcane, in crude juice samples from stalks. After removal of abiotic impurities and large molecular weight microorgani...

Transcriptome-Based Characterization of Interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in Lactose-Grown Chemostat Cocultures

PubMed Central

Mendes, Filipa; Sieuwerts, Sander; de Hulster, Erik; Almering, Marinka J. H.; Luttik, Marijke A. H.; Pronk, Jack T.; Smid, Eddy J.; Bron, Peter A.

2013-01-01

Mixed populations of Saccharomyces cerevisiae yeasts and lactic acid bacteria occur in many dairy, food, and beverage fermentations, but knowledge about their interactions is incomplete. In the present study, interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus, two microorganisms that co-occur in kefir fermentations, were studied during anaerobic growth on lactose. By combining physiological and transcriptome analysis of the two strains in the cocultures, five mechanisms of interaction were identified. (i) Lb. delbrueckii subsp. bulgaricus hydrolyzes lactose, which cannot be metabolized by S. cerevisiae, to galactose and glucose. Subsequently, galactose, which cannot be metabolized by Lb. delbrueckii subsp. bulgaricus, is excreted and provides a carbon source for yeast. (ii) In pure cultures, Lb. delbrueckii subsp. bulgaricus grows only in the presence of increased CO2 concentrations. In anaerobic mixed cultures, the yeast provides this CO2 via alcoholic fermentation. (iii) Analysis of amino acid consumption from the defined medium indicated that S. cerevisiae supplied alanine to the bacterium. (iv) A mild but significant low-iron response in the yeast transcriptome, identified by DNA microarray analysis, was consistent with the chelation of iron by the lactate produced by Lb. delbrueckii subsp. bulgaricus. (v) Transcriptome analysis of Lb. delbrueckii subsp. bulgaricus in mixed cultures showed an overrepresentation of transcripts involved in lipid metabolism, suggesting either a competition of the two microorganisms for fatty acids or a response to the ethanol produced by S. cerevisiae. This study demonstrates that chemostat-based transcriptome analysis is a powerful tool to investigate microbial interactions in mixed populations. PMID:23872557

Transcriptome-based characterization of interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus in lactose-grown chemostat cocultures.

PubMed

Mendes, Filipa; Sieuwerts, Sander; de Hulster, Erik; Almering, Marinka J H; Luttik, Marijke A H; Pronk, Jack T; Smid, Eddy J; Bron, Peter A; Daran-Lapujade, Pascale

2013-10-01

Mixed populations of Saccharomyces cerevisiae yeasts and lactic acid bacteria occur in many dairy, food, and beverage fermentations, but knowledge about their interactions is incomplete. In the present study, interactions between Saccharomyces cerevisiae and Lactobacillus delbrueckii subsp. bulgaricus, two microorganisms that co-occur in kefir fermentations, were studied during anaerobic growth on lactose. By combining physiological and transcriptome analysis of the two strains in the cocultures, five mechanisms of interaction were identified. (i) Lb. delbrueckii subsp. bulgaricus hydrolyzes lactose, which cannot be metabolized by S. cerevisiae, to galactose and glucose. Subsequently, galactose, which cannot be metabolized by Lb. delbrueckii subsp. bulgaricus, is excreted and provides a carbon source for yeast. (ii) In pure cultures, Lb. delbrueckii subsp. bulgaricus grows only in the presence of increased CO2 concentrations. In anaerobic mixed cultures, the yeast provides this CO2 via alcoholic fermentation. (iii) Analysis of amino acid consumption from the defined medium indicated that S. cerevisiae supplied alanine to the bacterium. (iv) A mild but significant low-iron response in the yeast transcriptome, identified by DNA microarray analysis, was consistent with the chelation of iron by the lactate produced by Lb. delbrueckii subsp. bulgaricus. (v) Transcriptome analysis of Lb. delbrueckii subsp. bulgaricus in mixed cultures showed an overrepresentation of transcripts involved in lipid metabolism, suggesting either a competition of the two microorganisms for fatty acids or a response to the ethanol produced by S. cerevisiae. This study demonstrates that chemostat-based transcriptome analysis is a powerful tool to investigate microbial interactions in mixed populations.

Persistence and Decontamination of Bacillus atrophaeus subsp. globigii Spores on Corroded Iron in a Model Drinking Water System▿

PubMed Central

Szabo, Jeffrey G.; Rice, Eugene W.; Bishop, Paul L.

2007-01-01

Persistence of Bacillus atrophaeus subsp. globigii spores on corroded iron coupons in drinking water was studied using a biofilm annular reactor. Spores were inoculated at 106 CFU/ml in the dechlorinated reactor bulk water. The dechlorination allowed for observation of the effects of hydraulic shear and biofilm sloughing on persistence. Approximately 50% of the spores initially adhered to the corroded iron surface were not detected after 1 month. Addition of a stable 10 mg/liter free chlorine residual after 1 month led to a 2-log10 reduction of adhered B. atrophaeus subsp. globigii, but levels on the coupons quickly stabilized thereafter. Increasing the free chlorine concentration to 25 or 70 mg/liter had no additional effect on inactivation. B. atrophaeus subsp. globigii spores injected in the presence of a typical distribution system chlorine residual (∼0.75 mg/liter) resulted in a steady reduction of adhered B. atrophaeus subsp. globigii over 1 month, but levels on the coupons eventually stabilized. Adding elevated chlorine levels (10, 25, and 70 mg/liter) after 1 month had no effect on the rate of inactivation. Decontamination with elevated free chlorine levels immediately after spore injection resulted in a 3-log10 reduction within 2 weeks, but the rate of inactivation leveled off afterward. This indicates that free chlorine did not reach portions of the corroded iron surface where B. atrophaeus subsp. globigii spores had adhered. B. atrophaeus subsp. globigii spores are capable of persisting for an extended time in the presence of high levels of free chlorine. PMID:17308186

Isolation of Mycobacterium avium subsp. paratuberculosis from Free-Ranging Birds and Mammals on Livestock Premises

PubMed Central

Corn, Joseph L.; Manning, Elizabeth J. B.; Sreevatsan, Srinand; Fischer, John R.

2005-01-01

Surveys for Mycobacterium avium subsp. paratuberculosis infection in free-ranging mammals and birds were conducted on nine dairy and beef cattle farms in Wisconsin and Georgia. Specimens were collected from 774 animals representing 25 mammalian and 22 avian species. Specimens of ileum, liver, intestinal lymph nodes, and feces were harvested from the larger mammals; a liver specimen and the gastrointestinal tract were harvested from birds and small mammals. Cultures were performed by using radiometric culture and acid-fast isolates were identified by 16S/IS900/IS1311 PCR and mycobactin dependency characteristics. M. avium subsp. paratuberculosis was cultured from tissues and feces from 39 samples from 30 animals representing nine mammalian and three avian species. The prevalence of infected wild animals by premises ranged from 2.7 to 8.3% in Wisconsin and from 0 to 6.0% in Georgia. Shedding was documented in seven (0.9%) animals: three raccoons, two armadillos, one opossum, and one feral cat. The use of two highly polymorphic short sequence repeat loci for analysis of 29 of the 39 strains identified 10 alleles. One allelic pattern broadly shared in domestic ruminants (“7,5”) appeared in approximately one-third of the wildlife M. avium subsp. paratuberculosis isolates studied. Given the few cases of shedding by free-ranging animals compared to the volume of contaminated manure produced by infected domestic ruminant livestock, contamination of the farm environment by infected wildlife was negligible. Wildlife may, however, have epidemiological significance for farms where M. avium subsp. paratuberculosis recently has been eliminated or on farms free of M. avium subsp. paratuberculosis but located in the geographic vicinity of farms with infected livestock. PMID:16269731

New Type of Antimicrobial Protein Produced by the Plant Pathogen Clavibacter michiganensis subsp. michiganensis

PubMed Central

Liu, Zhanliang; Ma, Ping; Holtsmark, Ingrid; Skaugen, Morten; Eijsink, Vincent G. H.

2013-01-01

It has previously been shown that the tomato pathogen Clavibacter michiganensis subsp. michiganensis secretes a 14-kDa protein, C. michiganensis subsp. michiganensis AMP-I (CmmAMP-I), that inhibits growth of Clavibacter michiganensis subsp. sepedonicus, the causal agent of bacterial ring rot of potato. Using sequences obtained from tryptic fragments, we have identified the gene encoding CmmAMP-I and we have recombinantly produced the protein with an N-terminal intein tag. The gene sequence showed that CmmAMP-I contains a typical N-terminal signal peptide for Sec-dependent secretion. The recombinant protein was highly active, with 50% growth inhibition (IC50) of approximately 10 pmol, but was not toxic to potato leaves or tubers. CmmAMP-I does not resemble any known protein and thus represents a completely new type of bacteriocin. Due to its high antimicrobial activity and its very narrow inhibitory spectrum, CmmAMP-1 may be of interest in combating potato ring rot disease. PMID:23851100

Salmonella enterica Suppresses Pectobacterium carotovorum subsp. carotovorum Population and Soft Rot Progression by Acidifying the Microaerophilic Environment

PubMed Central

Kwan, Grace; Charkowski, Amy O.; Barak, Jeri D.

2013-01-01

ABSTRACT Although enteric human pathogens are usually studied in the context of their animal hosts, a significant portion of their life cycle occurs on plants. Plant disease alters the phyllosphere, leading to enhanced growth of human pathogens; however, the impact of human pathogens on phytopathogen biology and plant health is largely unknown. To characterize the interaction between human pathogens and phytobacterial pathogens in the phyllosphere, we examined the interactions between Pectobacterium carotovorum subsp. carotovorum and Salmonella enterica or Escherichia coli O157:H7 with regard to bacterial populations, soft rot progression, and changes in local pH. The presence of P. carotovorum subsp. carotovorum enhanced the growth of both S. enterica and E. coli O157:H7 on leaves. However, in a microaerophilic environment, S. enterica reduced P. carotovorum subsp. carotovorum populations and soft rot progression by moderating local environmental pH. Reduced soft rot was not due to S. enterica proteolytic activity. Limitations on P. carotovorum subsp. carotovorum growth, disease progression, and pH elevation were not observed on leaves coinoculated with E. coli O157:H7 or when leaves were coinoculated with S. enterica in an aerobic environment. S. enterica also severely undermined the relationship between the phytobacterial population and disease progression of a P. carotovorum subsp. carotovorum budB mutant defective in the 2,3-butanediol pathway for acid neutralization. Our results show that S. enterica and E. coli O157:H7 interact differently with the enteric phytobacterial pathogen P. carotovorum subsp. carotovorum. S. enterica inhibition of soft rot progression may conceal a rapidly growing human pathogen population. Whereas soft rotted produce can alert consumers to the possibility of food-borne pathogens, healthy-looking produce may entice consumption of contaminated vegetables. PMID:23404399

Local genetic diversity of Xanthomonas citri subsp. citri in citrus orchards in northwest Paraná state, Brazil

USDA-ARS?s Scientific Manuscript database

Xanthomonas citri subsp. citri, causal agent of Asiatic citrus canker, is an important pathogen of citrus in Brazil and elsewhere. The genetic diversity of X. citri subsp. citri pathtype ‘A’ has not been studied in Brazil at a local scale (up to 300 km). A total of 40 isolates were collected from le...

Isolation of Bartonella vinsonii subsp. berkhoffii genotype II from a boy with epithelioid hemangioendothelioma and a dog with hemangiopericytoma.

PubMed

Breitschwerdt, Edward B; Maggi, Ricardo G; Varanat, Mrudula; Linder, Keith E; Weinberg, Guy

2009-06-01

In this report, we describe isolation of Bartonella vinsonii subsp. berkhoffii genotype II from a boy with epithelioid hemangioendothelioma and a dog with hemangiopericytoma. These results suggest that B. vinsonii subsp. berkhoffii may cause vasoproliferative lesions in both humans and dogs.

Genomic Diversity of Erwinia carotovora subsp. carotovora and Its Correlation with Virulence

PubMed Central

Yap, Mee-Ngan; Barak, Jeri D.; Charkowski, Amy O.

2004-01-01

We used genetic and biochemical methods to examine the genomic diversity of the enterobacterial plant pathogen Erwinia carotovora subsp. carotovora. The results obtained with each method showed that E. carotovora subsp. carotovora strains isolated from one ecological niche, potato plants, are surprisingly diverse compared to related pathogens. A comparison of 23 partial mdh sequences revealed a maximum pairwise difference of 10.49% and an average pairwise difference of 2.13%, values which are much greater than the maximum variation (1.81%) and average variation (0.75%) previously reported for Escherichia coli. Pulsed-field gel electrophoresis analysis of I-CeuI-digested genomic DNA revealed seven rrn operons in all E. carotovora subsp. carotovora strains examined except strain WPP17, which had only six copies. We identified 26 I-CeuI restriction fragment length polymorphism patterns and observed significant polymorphism in fragment sizes ranging from 100 to 450 kb for all strains. We detected large plasmids in two strains, including the model strain E. carotovora subsp. carotovora 71. The two least virulent strains had an unusual chromosomal structure, suggesting that a particular pulsotype is correlated with virulence. To compare chromosomal organization of multiple enterobacterial genomes, several genes were mapped onto I-CeuI fragments. We identified portions of the genome that appear to be conserved across enterobacteria and portions that have undergone genome rearrangements. We found that the least virulent strain, WPP17, failed to oxidize cellobiose and was missing several hrp and hrc genes. The unexpected variability among isolates obtained from clonal hosts in one region and in one season suggests that factors other than the host plant, potato, drive the evolution of this common environmental bacterium and key plant pathogen. PMID:15128563

Isolation of Bartonella vinsonii subsp. berkhoffii Genotype II from a Boy with Epithelioid Hemangioendothelioma and a Dog with Hemangiopericytoma▿

PubMed Central

Breitschwerdt, Edward B.; Maggi, Ricardo G.; Varanat, Mrudula; Linder, Keith E.; Weinberg, Guy

2009-01-01

In this report, we describe isolation of Bartonella vinsonii subsp. berkhoffii genotype II from a boy with epithelioid hemangioendothelioma and a dog with hemangiopericytoma. These results suggest that B. vinsonii subsp. berkhoffii may cause vasoproliferative lesions in both humans and dogs. PMID:19369441

Allelopathic activity of Nepeta nuda L. subsp. nuda water extracts

NASA Astrophysics Data System (ADS)

Dragoeva, Asya; Stoyanova, Zheni; Koleva, Vanya; Dragolova, Daniela

2017-03-01

Nepeta nuda subsp. nuda is a medicinal plant growing wild in Bulgaria. Different species of Nepeta genus have been reported to possess allelopathic potential. The present study was conducted to observe its phytotoxic effects on T. aestivum and C. sativus L. seeds in laboratory conditions. Nepeta water extracts (NWE) prepared from aerial parts of plants at concentrations 2, 4, 6, 8, 10, 12 and 14 g/l were tested. The rate of seed germination, the root and shoot length, fresh and dry weight of seedlings were observed after treatment with NWE. As a control served seeds treated with distilled water. Germination was not affected, but NWE showed deterioration in seedling growth. Roots were more affected than shoots. The fresh and dry weights were reduced upon treatment with the extracts tested. These negative effects were dose-dependent. The overall results indicate presence of water soluble allelochemicals in Nepeta nuda subsp. nuda.

Desulfovibrio oceani subsp. oceani sp. nov., subsp. nov. and Desulfovibrio oceani subsp. galateae subsp. nov., novel sulfate-reducing bacteria isolated from the oxygen minimum zone off the coast of Peru.

PubMed

Finster, Kai W; Kjeldsen, Kasper U

2010-03-01

Two deltaproteobacterial sulfate reducers, designated strain I.8.1(T) and I.9.1(T), were isolated from the oxygen minimum zone water column off the coast of Peru at 400 and 500 m water depth. The strains were Gram-negative, vibrio-shaped and motile. Both strains were psychrotolerant, grew optimally at 20 degrees C at pH 7.0-8.0 and at 2.5-3.5% NaCl (w/v). The strains grew by utilizing hydrogen/acetate, C(3-4) fatty acids, amino acids and glycerol as electron acceptors for sulfate reduction. Fumarate, lactate and pyruvate supported fermentative growth. Sulfate, sulfite, thiosulfate and taurin supported growth as electron acceptors. Both strains were catalase-positive and highly oxygen-tolerant, surviving 24 days of exposure to atmospheric concentrations. MK6 was the only respiratory quinone. The most prominent cellular fatty acid was iso-17:1-omega9c (18%) for strain I.8.1(T) and iso-17:0-omega9c (14%) for strain I.9.1(T). The G+C contents of their genomic DNA were 45-46 mol%. Phylogenetic analysis of 16S rRNA and dsrAB gene sequences showed that both strains belong to the genus Desulfovibrio. Desulfovibrio acrylicus DSM 10141(T) and Desulfovibrio marinisediminis JCM 14577(T) represented their closest validly described relatives with pairwise 16S rRNA gene sequence identities of 98-99%. The level of DNA-DNA hybridization between strains I.8.1(T) and I.9.1(T) was 30-38%. The two strains shared 10-26% DNA-DNA relatedness with D. acrylicus. Based on a polyphasic investigation it is proposed that strains I.8.1(T) and I.9.1(T) represent a novel species for which the name Desulfovibrio oceani sp. nov. is proposed with the two subspecies D. oceani subsp. oceani (type strain, I.8.1(T) = DSM 21390(T) = JCM 15970(T)) and D. oceani subsp. galateae (type strain, I.9.1(T) = DSM 21391(T) = JCM 15971(T)).

Reclassification of Lactobacillus kefirgranum Takizawa et al. 1994 as Lactobacillus kefiranofaciens subsp. kefirgranum subsp. nov. and emended description of L. kefiranofaciens Fujisawa et al. 1988.

PubMed

Vancanneyt, M; Mengaud, J; Cleenwerck, I; Vanhonacker, K; Hoste, B; Dawyndt, P; Degivry, M C; Ringuet, D; Janssens, D; Swings, J

2004-03-01

Fourteen homofermentative lactic acid bacteria that were isolated from kefir grains and kefir fermented milks were assigned to either Lactobacillus kefiranofaciens or Lactobacillus kefirgranum, based on their characteristic morphotypes, phenotypic features and SDS-PAGE profiles of whole-cell proteins. Further genotypic analyses on representative strains from both taxa demonstrated that L. kefiranofaciens and L. kefirgranum share 100 % 16S rDNA sequence similarity and belong phylogenetically to the Lactobacillus acidophilus species group. DNA-DNA binding values of >79 % and analogous DNA G+C contents of 37-38 mol% showed that the strains studied belonged to one species: L. kefirgranum is a later synonym of L. kefiranofaciens. An emended description is proposed for L. kefiranofaciens. Due to the specific morphological and biochemical characteristics of these taxa in kefir grain formation, it is proposed that L. kefirgranum should be reclassified as L. kefiranofaciens subsp. kefirgranum subsp. nov.

Heterologous expression of Oenococcus oeni malolactic enzyme in Lactobacillus plantarum for improved malolactic fermentation

PubMed Central

2012-01-01

Lactobacillus plantarum is involved in a multitude of food related industrial fermentation processes including the malolactic fermentation (MLF) of wine. This work is the first report on a recombinant L. plantarum strain successfully conducting MLF. The malolactic enzyme (MLE) from Oenococcus oeni was cloned into the lactobacillal expression vector pSIP409 which is based on the sakacin P operon of Lactobacillus sakei and expressed in the host strain L. plantarum WCFS1. Both recombinant and wild-type L. plantarum strains were tested for MLF using a buffered malic acid solution in absence of glucose. Under the conditions with L-malic acid as the only energy source and in presence of Mn2+ and NAD+, the recombinant L. plantarum and the wild-type strain converted 85% (2.5 g/l) and 51% (1.5 g/l), respectively, of L-malic acid in 3.5 days. Furthermore, the recombinant L. plantarum cells converted in a modified wine 15% (0.4 g/l) of initial L-malic acid concentration in 2 days. In conclusion, recombinant L. plantarum cells expressing MLE accelerate the malolactic fermentation. PMID:22452826

Lactic acid bacteria associated with a heat-processed pork product and sources of variation affecting chemical indices of spoilage and sensory characteristics.

PubMed

Laursen, B G; Byrne, D V; Kirkegaard, J B; Leisner, J J

2009-02-01

To evaluate the potential for developing a quality index for a Danish modified atmosphere packaged (MAP) heat-processed and naturally contaminated pork meat product stored at 5 degrees C. The composition of the predominating microflora and changes in contents of tyramine, arginine, organic acids and sensory characteristics were analysed. The microflora was predominated by Lactobacillus sakei, Leuconostoc carnosum and Carnobacterium divergens. The presence of each species varied between products and batches resulting in limited usefulness of the concentrations of these bacteria or their metabolites as indices of quality. Furthermore, the three species differed in their metabolic activities as shown by use of a model meat extract. However, when MAP storage of the processed pork product was followed by aerobic storage then acetic acid showed some potential as a chemical indicator of sensory quality. Variation in processing parameters and spoilage microbiota limited the usefulness of concentrations of micro-organisms and their metabolites as indices of spoilage for the studied processed MAP pork product. The present study contributes to an understanding of the difficulties experienced in developing quality indices to be used in the control of microbial spoilage of processed MAP meat products.

Investigation of lactic acid bacterial strains for meat fermentation and the product's antioxidant and angiotensin-I-converting-enzyme inhibitory activities.

PubMed

Takeda, Shiro; Matsufuji, Hisashi; Nakade, Koji; Takenoyama, Shin-Ichi; Ahhmed, Abdulatef; Sakata, Ryoichi; Kawahara, Satoshi; Muguruma, Michio

2017-03-01

In the lactic acid bacteria (LAB) strains screened from our LAB collection, Lactobacillus (L.) sakei strain no. 23 and L. curvatus strain no. 28 degraded meat protein and tolerated salt and nitrite in vitro. Fermented sausages inoculated strains no. 23 and no. 28 showed not only favorable increases in viable LAB counts and reduced pH, but also the degradation of meat protein. The sausages fermented with these strains showed significantly higher antioxidant activity than those without LAB or fermented by each LAB type strain. Angiotensin-I-converting-enzyme (ACE) inhibitory activity was also significantly higher in the sausages fermented with strain no. 23 than in those fermented with the type strain. Higher ACE inhibitory activity was also observed in the sausages fermented with strain no. 28, but did not differ significantly from those with the type strain. An analysis of the proteolysis and degradation products formed by each LAB in sausages suggested that those bioactivities yielded fermentation products such as peptides. Therefore, LAB starters that can adequately ferment meat, such as strains no. 23 and no. 28, should contribute to the production of bioactive compounds in meat products. © 2016 Japanese Society of Animal Science.

Construction of a food-grade cloning vector for Lactobacillus plantarum and its utilization in a food model.

PubMed

Rattanachaikunsopon, Pongsak; Phumkhachorn, Parichat

2012-01-01

The development of Lactobacillus plantarum to be used in starter cultures in the food industry has been limited because of the lack of a food-grade cloning vector for the bacterium. In this study, the plasmid pFLP1 was constructed by joining 2 DNA fragments derived from food-approved organisms. The 5.2-kb BamHI/KpnI DNA fragment of pRV566 containing the theta-type replicon of Lactobacillus sakei was ligated to the BamHI/KpnI DNA fragment of a 2.9-kb lactococcal cadmium resistance determinant amplified from pND918. The 8.1-kb newly constructed plasmid could transform L. plantarum N014, a bacteriocin-producing bacteria originally isolated from nham, a traditional Thai fermented sausage. The resulting transformant, L. plantarum N014-FLP, and its parent strain were shown to be very similar in growth rate and bacteriocin activity. In addition, the plasmid was very stable in its host bacteria under nonselective pressure for 100 generations in MRS medium and for 5 days in a nham model. These results suggest that pFLP1 is a potential food-grade cloning vector for L. plantarum.

Metagenomic approach reveals microbial diversity and predictive microbial metabolic pathways in Yucha, a traditional Li fermented food.

PubMed

Zhang, Jiachao; Wang, Xiaoru; Huo, Dongxue; Li, Wu; Hu, Qisong; Xu, Chuanbiao; Liu, Sixin; Li, Congfa

2016-08-31

Yucha is a typical traditional fermented food of the Li population in the Hainan province of China, and it is made up of cooked rice and fresh fish. In the present study, metagenomic approach and culture-dependent technology were applied to describe the diversity of microbiota and identify beneficial microbes in the Yucha. At the genus level, Lactobacillus was the most abundant genus (43.82% of the total reads), followed by Lactococcus, Enterococcus, Vibrio, Weissella, Pediococcus, Enterobacter, Salinivibrio, Acinetobacter, Macrococcus, Kluyvera and Clostridium; this result was confirmed by q-PCR. PCoA based on Weighted UniFrac distances showed an apparent clustering pattern for Yucha samples from different locations, and Lactobacillus sakei, Lactobacillus saniviri and Staphylococcus sciuri represented OTUs according to the major identified markers. At the microbial functional level, it was observed that there was an enrichment of metabolic functional features, including amino acid and carbohydrate metabolism, which implied that the microbial metabolism in the Yucha samples tended to be vigorous. Accordingly, we further investigated the correlation between the predominant microbes and metabolic functional features. Thirteen species of Lactobacillus (147 strains) were isolated, and Lactobacillus plantarum (60 isolates) and Lactobacillus pentosus (34 isolates) were isolated from every sample.

Factors Affecting Exocellular Polysaccharide Production by Lactobacillus delbrueckii subsp. bulgaricus Grown in a Chemically Defined Medium†

PubMed Central

Petry, Sandrine; Furlan, Sylviane; Crepeau, Marie-Jeanne; Cerning, Jutta; Desmazeaud, Michel

2000-01-01

We developed a chemically defined medium (CDM) containing lactose or glucose as the carbon source that supports growth and exopolysaccharide (EPS) production of two strains of Lactobacillus delbrueckii subsp. bulgaricus. The factors found to affect EPS production in this medium were oxygen, pH, temperature, and medium constituents, such as orotic acid and the carbon source. EPS production was greatest during the stationary phase. Composition analysis of EPS isolated at different growth phases and produced under different fermentation conditions (varying carbon source or pH) revealed that the component sugars were the same. The EPS from strain L. delbrueckii subsp. bulgaricus CNRZ 1187 contained galactose and glucose, and that of strain L. delbrueckii subsp. bulgaricus CNRZ 416 contained galactose, glucose, and rhamnose. However, the relative proportions of the individual monosaccharides differed, suggesting that repeating unit structures can vary according to specific medium alterations. Under pH-controlled fermentation conditions, L. delbrueckii subsp. bulgaricus strains produced as much EPS in the CDM as in milk. Furthermore, the relative proportions of individual monosaccharides of EPS produced in pH-controlled CDM or in milk were very similar. The CDM we developed may be a useful model and an alternative to milk in studies of EPS production. PMID:10919802

Inhibition of protein glycation by essential oils of branchlets and fruits of Juniperus communis subsp. hemisphaerica

PubMed Central

Asgary, S.; Naderi, G.A.; Shams Ardekani, M.R.; Sahebkar, A.; Airin, A.; Aslani, S.; Kasher, T.; Emami, S.A.

2014-01-01

Oxidative stress and protein glycation play pivotal roles in the pathophysiology of diabetes mellitus and its vascular complications. The present study aimed to investigate the anti-glycation properties of essential oils obtained from different parts of Juniperus communis subsp. hemisphaerica. The branchlets of male tree (BMT) and branchlets of female (BFT) tree, and fruits of J. communis subsp. hemisphaerica were extracted using steam distillation method. The oils were phytochemically analyzed using gas chromatography-mass spectrometry. Anti-glycation properties were evaluated using hemoglobin and insulin glycation assays. Overall, 18 volatile components were identified in the J. communis subsp. hemisphaerica oils, amounting to 82.1%, 100.0% and 96.4% of the BMT, BFT and fruit oils, respectively. Promising inhibitory activity was observed from all concentrations of the tested oils in the hemoglobin and insulin glycation assays. The inhibitory activities peaked to 89.9% (BFT oil; 200 μg mL-1) and 81.0% (BFT oil; 600 μg mL-1) in the hemoglobin and insulin glycation assays, respectively. The evidence from this study suggests that essential oils obtained from the fruits and branchlets of J. communis subsp. hemisphaerica possess anti-glycation properties. These activities may find implication for the prevention and treatment of diabetic complications. PMID:25657787

Inhibition of protein glycation by essential oils of branchlets and fruits of Juniperus communis subsp. hemisphaerica.

PubMed

Asgary, S; Naderi, G A; Shams Ardekani, M R; Sahebkar, A; Airin, A; Aslani, S; Kasher, T; Emami, S A

2014-01-01

Oxidative stress and protein glycation play pivotal roles in the pathophysiology of diabetes mellitus and its vascular complications. The present study aimed to investigate the anti-glycation properties of essential oils obtained from different parts of Juniperus communis subsp. hemisphaerica. The branchlets of male tree (BMT) and branchlets of female (BFT) tree, and fruits of J. communis subsp. hemisphaerica were extracted using steam distillation method. The oils were phytochemically analyzed using gas chromatography-mass spectrometry. Anti-glycation properties were evaluated using hemoglobin and insulin glycation assays. Overall, 18 volatile components were identified in the J. communis subsp. hemisphaerica oils, amounting to 82.1%, 100.0% and 96.4% of the BMT, BFT and fruit oils, respectively. Promising inhibitory activity was observed from all concentrations of the tested oils in the hemoglobin and insulin glycation assays. The inhibitory activities peaked to 89.9% (BFT oil; 200 μg mL(-1)) and 81.0% (BFT oil; 600 μg mL(-1)) in the hemoglobin and insulin glycation assays, respectively. The evidence from this study suggests that essential oils obtained from the fruits and branchlets of J. communis subsp. hemisphaerica possess anti-glycation properties. These activities may find implication for the prevention and treatment of diabetic complications.

Mycobacterium avium subsp. paratuberculosis infection, immunology and pathology of livestock

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) infection in ruminants leads to a chronic and progressive enteric disease (Johne’s disease) that results in loss of intestinal function, poor body condition, and eventual death. Transmission is primarily through a fecal-oral route in neonates but con...

Understanding the ontogeny and succession of Bacillus velezensis and B. subtilis subsp. subtilis by focusing on kimchi fermentation.

PubMed

Cho, Min Seok; Jin, Yong Ju; Kang, Bo Kyoung; Park, Yu Kyoung; Kim, ChangKug; Park, Dong Suk

2018-05-04

Bacillus subtilis and B. velezensis are frequently isolated from various niches, including fermented foods, water, and soil. Within the Bacillus subtilis group, B. velezensis and B. subtilis subsp. subtilis have received significant attention as biological resources for biotechnology-associated industries. Nevertheless, radical solutions are urgently needed to identify microbes during their ecological succession to accurately confirm their action at the species or subspecies level in diverse environments, such as fermented materials. Thus, in this study, previously published genome data of the B. subtilis group were compared to exploit species- or subspecies-specific genes for use as improved qPCR targets to detect B. velezensis and B. subtilis subsp. subtilis in kimchi samples. In silico analyses of the selected genes and designed primer sequences, in conjunction with SYBR Green real-time PCR, confirmed the robustness of this newly developed assay. Consequently, this study will allow for new insights into the ontogeny and succession of B. velezensis and B. subtilis subsp. subtilis in various niches. Interestingly, in white kimchi without red pepper powder, neither B. subtilis subsp. subtilis nor B. velezensis was detected.

Geography of Genetic Structure in Barley Wild Relative Hordeum vulgare subsp. spontaneum in Jordan.

PubMed

Thormann, Imke; Reeves, Patrick; Reilley, Ann; Engels, Johannes M M; Lohwasser, Ulrike; Börner, Andreas; Pillen, Klaus; Richards, Christopher M

2016-01-01

Informed collecting, conservation, monitoring and utilization of genetic diversity requires knowledge of the distribution and structure of the variation occurring in a species. Hordeum vulgare subsp. spontaneum (K. Koch) Thell., a primary wild relative of barley, is an important source of genetic diversity for barley improvement and co-occurs with the domesticate within the center of origin. We studied the current distribution of genetic diversity and population structure in H. vulgare subsp. spontaneum in Jordan and investigated whether it is correlated with either spatial or climatic variation inferred from publically available climate layers commonly used in conservation and ecogeographical studies. The genetic structure of 32 populations collected in 2012 was analyzed with 37 SSRs. Three distinct genetic clusters were identified. Populations were characterized by admixture and high allelic richness, and genetic diversity was concentrated in the northern part of the study area. Genetic structure, spatial location and climate were not correlated. This may point out a limitation in using large scale climatic data layers to predict genetic diversity, especially as it is applied to regional genetic resources collections in H. vulgare subsp. spontaneum.

Lymphoproliferative and gamma interferon responses to stress-regulated Mycobacterium avium subsp. paratuberculosis recombinant proteins.

PubMed

Gurung, Ratna B; Begg, Douglas J; Purdie, Auriol C; de Silva, Kumudika; Bannantine, John P; Whittington, Richard J

2014-06-01

Johne's disease in ruminants is a chronic infection of the intestines caused by Mycobacterium avium subsp. paratuberculosis. An important strategy to control disease is early detection, and a potentially efficient method for early detection is measurement of cell-mediated immune responses developed by the host in response to exposure or infection. One method is to measure lymphoproliferation and cytokine release from the host cells when exposed to the organism or parts of the organism. In this study, 10 recombinant M. avium subsp. paratuberculosis proteins known to be upregulated under in vitro stress conditions were evaluated by examining their ability to evoke memory as a result of exposure by vaccination or oral challenge with live Mycobacterium avium subsp. paratuberculosis. Out of 10 proteins, MAP2698c was found to induce higher cell-mediated immune responses in vaccinated and challenged sheep in comparison to healthy controls. The findings suggest that not all stress-regulated proteins have the diagnostic potential to detect cell-mediated immune responses in ovine paratuberculosis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Lymphoproliferative and Gamma Interferon Responses to Stress-Regulated Mycobacterium avium subsp. paratuberculosis Recombinant Proteins

PubMed Central

Gurung, Ratna B.; Begg, Douglas J.; Purdie, Auriol C.; de Silva, Kumudika; Bannantine, John P.

2014-01-01

Johne's disease in ruminants is a chronic infection of the intestines caused by Mycobacterium avium subsp. paratuberculosis. An important strategy to control disease is early detection, and a potentially efficient method for early detection is measurement of cell-mediated immune responses developed by the host in response to exposure or infection. One method is to measure lymphoproliferation and cytokine release from the host cells when exposed to the organism or parts of the organism. In this study, 10 recombinant M. avium subsp. paratuberculosis proteins known to be upregulated under in vitro stress conditions were evaluated by examining their ability to evoke memory as a result of exposure by vaccination or oral challenge with live Mycobacterium avium subsp. paratuberculosis. Out of 10 proteins, MAP2698c was found to induce higher cell-mediated immune responses in vaccinated and challenged sheep in comparison to healthy controls. The findings suggest that not all stress-regulated proteins have the diagnostic potential to detect cell-mediated immune responses in ovine paratuberculosis. PMID:24695774

Genetic Variability and Population Structure of Disanthus cercidifolius subsp. longipes (Hamamelidaceae) Based on AFLP Analysis

PubMed Central

Yu, Yi; Fan, Qiang; Shen, Rujiang; Guo, Wei; Jin, Jianhua; Cui, Dafang; Liao, Wenbo

2014-01-01

Disanthus cercidifolius subsp. longipes is an endangered species in China. Genetic diversity and structure analysis of this species was investigated using amplified fragments length polymorphism (AFLP) fingerprinting. Nei's gene diversity ranged from 0.1290 to 0.1394. The AMOVA indicated that 75.06% of variation was distributed within populations, while the between-group component 5.04% was smaller than the between populations-within-group component 19.90%. Significant genetic differentiation was detected between populations. Genetic and geographical distances were not correlated. PCA and genetic structure analysis showed that populations from East China were together with those of the Nanling Range. These patterns of genetic diversity and levels of genetic variation may be the result of D. c. subsp. longipes restricted to several isolated habitats and “excess flowers production, but little fruit set”. It is necessary to protect all existing populations of D. c. subsp. longipes in order to preserve as much genetic variation as possible. PMID:25250583

Interaction between 2,4-Diacetylphloroglucinol- and Hydrogen Cyanide-Producing Pseudomonas brassicacearum LBUM300 and Clavibacter michiganensis subsp. michiganensis in the Tomato Rhizosphere

PubMed Central

Paulin, Mélanie M.; Novinscak, Amy; Lanteigne, Carine; Gadkar, Vijay J.

2017-01-01

ABSTRACT We have previously demonstrated that inoculation of tomato plants with 2,4-diacetylphloroglucinol (DAPG)- and hydrogen cyanide (HCN)-producing Pseudomonas brassicacearum LBUM300 could significantly reduce bacterial canker symptoms caused by Clavibacter michiganensis subsp. michiganensis. In this study, in order to better characterize the population dynamics of LBUM300 in the rhizosphere of tomato plants, we characterized the role played by DAPG and HCN production by LBUM300 on rhizosphere colonization of healthy and C. michiganensis subsp. michiganensis-infected tomato plants. The impact of C. michiganensis subsp. michiganensis presence on the expression of DAPG and HCN biosynthetic genes in the rhizosphere was also examined. In planta assays were performed using combinations of C. michiganensis subsp. michiganensis and wild-type LBUM300 or DAPG (LBUM300ΔphlD) or HCN (LBUM300ΔhcnC) isogenic mutant strains. Populations of LBUM300 and phlD and hcnC gene expression levels were quantified in rhizosphere soil at several time points up to 264 h postinoculation using culture-independent quantitative PCR (qPCR) and reverse transcriptase quantitative PCR (RT-qPCR) TaqMan assays, respectively. The presence of C. michiganensis subsp. michiganensis significantly increased rhizospheric populations of LBUM300. In C. michiganensis subsp. michiganensis-infected tomato rhizospheres, the populations of wild-type LBUM300 and strain LBUM300ΔhcnC, both producing DAPG, were significantly higher than the population of strain LBUM300ΔphlD. A significant upregulation of phlD expression was observed in the presence of C. michiganensis subsp. michiganensis, while hcnC expression was only slightly increased in the mutant strain LBUM300ΔphlD when C. michiganensis subsp. michiganensis was present. Additionally, biofilm production was found to be significantly reduced in strain LBUM300ΔphlD compared to the wild-type and LBUM300ΔhcnC strains. IMPORTANCE The results of this study

Four new diterpenoid alkaloids from Aconitum japonicum subsp. subcuneatum.

PubMed

Yamashita, Hiroshi; Takeda, Keiko; Haraguchi, Machiko; Abe, Yuki; Kuwahara, Natsumi; Suzuki, Shota; Terui, Ayaka; Masaka, Takumi; Munakata, Naoko; Uchida, Mariko; Nunokawa, Masashi; Kaneda, Kyousuke; Goto, Masuo; Lee, Kuo-Hsiung; Wada, Koji

2018-01-01

Diterpenoid alkaloids with remarkable chemical properties and biological activities are frequently found in plants of the genera Aconitum, Delphinium, and Garrya. Accordingly, several diterpenoid alkaloid constituents of Aconitum and Delphinium plants as well as their derivatives exhibited cytotoxic activity against lung, prostate, nasopharyngeal, and vincristine-resistant nasopharyngeal cancer cell lines. Four new C 19 -diterpenoid alkaloids, 14-anisoyllasianine (1), 14-anisoyl-N-deethylaconine (2), N-deethylaljesaconitine A (3), and N-deethylnevadensine (4), together with 17 known C 19 - and C 20 -diterpenoid alkaloids, were isolated in a phytochemical investigation of rhizoma of Aconitum japonicum THUNB. subsp. subcuneatum (NAKAI) KADOTA. Their structures were elucidated by extensive spectroscopic methods including NMR (1D and 2D), IR, and MS (HRMS). Eight known diterpenoid alkaloids, lipoaconitine, lipomesaconitine, aconine, nevadenine, talatisamine, nevadensine, ryosenamine, and dehydrolucidusculine, were isolated the first time from A. japonicum subsp. subcuneatum. Three of the new C 19 -diterpenoid alkaloids (1, 3, 4) and six of the known diterpenoid alkaloids were evaluated for cytotoxic activity against five human tumor cell lines.

Culture- and quantitative IS900 real-time PCR-based analysis of the persistence of Mycobacterium avium subsp. paratuberculosis in a controlled dairy cow farm environment.

PubMed

Moravkova, M; Babak, V; Kralova, A; Pavlik, I; Slana, I

2012-09-01

The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculosis in environmental samples taken from a Holstein farm with a long history of clinical paratuberculosis. A herd of 606 head was eradicated, and mechanical cleaning and disinfection with chloramine B with ammonium (4%) was carried out on the farm; in the surrounding areas (on the field and field midden) lime was applied. Environmental samples were collected before and over a period of 24 months after destocking. Only one sample out of 48 (2%) examined on the farm (originating from a waste pit and collected before destocking) was positive for M. avium subsp. paratuberculosis by cultivation on solid medium (Herrold's egg yolk medium). The results using real-time quantitative PCR (qPCR) showed that a total of 81% of environmental samples with an average mean M. avium subsp. paratuberculosis cell number of 3.09 × 10(3) were positive for M. avium subsp. paratuberculosis before destocking compared to 43% with an average mean M. avium subsp. paratuberculosis cell number of 5.86 × 10(2) after 24 months. M. avium subsp. paratuberculosis-positive samples were detected in the cattle barn as well as in the calf barn and surrounding areas. M. avium subsp. paratuberculosis was detected from different matrices: floor and instrument scrapings, sediment, or scraping from watering troughs, waste pits, and cobwebs. M. avium subsp. paratuberculosis DNA was also detected in soil and plants collected on the field midden and the field 24 months after destocking. Although the proportion of positive samples decreased from 64% to 23% over time, the numbers of M. avium subsp. paratuberculosis cells were comparable.

Culture- and Quantitative IS900 Real-Time PCR-Based Analysis of the Persistence of Mycobacterium avium subsp. paratuberculosis in a Controlled Dairy Cow Farm Environment

PubMed Central

Moravkova, M.; Babak, V.; Kralova, A.; Pavlik, I.

2012-01-01

The aim of this study was to monitor the persistence of Mycobacterium avium subsp. paratuberculosis in environmental samples taken from a Holstein farm with a long history of clinical paratuberculosis. A herd of 606 head was eradicated, and mechanical cleaning and disinfection with chloramine B with ammonium (4%) was carried out on the farm; in the surrounding areas (on the field and field midden) lime was applied. Environmental samples were collected before and over a period of 24 months after destocking. Only one sample out of 48 (2%) examined on the farm (originating from a waste pit and collected before destocking) was positive for M. avium subsp. paratuberculosis by cultivation on solid medium (Herrold's egg yolk medium). The results using real-time quantitative PCR (qPCR) showed that a total of 81% of environmental samples with an average mean M. avium subsp. paratuberculosis cell number of 3.09 × 103 were positive for M. avium subsp. paratuberculosis before destocking compared to 43% with an average mean M. avium subsp. paratuberculosis cell number of 5.86 × 102 after 24 months. M. avium subsp. paratuberculosis-positive samples were detected in the cattle barn as well as in the calf barn and surrounding areas. M. avium subsp. paratuberculosis was detected from different matrices: floor and instrument scrapings, sediment, or scraping from watering troughs, waste pits, and cobwebs. M. avium subsp. paratuberculosis DNA was also detected in soil and plants collected on the field midden and the field 24 months after destocking. Although the proportion of positive samples decreased from 64% to 23% over time, the numbers of M. avium subsp. paratuberculosis cells were comparable. PMID:22773642

Derivation of Mutants of Erwinia carotovora subsp. betavasculorum Deficient in Export of Pectolytic Enzymes with Potential for Biological Control of Potato Soft Rot

PubMed Central

Costa, José M.; Loper, Joyce E.

1994-01-01

Erwinia carotovora subsp. betavasculorum Ecb168 produces an antibiotic(s) that suppresses growth of the related bacterium Erwinia carotovora subsp. carotovora in culture and in wounds of potato tubers. Strain Ecb168 also produces and secretes pectolytic enzymes and causes a vascular necrosis and root rot of sugar beet. Genes (out) involved in secretion of pectolytic enzymes by Ecb168 were localized to two HindIII fragments (8.5 and 10.5 kb) of Ecb168 genomic DNA by hybridization to the cloned out region of E. carotovora subsp. carotovora and by complementation of Out- mutants of E. carotovora subsp. carotovora. Out- mutants of Ecb168, which did not secrete pectate lyase into the culture medium, were obtained when deletions internal to either HindIII fragment were introduced into the genome of Ecb168 through marker exchange mutagenesis. Out- mutants of Ecb168 were complemented to the Out+ phenotype by introduction of the corresponding cloned HindIII fragment. Out- mutants of Ecb168 were less virulent than the Out+ parental strain on potato tubers. Strain Ecb168 and Out- derivatives inhibited the growth of E. carotovora subsp. carotovora in culture, indicating that the uncharacterized antibiotic(s) responsible for antagonism was exported through an out-independent mechanism. Strain Ecb168 and Out- derivatives reduced the establishment of large populations of E. carotovora subsp. carotovora in wounds of potato tubers and suppressed tuber soft rot caused by E. carotovora subsp. carotovora. PMID:16349316

Antioxidant Activity of the Essential Oils of Different Parts of Juniperus excelsa M. Bieb. subsp. excelsa and J. excelsa M. Bieb. subsp. polycarpos (K. Koch) Takhtajan (Cupressaceae)

PubMed Central

Emami, Sayyed Ahmad; Abedindo, Bibi Fatemeh; Hassanzadeh-Khayyat, Mohammad

2011-01-01

The essential oils of branchlets and fruits of Juniperus excelsa subsp. excelsa and Juniperus excelsa subsp. polycarpos were examined for their antioxidant activity. The compositions of the essential oils were studied by GC and GC-MS. To evaluation the antioxidants activity of the volatile oils, pure components and positive controls at different concentrations, thin-layer chromatography (TLC) screening methods, diphenylpicrylhydrazyl (DPPH) assay, deoxyribose degradation test and modified deoxyribose degradation test were employed. The results of the present study demonstrate some antioxidant activity for the tested essential oils obtained from various parts of both plants. It indicates that the use of these essential oils, in very low concentrations, may be useful as a natural preservative. However before any final conclusion, it is suggested that the antioxidant activity of these oils should also be evaluated by using lipid solvent system methods. PMID:24250416

A quantitative and direct PCR assay for the subspecies-specific detection of Clavibacter michiganensis subsp. michiganensis based on a ferredoxin reductase gene.

PubMed

Cho, Min Seok; Lee, Jang Ha; Her, Nam Han; Kim, Changkug; Seol, Young-Joo; Hahn, Jang Ho; Baeg, Ji Hyoun; Kim, Hong Gi; Park, Dong Suk

2012-06-01

The Gram-positive bacterium Clavibacter michiganensis subsp. michiganensis is the causal agent of canker disease in tomato. Because it is very important to control newly introduced inoculum sources from commercial materials, the specific detection of this pathogen in seeds and seedlings is essential for effective disease control. In this study, a novel and efficient assay for the detection and quantitation of C. michiganensis subsp. michiganensis in symptomless tomato and red pepper seeds was developed. A pair of polymerase chain reaction (PCR) primers (Cmm141F/R) was designed to amplify a specific 141 bp fragment on the basis of a ferredoxin reductase gene of C. michiganensis subsp. michiganensis NCPPB 382. The specificity of the primer set was evaluated using purified DNA from 16 isolates of five C. michiganensis subspecies, one other Clavibacter species, and 17 other reference bacteria. The primer set amplified a single band of expected size from the genomic DNA obtained from the C. michiganensis subsp. michiganensis strains but not from the other C. michiganensis subspecies or from other Clavibacter species. The detection limit was a single cloned copy of the ferredoxin reductase gene of C. michiganensis subsp. michiganensis. In conclusion, this quantitative direct PCR assay can be applied as a practical diagnostic method for epidemiological research and the sanitary management of seeds and seedlings with a low level or latent infection of C. michiganensis subsp. michiganensis.

Rapid Assessment of the Viability of Mycobacterium avium subsp. paratuberculosis Cells after Heat Treatment, Using an Optimized Phage Amplification Assay▿

PubMed Central

Foddai, Antonio; Elliott, Christopher T.; Grant, Irene R.

2010-01-01

Thermal inactivation experiments were carried out to assess the utility of a recently optimized phage amplification assay to accurately enumerate viable Mycobacterium avium subsp. paratuberculosis cells in milk. Ultra-heat-treated (UHT) whole milk was spiked with large numbers of M. avium subsp. paratuberculosis organisms (106 to 107 CFU/ml) and dispensed in 100-μl aliquots in thin-walled 200-μl PCR tubes. A Primus 96 advanced thermal cycler (Peqlab, Erlangen, Germany) was used to achieve the following time and temperature treatments: (i) 63°C for 3, 6, and 9 min; (ii) 68°C for 20, 40, and 60 s; and (iii) 72°C for 5, 10, 15, and 25 s. After thermal stress, the number of surviving M. avium subsp. paratuberculosis cells was assessed by both phage amplification assay and culture on Herrold's egg yolk medium (HEYM). A high correlation between PFU/ml and CFU/ml counts was observed for both unheated (r2 = 0.943) and heated (r2 = 0.971) M. avium subsp. paratuberculosis cells. D and z values obtained using the two types of counts were not significantly different (P > 0.05). The D68°C, mean D63°C, and D72°C for four M. avium subsp. paratuberculosis strains were 81.8, 9.8, and 4.2 s, respectively, yielding a mean z value of 6.9°C. Complete inactivation of 106 to 107 CFU of M. avium subsp. paratuberculosis/ml milk was not observed for any of the time-temperature combinations studied; 5.2- to 6.6-log10 reductions in numbers were achieved depending on the temperature and time. Nonlinear thermal inactivation kinetics were consistently observed for this bacterium. This study confirms that the optimized phage assay can be employed in place of conventional culture on HEYM to speed up the acquisition of results (48 h instead of a minimum of 6 weeks) for inactivation experiments involving M. avium subsp. paratuberculosis-spiked samples. PMID:20097817

Comparative Genomic Analyses of Clavibacter michiganensis subsp. insidiosus and Pathogenicity on Medicago truncatula.

PubMed

Lu, You; Ishimaru, Carol A; Glazebrook, Jane; Samac, Deborah A

2018-02-01

Clavibacter michiganensis is the most economically important gram-positive bacterial plant pathogen, with subspecies that cause serious diseases of maize, wheat, tomato, potato, and alfalfa. Much less is known about pathogenesis involving gram-positive plant pathogens than is known for gram-negative bacteria. Comparative genome analyses of C. michiganensis subspecies affecting tomato, potato, and maize have provided insights on pathogenicity. In this study, we identified strains of C. michiganensis subsp. insidiosus with contrasting pathogenicity on three accessions of the model legume Medicago truncatula. We generated complete genome sequences for two strains and compared these to a previously sequenced strain and genome sequences of four other subspecies. The three C. michiganensis subsp. insidiosus strains varied in gene content due to genome rearrangements, most likely facilitated by insertion elements, and plasmid number, which varied from one to three depending on strain. The core C. michiganensis genome consisted of 1,917 genes, with 379 genes unique to C. michiganensis subsp. insidiosus. An operon for synthesis of the extracellular blue pigment indigoidine, enzymes for pectin degradation, and an operon for inositol metabolism are among the unique features. Secreted serine proteases belonging to both the pat-1 and ppa families were present but highly diverged from those in other subspecies.

Geography of Genetic Structure in Barley Wild Relative Hordeum vulgare subsp. spontaneum in Jordan

PubMed Central

Reeves, Patrick; Reilley, Ann; Engels, Johannes M. M.; Lohwasser, Ulrike; Börner, Andreas; Pillen, Klaus; Richards, Christopher M.

2016-01-01

Informed collecting, conservation, monitoring and utilization of genetic diversity requires knowledge of the distribution and structure of the variation occurring in a species. Hordeum vulgare subsp. spontaneum (K. Koch) Thell., a primary wild relative of barley, is an important source of genetic diversity for barley improvement and co-occurs with the domesticate within the center of origin. We studied the current distribution of genetic diversity and population structure in H. vulgare subsp. spontaneum in Jordan and investigated whether it is correlated with either spatial or climatic variation inferred from publically available climate layers commonly used in conservation and ecogeographical studies. The genetic structure of 32 populations collected in 2012 was analyzed with 37 SSRs. Three distinct genetic clusters were identified. Populations were characterized by admixture and high allelic richness, and genetic diversity was concentrated in the northern part of the study area. Genetic structure, spatial location and climate were not correlated. This may point out a limitation in using large scale climatic data layers to predict genetic diversity, especially as it is applied to regional genetic resources collections in H. vulgare subsp. spontaneum. PMID:27513459

Peritonitis in a llama caused by Streptococcus equi subsp. zooepidemicus.

PubMed Central

Hewson, J; Cebra, C K

2001-01-01

A 7-month-old, male llama was diagnosed with peritonitis caused by Streptococcus equi subsp. zooepidemicus. Clinical findings, medical treatment, and case outcome are described. Hematogenous dissemination from suspected pneumonia is proposed as the route of infection in this case. Possible transmission of the organism through contact with horses is discussed. PMID:11424579

The Genome Sequence of the Tomato-Pathogenic Actinomycete Clavibacter michiganensis subsp. michiganensis NCPPB382 Reveals a Large Island Involved in Pathogenicity▿ †

PubMed Central

Gartemann, Karl-Heinz; Abt, Birte; Bekel, Thomas; Burger, Annette; Engemann, Jutta; Flügel, Monika; Gaigalat, Lars; Goesmann, Alexander; Gräfen, Ines; Kalinowski, Jörn; Kaup, Olaf; Kirchner, Oliver; Krause, Lutz; Linke, Burkhard; McHardy, Alice; Meyer, Folker; Pohle, Sandra; Rückert, Christian; Schneiker, Susanne; Zellermann, Eva-Maria; Pühler, Alfred; Eichenlaub, Rudolf; Kaiser, Olaf; Bartels, Daniela

2008-01-01

Clavibacter michiganensis subsp. michiganensis is a plant-pathogenic actinomycete that causes bacterial wilt and canker of tomato. The nucleotide sequence of the genome of strain NCPPB382 was determined. The chromosome is circular, consists of 3.298 Mb, and has a high G+C content (72.6%). Annotation revealed 3,080 putative protein-encoding sequences; only 26 pseudogenes were detected. Two rrn operons, 45 tRNAs, and three small stable RNA genes were found. The two circular plasmids, pCM1 (27.4 kbp) and pCM2 (70.0 kbp), which carry pathogenicity genes and thus are essential for virulence, have lower G+C contents (66.5 and 67.6%, respectively). In contrast to the genome of the closely related organism Clavibacter michiganensis subsp. sepedonicus, the genome of C. michiganensis subsp. michiganensis lacks complete insertion elements and transposons. The 129-kb chp/tomA region with a low G+C content near the chromosomal origin of replication was shown to be necessary for pathogenicity. This region contains numerous genes encoding proteins involved in uptake and metabolism of sugars and several serine proteases. There is evidence that single genes located in this region, especially genes encoding serine proteases, are required for efficient colonization of the host. Although C. michiganensis subsp. michiganensis grows mainly in the xylem of tomato plants, no evidence for pronounced genome reduction was found. C. michiganensis subsp. michiganensis seems to have as many transporters and regulators as typical soil-inhabiting bacteria. However, the apparent lack of a sulfate reduction pathway, which makes C. michiganensis subsp. michiganensis dependent on reduced sulfur compounds for growth, is probably the reason for the poor survival of C. michiganensis subsp. michiganensis in soil. PMID:18192381

Draft genome sequence of Xylella fastidiosa subsp. fastidiosa strain Stag’s Leap

USDA-ARS?s Scientific Manuscript database

Xylella fastidiosa subsp. fastidiosa causes Pierce’s disease of grapevine. Presented here is the draft genome sequence of the Stag’s Leap strain, previously used in pathogenicity/virulence assays to evaluate grapevine germplasm bearing Pierce’s disease....

Bartonella vinsonii subsp. berkhoffii endocarditis in a dog from Saskatchewan

PubMed Central

Cockwill, Ken R.; Taylor, Susan M.; Philibert, Helene M.; Breitschwerdt, Edward B.; Maggi, Ricardo G.

2007-01-01

A dog referred for lameness was diagnosed with culture-negative endocarditis. Antibodies to Bartonella spp. were detected. Antibiotic treatment resulted in transient clinical improvement, but the dog developed cardiac failure and was euthanized. Bartonella vinsonii subsp. berkhoffii genotype IV was identified within the aortic heart valve lesions by PCR amplification and DNA sequencing. PMID:17824328

In Silico Identification of Epitopes in Mycobacterium avium subsp. paratuberculosis Proteins That Were Upregulated under Stress Conditions

PubMed Central

Gurung, Ratna B.; Purdie, Auriol C.; Begg, Douglas J.

2012-01-01

Johne's disease in ruminants is caused by Mycobacterium avium subsp. paratuberculosis. Diagnosis of M. avium subsp. paratuberculosis infection is difficult, especially in the early stages. To date, ideal antigen candidates are not available for efficient immunization or immunodiagnosis. This study reports the in silico selection and subsequent analysis of epitopes of M. avium subsp. paratuberculosis proteins that were found to be upregulated under stress conditions as a means to identify immunogenic candidate proteins. Previous studies have reported differential regulation of proteins when M. avium subsp. paratuberculosis is exposed to stressors which induce a response similar to dormancy. Dormancy may be involved in evading host defense mechanisms, and the host may also mount an immune response against these proteins. Twenty-five M. avium subsp. paratuberculosis proteins that were previously identified as being upregulated under in vitro stress conditions were analyzed for B and T cell epitopes by use of the prediction tools at the Immune Epitope Database and Analysis Resource. Major histocompatibility complex class I T cell epitopes were predicted using an artificial neural network method, and class II T cell epitopes were predicted using the consensus method. Conformational B cell epitopes were predicted from the relevant three-dimensional structure template for each protein. Based on the greatest number of predicted epitopes, eight proteins (MAP2698c [encoded by desA2], MAP2312c [encoded by fadE19], MAP3651c [encoded by fadE3_2], MAP2872c [encoded by fabG5_2], MAP3523c [encoded by oxcA], MAP0187c [encoded by sodA], and the hypothetical proteins MAP3567 and MAP1168c) were identified as potential candidates for study of antibody- and cell-mediated immune responses within infected hosts. PMID:22496492

Effective heat inactivation of Mycobacterium avium subsp. paratuberculosis in raw milk contaminated with naturally infected feces.

PubMed

Rademaker, Jan L W; Vissers, Marc M M; Te Giffel, Meike C

2007-07-01

The effectiveness of high-temperature, short holding time (HTST) pasteurization and homogenization with respect to inactivation of Mycobacterium avium subsp. paratuberculosis was evaluated quantitatively. This allowed a detailed determination of inactivation kinetics. High concentrations of feces from cows with clinical symptoms of Johne's disease were used to contaminate raw milk in order to realistically mimic possible incidents most closely. Final M. avium subsp. paratuberculosis concentrations varying from 10(2) to 3.5 x 10(5) cells per ml raw milk were used. Heat treatments including industrial HTST were simulated on a pilot scale with 22 different time-temperature combinations, including 60 to 90 degrees C at holding (mean residence) times of 6 to 15 s. Following 72 degrees C and a holding time of 6 s, 70 degrees C for 10 and 15 s, or under more stringent conditions, no viable M. avium subsp. paratuberculosis cells were recovered, resulting in >4.2- to >7.1-fold reductions, depending on the original inoculum concentrations. Inactivation kinetic modeling of 69 quantitative data points yielded an E(a) of 305,635 J/mol and an lnk(0) of 107.2, corresponding to a D value of 1.2 s at 72 degrees C and a Z value of 7.7 degrees C. Homogenization did not significantly affect the inactivation. The conclusion can be drawn that HTST pasteurization conditions equal to 15 s at > or =72 degrees C result in a more-than-sevenfold reduction of M. avium subsp. paratuberculosis.

Effective Heat Inactivation of Mycobacterium avium subsp. paratuberculosis in Raw Milk Contaminated with Naturally Infected Feces▿

PubMed Central

Rademaker, Jan L. W.; Vissers, Marc M. M.; te Giffel, Meike C.

2007-01-01

The effectiveness of high-temperature, short holding time (HTST) pasteurization and homogenization with respect to inactivation of Mycobacterium avium subsp. paratuberculosis was evaluated quantitatively. This allowed a detailed determination of inactivation kinetics. High concentrations of feces from cows with clinical symptoms of Johne's disease were used to contaminate raw milk in order to realistically mimic possible incidents most closely. Final M. avium subsp. paratuberculosis concentrations varying from 102 to 3.5 × 105 cells per ml raw milk were used. Heat treatments including industrial HTST were simulated on a pilot scale with 22 different time-temperature combinations, including 60 to 90°C at holding (mean residence) times of 6 to 15 s. Following 72°C and a holding time of 6 s, 70°C for 10 and 15 s, or under more stringent conditions, no viable M. avium subsp. paratuberculosis cells were recovered, resulting in >4.2- to >7.1-fold reductions, depending on the original inoculum concentrations. Inactivation kinetic modeling of 69 quantitative data points yielded an Ea of 305,635 J/mol and an lnk0 of 107.2, corresponding to a D value of 1.2 s at 72°C and a Z value of 7.7°C. Homogenization did not significantly affect the inactivation. The conclusion can be drawn that HTST pasteurization conditions equal to 15 s at ≥72°C result in a more-than-sevenfold reduction of M. avium subsp. paratuberculosis. PMID:17496131

Envelope protein complexes of Mycobacterium avium subsp. paratuberculosis and their antigenicity

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne’s disease, a chronic enteric disease of ruminant animals. In the present study, blue native PAGE electrophoresis and 2D SDS-PAGE were used to separate MAP envelope protein complexes, followed by mass spectrometry (MS) ...

Draft Genome Sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a Citrate-Fermenting Strain

PubMed Central

Zuljan, Federico; Espariz, Martín; Blancato, Victor S.; Esteban, Luis; Alarcón, Sergio

2016-01-01

We report the draft genome sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a natural strain isolated from artisanal cheese from northwest Argentina. L. lactis subsp. lactis bv. diacetylactis is one of the most important microorganisms used as starter culture around the world. The CRL264 strain constitutes a model microorganism in the studies on the generation of aroma compounds (diacetyl, acetoin, and 2,3-butanediol) by lactic acid bacteria. Our genome analysis shows similar genetic organization to other available genomes of L. lactis bv. diacetylactis strains. PMID:26847906

Composition and potency characterization of Mycobacterium avium subsp. paratuberculosis purified protein derivatives

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) purified protein derivatives (PPDs) are immunologic reagents prepared from cultured filtrates of the type strain ATCC 19698. Traditional production consists of floating culture incubation at 37oC, organism inactivation by autoclaving, coarse filtrat...

Survival of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus in the Terminal Ileum of Fistulated Göttingen Minipigs

PubMed Central

Lick, Sonja; Drescher, Karsten; Heller, Knut J.

2001-01-01

The ability of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus administered in yogurt to survive the passage through the upper gastrointestinal tract was investigated with Göttingen minipigs that were fitted with ileum T-cannulas. After ingestion of yogurt containing viable microorganisms, ileostomy samples were collected nearly every hour beginning 3 h after food uptake. Living L. delbrueckii subsp. bulgaricus and S. thermophilus were detected in the magnitude of 106 to 107 per gram of intestinal contents (wet weight) in all animals under investigation. A calculation of the minimum amount of surviving bacteria that had been administered is presented. Total DNA extracted from ileostomy samples was subjected to PCR, which was species specific for L. delbrueckii and S. thermophilus and subspecies specific for L. delbrueckii subsp. bulgaricus. All three bacterial groups could be detected by PCR after yogurt uptake but not after uptake of a semisynthetic diet. One pig apparently had developed an endogenous L. delbrueckii flora. When heat-treated yogurt was administered, L. delbrueckii was detected in all animals. S. thermophilus or L. delbrueckii subsp. bulgaricus was not detected, indicating that heat-inactivated cells and their DNAs had already been digested and their own L. delbrueckii flora had been stimulated for growth. PMID:11526016

Cytogenetic characterization of Amaranthus caudatus L. and Amaranthus hybridus subsp. cruentus (L.) Thell.

PubMed

Prajitha, V; Thoppil, J E

2018-02-01

The present study is aimed to identify genetic variability between two species of Amaranthus viz., A. caudatus and A. hybridus subsp. cruentus, two economically important species, cultivated mainly for grain production. Karyomorphological studies in Amaranthus are scarce, probably due to higher number of small sized chromosomes. Karyomorphological studies were conducted using mitotic squash preparation of young healthy root tips. Karyological parameters and karyotypic formula were established using various software programs and tabulated the karyomorphometric and asymmetry indices viz., Disparity index, Variation coefficient, Total forma percentage, Karyotype asymmetry index, Syi index, Rec index, Interchromosomal and Intrachromosomal asymmetry index and Degree of asymmetry of karyotypes. The mitotic chromosome number observed for A. caudatus was 2n = 32 with a gametic number n = 16 and A. hybridus subsp. cruentus was 2n = 34 with a gametic number n = 17. In A. caudatus the chromosome length during somatic metaphase ranged from 0.8698 to 1.7722 μm with a total length of 39.1412 μm. In A. hybridus subsp. cruentus the length of chromosome ranged from 0.7756 to 1.9421 μm with a total length of 44.9922 μm. Various karyomorphometry and asymmetry indices analyzed revealed the extend of interspecific variation and their evolutionary status.

Stawamycin analog, JBIR-11 from Streptomyces viridochromogenes subsp. sulfomycini NBRC 13830.

PubMed

Izumikawa, Miho; Komaki, Hisayuki; Hashimoto, Junko; Takagi, Motoki; Shin-ya, Kazuo

2008-05-01

A stawamycin analog, JBIR-11 (1) was isolated from mycelium of Streptomyces viridochromogenes subsp. sulfomycini NBRC 13830. The structure was determined on the basis of the spectroscopic data. Compound 1 exhibited growth inhibitory effect against human fibrosarcoma HT1080 cells with an IC50 value of 25 microM.

H(+) -ATPase-defective variants of Lactobacillus delbrueckii subsp. bulgaricus contribute to inhibition of postacidification of yogurt during chilled storage.

PubMed

Wang, Xinhui; Ren, Hongyang; Liu, Dayu; Wang, Bing; Zhu, Wenyou; Wang, Wei

2013-02-01

Continued acid production by Lactobacillus delbrueckii subsp. bulgaricus during the chilled storage of yogurt is the major cause of postacidification, resulting in a short shelf life. Two H(+) -ATPase defective variants of L. delbrueckii subsp. bulgaricus were successfully isolated and their H(+) -ATPase activities were reduced by 51.3% and 34.3%, respectively. It was shown that growth and acid production of variants were remarkably inhibited. The variants were more sensitive to acidic condition and had a significant rate for inactivation of H(+) -ATPase by N, N-dicyclohexylcarbodiimide (DCCD), along with a low H(+) -extrusion, suggesting that H(+) -ATPase is direct response for H(+) -extrusion. In addition, the variants were also more sensitive to NaCl, while H(+) -ATPase activities of variants and parent strain were significantly enhanced by NaCl stress. Obviously, H(+) -ATPase might be involved in Na(+) transportation. Furthermore, variants were inoculated in fermented milk to ferment yogurt. There was no significant difference in flavor, whereas the postacidification of yogurt during chilled storage was remarkably inhibited. It is suggested that application of L. delbrueckii subsp. bulgaricus with reduced H(+) -ATPase activity in yogurt fermentation is one of effect, economic and simple avenues of inhibiting postacidification of yogurt during refrigerated storage, giving a longer shelf life. During yogurt fermentation, continued acid production by Lactobacillus delbrueckii subsp. bulgaricus during the chilled storage of yogurt leads to milk fermentation with high postacidification, resulting in a short shelf life. In this work, 2 acid-sensitive variant strains of L. delbrueckii subsp. bulgaricus were isolated. The characteristics related to H(+) -ATPase were compared and it was observed that milk fermented by the variants had lower postacidification, giving a longer shelf life. Application of L. delbrueckii subsp. bulgaricus with reduced H(+) -ATPase activity

Profiles of Volatile Flavor Compounds in Milk Fermented with Different Proportional Combinations of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus.

PubMed

Dan, Tong; Wang, Dan; Wu, Shimei; Jin, Rulin; Ren, Weiyi; Sun, Tiansong

2017-09-29

Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus are key factors in the fermentation process and the final quality of dairy products worldwide. This study was performed to investigate the effects of the proportions of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus isolated from traditionally fermented dairy products in China and Mongolia on the profile of volatile compounds produced in samples. Six proportional combinations (1:1, 1:10, 1:50, 1:100, 1:1000, and 1:10,000) of L. delbrueckii subsp. bulgaricus IMAU20401 to S. thermophilus ND03 were considered, and the volatiles were identified and quantified by solid-phase microextraction and gas chromatography-mass spectrometry (SPME-GC-MS) against an internal standard. In total, 89 volatile flavor compounds, consisting of aldehydes, ketones, acids, alcohols, esters, and aromatic hydrocarbons, were identified. Among these, some key flavor volatile compounds were identified, including acetaldehyde, 3-methylbutanal, acetoin, 2-heptanone, acetic acid, butanoic acid, and 3-methyl-1-butanol. The of L. delbrueckii subsp. bulgaricus IMAU20401 to S. thermophilus ND03 influenced the type and concentration of volatiles produced. In particular, aldehydes and ketones were present at higher concentrations in the 1:1000 treatment combination than in the other combinations. Our findings emphasize the importance of selecting the appropriate proportions of L. delbrueckii subsp. bulgaricus and S. thermophilus for the starter culture in determining the final profile of volatiles and the overall flavor of dairy products.

[The occurrence of campylobacter fetus subsp. jejuni and Salmonella bacteria in some wild birds (author's transl)].

PubMed

Rosef, O

1981-12-01

An investigation was carried out into the occurrence of Campylobacter fetus subsp. jejuni and Salmonella species in some wild birds. A total of 129 birds was examined, consisting of 71 pigeons, 54 seagulls, three crows and one raven. Campylobacter bacteria were isolated from 32 birds (24.8%), of which three were pigeons, 27 seagulls and two were crows. Of the 27 Campylobacter strains isolated from seagulls, four had the biochemical characteristics of the NARTC biotype described by Skirrow and Benjamin, seven were grouped as Campylobacter coli biotype and 16 as the biotype of Campylobacter jejuni. All the strains isolated from crows and pigeons had the biochemical characteristics of Campylobacter jejuni biotypes. Salmonella bacteria were isolated from the intestinal contents of two of the 54 seagulls (3.7%), and were identified serologically as Salmonella indiana and Salmonella typhimurium. One seagull was found to be a carrier of both Campylobacter fetus subsp. jejuni and Salmonella typhimurium. A correlation could not be demonstrated between the occurrence of Salmonella bacteria and Campylobacter fetus subsp. jejuni.

Specific Discrimination of Three Pathogenic Salmonella enterica subsp. enterica Serotypes by carB-Based Oligonucleotide Microarray

PubMed Central

Shin, Hwa Hui; Hwang, Byeong Hee; Seo, Jeong Hyun

2014-01-01

It is important to rapidly and selectively detect and analyze pathogenic Salmonella enterica subsp. enterica in contaminated food to reduce the morbidity and mortality of Salmonella infection and to guarantee food safety. In the present work, we developed an oligonucleotide microarray containing duplicate specific capture probes based on the carB gene, which encodes the carbamoyl phosphate synthetase large subunit, as a competent biomarker evaluated by genetic analysis to selectively and efficiently detect and discriminate three S. enterica subsp. enterica serotypes: Choleraesuis, Enteritidis, and Typhimurium. Using the developed microarray system, three serotype targets were successfully analyzed in a range as low as 1.6 to 3.1 nM and were specifically discriminated from each other without nonspecific signals. In addition, the constructed microarray did not have cross-reactivity with other common pathogenic bacteria and even enabled the clear discrimination of the target Salmonella serotype from a bacterial mixture. Therefore, these results demonstrated that our novel carB-based oligonucleotide microarray can be used as an effective and specific detection system for S. enterica subsp. enterica serotypes. PMID:24185846

Specific discrimination of three pathogenic Salmonella enterica subsp. enterica serotypes by carB-based oligonucleotide microarray.

PubMed

Shin, Hwa Hui; Hwang, Byeong Hee; Seo, Jeong Hyun; Cha, Hyung Joon

2014-01-01

It is important to rapidly and selectively detect and analyze pathogenic Salmonella enterica subsp. enterica in contaminated food to reduce the morbidity and mortality of Salmonella infection and to guarantee food safety. In the present work, we developed an oligonucleotide microarray containing duplicate specific capture probes based on the carB gene, which encodes the carbamoyl phosphate synthetase large subunit, as a competent biomarker evaluated by genetic analysis to selectively and efficiently detect and discriminate three S. enterica subsp. enterica serotypes: Choleraesuis, Enteritidis, and Typhimurium. Using the developed microarray system, three serotype targets were successfully analyzed in a range as low as 1.6 to 3.1 nM and were specifically discriminated from each other without nonspecific signals. In addition, the constructed microarray did not have cross-reactivity with other common pathogenic bacteria and even enabled the clear discrimination of the target Salmonella serotype from a bacterial mixture. Therefore, these results demonstrated that our novel carB-based oligonucleotide microarray can be used as an effective and specific detection system for S. enterica subsp. enterica serotypes.

Complete genome sequence of Polynucleobacter necessarius subsp. asymbioticus type strain (QLW-P1DMWA-1T)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Meincke, Linda; Copeland, A; Lapidus, Alla L.

2012-01-01

Polynucleobacter necessarius subsp. asymbioticus Hahn et al. 2009 is one of currently two subspecies of P. necessarius. While P. necessarius subsp. asymbioticus is a free-living bacterium, the closely related second subspecies, P. necessarius subsp. necessarius is an obligate endosymbiont living in the cytoplasm of freshwater ciliates of the genus Euplotes aediculatus. The two P. necessarius subspecies were the closest thus far reported phylogenetic neighbors that differ in their lifestyle as obligately free-living vs. obligate endosymbiontic, and they are the only members of the genus Polynucleobacter with completely sequenced genomes. The genome-sequenced strain represents a group of closely related strains notmore » distinguishable by 16S rRNA, 16S-23S ITS or glnA sequences, which is persistent in the home habitat of the strain and frequently contributes > 10% of total bacterial numbers in water samples of the habitat. The 2,159,490 bp long chromosome with a total of 2,088 protein-coding and 48 RNA genes was sequenced as part of the DOE Joint Genome Institute Community Sequencing Program 2006.« less

Morphological leaf variability in natural populations of Pistacia atlantica Desf. subsp. atlantica along climatic gradient: new features to update Pistacia atlantica subsp. atlantica key.

PubMed

El Zerey-Belaskri, Asma; Benhassaini, Hachemi

2016-04-01

The effect of bioclimate range on the variation in Pistacia atlantica Desf. subsp. atlantica leaf morphology was studied on 16 sites in Northwest Algeria. The study examined biometrically mature leaves totaling 3520 compound leaves. Fifteen characters (10 quantitative and 5 qualitative) were assessed on each leaf. For each quantitative character, the nested analysis of variance (ANOVA) was used to examine relative magnitude of variation at each level of the nested hierarchy. The correlation between the climatic parameters and the leaf morphology was examined. The statistical analysis applied on the quantitative leaf characters showed highly significant variation at the within-site level and between-site variation. The correlation coefficient (r) showed also an important correlation between climatic parameters and leaf morphology. The results of this study exhibited several values reported for the first time on the species, such as the length and the width of the leaf (reaching up to 24.5 cm/21.9 cm), the number of leaflets (up to 18 leaflets/leaf), and the petiole length of the terminal leaflet (reaching up to 3.4 cm). The original findings of this study are used to update the P. atlantica subsp. atlantica identification key.

Complete genome sequence of Campylobacter fetus subsp. testudinum type strain 03-427T

USDA-ARS?s Scientific Manuscript database

Campylobacter fetus subsp. testudinum has been isolated from reptiles and humans. This Campylobacter subspecies is genetically distinct from other C. fetus subspecies. Here we present the first whole genome sequence for this C. fetus subspecies....

Quorum sensing in the plant pathogen Erwinia carotovora subsp. carotovora: the role of expR(Ecc).

PubMed

Andersson, R A; Eriksson, A R; Heikinheimo, R; Mäe, A; Pirhonen, M; Kõiv, V; Hyytiäinen, H; Tuikkala, A; Palva, E T

2000-04-01

The production of the main virulence determinants of the plant pathogen Erwinia carotovora subsp. carotovora, the extracellular cell wall-degrading enzymes, is partly controlled by the diffusible signal molecule N-(3-oxohexanoyl)-L-homoserine lactone (OHHL). OHHL is synthesized by the product of the expI/carI gene. Linked to expI we found a gene encoding a putative transcriptional regulator of the LuxR-family. This gene, expR(Ecc), is transcribed convergently to the expI gene and the two open reading frames are partially overlapping. The ExpR(Ecc) protein showed extensive amino acid sequence similarity to the repressor EsaR from Pantoea stewartii subsp. stewartii (formerly Erwinia stewartii subsp. stewartii) and to the ExpR(Ech) protein of Erwinia chrysanthemi. Inactivation of the E. carotovora subsp. carotovora expR(Ecc) gene caused no decrease in virulence or production of virulence determinants in vitro. In contrast, there was a slight increase in the maceration capacity of the mutant strain. The effects of ExpR(Ecc) were probably mediated by changes in OHHL levels. Inactivation of expR(Ecc) resulted in increased OHHL levels during early logarithmic growth. In addition, overexpression of expR(Ecc) caused a clear decrease in the production of virulence determinants and part of this effect was likely to be caused by OHHL binding to ExpR(Ecc). ExpR(Ecc) did not appear to exhibit transcriptional regulation of expI, but the effect on OHHL was apparently due to other mechanisms.

Geobacter sulfurreducens subsp. ethanolicus, subsp. nov., an ethanol-utilizing dissimilatory Fe(III)-reducing bacterium from a lotus field.

PubMed

Viulu, Samson; Nakamura, Kohei; Kojima, Akihiro; Yoshiyasu, Yuki; Saitou, Sakiko; Takamizawa, Kazuhiro

2013-01-01

An ethanol-utilizing Fe(III)-reducing bacterial strain, OSK2A(T), was isolated from a lotus field in Aichi, Japan. Phylogenetic analysis of the 16S rRNA gene sequences of OSK2A(T) and related strains placed it within Geobacter sulfurreducens PCA(T). Strain OSK2A(T) was shown to be a Gram-negative, motile, rod-shaped bacterium, strictly anaerobic, 0.76-1.65 µm long and 0.28-0.45 μm wide. Its growth occurred at 20-40℃, pH 6.0-8.1, and it tolerated up to 1% NaCl. The G+C content of the genomic DNA was 61.2 mol% and DNA-DNA hybridization value with Geobacter sulfurreducens PCA(T) was 60.7%. The major respiratory quinone was MK-8. The major fatty acids were 16：1 ω7c, 16：0, 14：0, 15：0 iso, 16：1 ω5c, and 18：1 ω7c. Strain OSK2A(T) could utilize H2, ethanol, acetate, lactate, pyruvate, and formate as substrates with Fe(III)-citrate as electron acceptor. Amorphous Fe(III) hydroxide, Fe(III)-NTA, fumarate, malate, and elemental sulfur were utilized as electron acceptors with either acetate or ethanol as substrates. Results obtained from physiological, DNA-DNA hybridization, and chemotaxonomic tests support genotypic and phenotypic differentiation of strain OSK2A(T) from its closest relative. The isolate is assigned as a novel subspecies with the name Geobacter sulfurreducens subsp. ethanolicus, subsp. nov. (type strain OSK2A(T)=DSMZ 26126(T)=JCM 18752(T)).

Draft Genome Sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a Citrate-Fermenting Strain.

PubMed

Zuljan, Federico; Espariz, Martín; Blancato, Victor S; Esteban, Luis; Alarcón, Sergio; Magni, Christian

2016-02-04

We report the draft genome sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a natural strain isolated from artisanal cheese from northwest Argentina. L. lactis subsp. lactis bv. diacetylactis is one of the most important microorganisms used as starter culture around the world. The CRL264 strain constitutes a model microorganism in the studies on the generation of aroma compounds (diacetyl, acetoin, and 2,3-butanediol) by lactic acid bacteria. Our genome analysis shows similar genetic organization to other available genomes of L. lactis bv. diacetylactis strains. Copyright © 2016 Zuljan et al.

Environmental contamination with Mycobacterium avium subsp. paratuberculosis in endemically infected dairy herds

USDA-ARS?s Scientific Manuscript database

Environmental contamination with Mycobacterium avium subsp. paratuberculosis (MAP) is thought to be the primary source of infection for dairy cattle. The exact link between fecal shedding of MAP by individual cows and environmental contamination levels at the herd level was explored with a cross-se...

A 38-Kilobase Pathogenicity Island Specific for Mycobacterium avium subsp. paratuberculosis Encodes Cell Surface Proteins Expressed in the Host

PubMed Central

Stratmann, Janin; Strommenger, Birgit; Goethe, Ralph; Dohmann, Karen; Gerlach, Gerald-F.; Stevenson, Karen; Li, Ling-ling; Zhang, Qing; Kapur, Vivek; Bull, Tim J.

2004-01-01

We have used representational difference analysis to identify a novel Mycobacterium avium subsp. paratuberculosis-specific ABC transporter operon (mpt), which comprises six open reading frames designated mptA to -F and is immediately preceded by two putative Fur boxes. Functional genomics revealed that the mpt operon is flanked on one end by a fep cluster encoding proteins involved in the uptake of Fe3+ and on the other end by a sid cluster encoding non-ribosome-dependent heterocyclic siderophore synthases. Together these genes form a 38-kb M. avium subsp. paratuberculosis-specific locus flanked by an insertion sequence similar to IS1110. Expression studies using Western blot analyses showed that MptC is present in the envelope fraction of M. avium subsp. paratuberculosis. The MptD protein was shown to be surface exposed, using a specific phage (fMptD) isolated from a phage-peptide library, by differential screening of Mycobacterium smegmatis transformants. The phage fMptD-derived peptide could be used in a peptide-mediated capture PCR with milk from infected dairy herds, thereby showing surface-exposed expression of the MptD protein in the host. Together, these data suggest that the 38-kb locus constitutes an M. avium subsp. paratuberculosis pathogenicity island. PMID:14977927

Prophage Lysin Ply30 Protects Mice from Streptococcus suis and Streptococcus equi subsp. zooepidemicus Infections

PubMed Central

Tang, Fang; Li, Dezhi; Wang, Haojin; Ma, Zhe; Lu, Chengping

2015-01-01

Streptococcus suis and Streptococcus equi subsp. zooepidemicus are capable of infecting humans and various animals, causing significant problems for the worldwide swine industry. As antibiotic resistance has increased, lysosomal enzymes encoded by phages have shown potential for use against pathogenic bacteria. In this study, a novel bacteriophage lysin, Ply30, encoded by the S. suis prophage phi30c, was recombinantly expressed and purified. Ply30 showed high bacteriolysis activity on S. suis and S. equi subsp. zooepidemicus in vitro. The ratio of the optical density at 600 nm (OD600) with treatment versus the OD600 with no treatment for most tested S. suis and S. equi subsp. zooepidemicus strains decreased from 1 to <0.3 and <0.5, respectively, within 1 h. The results of plate viability assays showed that treated bacteria suffered a 1- to 2-log decrease in CFU within 1 h. The optimal concentration of Ply30 was 50 μg/ml, and the optimal pH was 7. Moreover, Ply30 maintained high activity over a wide pH range (pH 6 to 10). The MICs of Ply30 against Streptococcus strains ranged from 16 to 512 μg/ml. In vivo, a 2-mg dose of Ply30 protected 90% (9/10 mice) of mice from infection with S. equi subsp. zooepidemicus and 80% (8/10 mice) of mice from infection with S. suis. Seven days after lysin Ply30 treatment, bacterial loads were significantly decreased in all tested organs and blood compared with those at 1 h postinfection without Ply30 treatment. Ply30 showed in vitro and in vivo antimicrobial efficiency and protected mice against two kinds of bacterial infections, indicating that Ply30 may be an effective therapeutic against streptococci. PMID:26253669

Prophage lysin Ply30 protects mice from Streptococcus suis and Streptococcus equi subsp. zooepidemicus infections.

PubMed

Tang, Fang; Li, Dezhi; Wang, Haojin; Ma, Zhe; Lu, Chengping; Dai, Jianjun

2015-11-01

Streptococcus suis and Streptococcus equi subsp. zooepidemicus are capable of infecting humans and various animals, causing significant problems for the worldwide swine industry. As antibiotic resistance has increased, lysosomal enzymes encoded by phages have shown potential for use against pathogenic bacteria. In this study, a novel bacteriophage lysin, Ply30, encoded by the S. suis prophage phi30c, was recombinantly expressed and purified. Ply30 showed high bacteriolysis activity on S. suis and S. equi subsp. zooepidemicus in vitro. The ratio of the optical density at 600 nm (OD600) with treatment versus the OD600 with no treatment for most tested S. suis and S. equi subsp. zooepidemicus strains decreased from 1 to <0.3 and <0.5, respectively, within 1 h. The results of plate viability assays showed that treated bacteria suffered a 1- to 2-log decrease in CFU within 1 h. The optimal concentration of Ply30 was 50 μg/ml, and the optimal pH was 7. Moreover, Ply30 maintained high activity over a wide pH range (pH 6 to 10). The MICs of Ply30 against Streptococcus strains ranged from 16 to 512 μg/ml. In vivo, a 2-mg dose of Ply30 protected 90% (9/10 mice) of mice from infection with S. equi subsp. zooepidemicus and 80% (8/10 mice) of mice from infection with S. suis. Seven days after lysin Ply30 treatment, bacterial loads were significantly decreased in all tested organs and blood compared with those at 1 h postinfection without Ply30 treatment. Ply30 showed in vitro and in vivo antimicrobial efficiency and protected mice against two kinds of bacterial infections, indicating that Ply30 may be an effective therapeutic against streptococci. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Genome-wide transcriptome analysis of Clavibacter michiganensis subsp. michiganensis grown in xylem mimicking medium.

PubMed

Hiery, Eva; Adam, Susanne; Reid, Stephen; Hofmann, Jörg; Sonnewald, Sophia; Burkovski, Andreas

2013-12-01

The interaction between Clavibacter michiganensis subsp. michiganensis with its host, the tomato plant (Solanum lycopersicum), is poorly understood and only few virulence factors are known. While studying of the bacteria in planta is time-consuming and difficult, the analysis in vitro would facilitate research. Therefore, a xylem mimicking medium (XMM) for C. michiganensis subsp. michiganensis was established in this study based on an apoplast medium for Xanthomonas campestris pv. vesicatoria. In contrast to the apoplast medium, XMM contains no sugars, but amino acids which serve as nitrogen and carbon source. As a result, growth in XMM induced transcriptional changes of genes encoding putative sugar, amino acid and iron uptake systems. In summary, mRNA levels of about 8% of all C. michiganensis subsp. michiganensis genes were changed when XMM-grown bacteria were compared to M9 minimal medium-grown cells. Almost no transcriptional changes of genes encoding hydrolytic enzymes were detected, leading to the idea that XMM reflects the situation in the beginning of infection and therefore allows the characterization of virulence factors in this early stage of infection. The addition of the plant wound substance acetosyringone to the XMM medium led to a change in transcript amount, including genes coding for proteins involved in protein transport, iron uptake and regulation processes. Copyright © 2013 Elsevier B.V. All rights reserved.

Isolation and characterization of lactic acid bacteria from pobuzihi (fermented cummingcordia), a traditional fermented food in Taiwan.

PubMed

Chen, Yi-Sheng; Wu, Hui-Chung; Wang, Chiung-Mei; Lin, Chia-Chun; Chen, Yi-Ting; Jhong, Yu-Jyun; Yanagida, Fujitoshi

2013-03-01

Lactobacillus pobuzihii is a novel species which has been previously found in pobuzihi (fermented cummingcordia), a traditional fermented food in Taiwan. However, the lactic acid bacteria (LAB) microflora in pobuzihi has not been studied in detail. In this study, LAB from pobuzihi were isolated, identified, and characterized. A total of 196 LAB were isolated; 79 cultures were isolated from the sample collected from a manufacturing factory, 38 from pobuzihi samples collected from 4 different markets, and 79 from 2 fresh cummingcordia samples. These isolates were characterized phenotypically and then divided into eight groups (A to H) by restriction fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. Lactobacillus plantarum was the most abundant LAB found in most samples during the fermentation of pobuzihi. On the other hand, Enterococcus casseliflavus and Weissella cibaria were, respectively, the major species found in the two fresh cummingcordia samples. A potential novel species or subspecies of lactococcal strain was found. In addition, seven L. plantarum and five W. cibaria strains showed inhibitory activity against the indicator strain Lactobacillus sakei JCM 1157(T). This is the first report describing the distribution and varieties of LAB existing in the pobuzihi during its fermentation process and the final product on the market.

Metagenomic approach reveals microbial diversity and predictive microbial metabolic pathways in Yucha, a traditional Li fermented food

PubMed Central

Zhang, Jiachao; Wang, Xiaoru; Huo, Dongxue; Li, Wu; Hu, Qisong; Xu, Chuanbiao; Liu, Sixin; Li, Congfa

2016-01-01

Yucha is a typical traditional fermented food of the Li population in the Hainan province of China, and it is made up of cooked rice and fresh fish. In the present study, metagenomic approach and culture-dependent technology were applied to describe the diversity of microbiota and identify beneficial microbes in the Yucha. At the genus level, Lactobacillus was the most abundant genus (43.82% of the total reads), followed by Lactococcus, Enterococcus, Vibrio, Weissella, Pediococcus, Enterobacter, Salinivibrio, Acinetobacter, Macrococcus, Kluyvera and Clostridium; this result was confirmed by q-PCR. PCoA based on Weighted UniFrac distances showed an apparent clustering pattern for Yucha samples from different locations, and Lactobacillus sakei, Lactobacillus saniviri and Staphylococcus sciuri represented OTUs according to the major identified markers. At the microbial functional level, it was observed that there was an enrichment of metabolic functional features, including amino acid and carbohydrate metabolism, which implied that the microbial metabolism in the Yucha samples tended to be vigorous. Accordingly, we further investigated the correlation between the predominant microbes and metabolic functional features. Thirteen species of Lactobacillus (147 strains) were isolated, and Lactobacillus plantarum (60 isolates) and Lactobacillus pentosus (34 isolates) were isolated from every sample. PMID:27578483

Gene expression of enzymes involved in utilization of xylooligosaccharides by Lactobacillus strains

PubMed Central

Maria, Ananieva; Margarita, Tzenova; IIlia, Iliev; Iskra, Ivanova

2014-01-01

Prebiotics are defined as food components that confer health benefits on the host through modulation of the microbiota. Xylooligosaccharides (XOS) are non-digestible oligosaccharides that have recently received increasing attention as potential prebiotic candidates. XOS are sugar oligomers composed of 1,4-linked xylopyranosyl backbone and are obtained by either chemical or, more commonly, enzymatic hydrolysis of xylan polysaccharides, extracted from the plant cell wall. The bifidogenic effect of XOS was demonstrated by both in vitro studies and small-scale in vivo human studies. Some intestinal bacterial strains are able to grow on XOS, yet numerous studies have demonstrated that the ability to utilize these oligosaccharides varies considerably among these bacteria. The aim of this study is to investigate the ability of several strains Lactobacillus to use XOS. Fifteen Lactobacillus strains, allifiated to L. plantarum, L. brevis and L. sakei, were studied. Screening procedure was performed for the ability of the strains to utilize XOS as an alternative carbon source. Only some of them utilize XOS. The growth kinetics show the presence of two lag phases, indicating that these bacteria utilize probably some monosaccharides present in the used XOS. XOS were fermented with high specificity by Bifidobacteria strains, but Lactobacilli did not metabolize XOS efficiently. PMID:26019582

Construction of genetically engineered Candida tropicalis for conversion of l-arabinose to l-ribulose.

PubMed

Yeo, In-Seok; Shim, Woo-Yong; Kim, Jung Hoe

2018-05-20

For the biological production of l-ribulose, conversion by enzymes or resting cells has been investigated. However, expensive or concentrated substrates, an additional purification step to remove borate and the requirement for cell cultivation and harvest steps before utilization of resting cells make the production process complex and unfavorable. Microbial fermentation may help overcome these limitations. In this study, we constructed a genetically engineered Candida tropicalis strain to produce l-ribulose by fermentation with a glucose/l-arabinose mixture. For the uptake of l-arabinose as a substrate and conversion of l-arabinose to l-ribulose, two heterologous genes coding for l-arabinose transporter and l-arabinose isomerase, were constitutively expressed in C. tropicalis under the GAPDH promoter. The Arabidopsis thaliana-originated l-arabinose transporter gene (STP2)-expressing strain exhibited a high l-arabinose uptake rate of 0.103 g/g cell/h and the expression of l-arabinose isomerase from Lactobacillus sakei 23 K showed 30% of conversion (9 g/L) from 30 g/L of l-arabinose. This genetically engineered strain can be used for l-ribulose production by fermentation using mixed sugars of glucose and l-arabinose. Copyright © 2018 Elsevier B.V. All rights reserved.

Aerobic metabolism in the genus Lactobacillus: impact on stress response and potential applications in the food industry.

PubMed

Zotta, T; Parente, E; Ricciardi, A

2017-04-01

This review outlines the recent advances in the knowledge on aerobic and respiratory growth of lactic acid bacteria, focusing on the features of respiration-competent lactobacilli. The species of the genus Lactobacillus have been traditionally classified as oxygen-tolerant anaerobes, but it has been demonstrated that several strains are able to use oxygen as a substrate in reactions mediated by flavin oxidases and, in some cases, to synthesize a minimal respiratory chain. The occurrence of genes related to aerobic and respiratory metabolism and to oxidative stress response apparently correlates with the taxonomic position of lactobacilli. Members of the ecologically versatile Lactobacillus casei, L. plantarum and L. sakei groups are apparently best equipped to deal with aerobic/respiratory growth. The shift from anaerobic growth to aerobic (oxygen) and/or respiratory promoting (oxygen, exogenous haem and menaquinone) conditions offers physiological advantages and affects the pattern of metabolite production in several species. Even if this does not result in dramatic increases in biomass production and growth rate, cells grown in these conditions have improved tolerance to heat and oxidative stresses. An overview of benefits and of the potential applications of Lactobacillus cultures grown under aerobic or respiratory conditions is also discussed. © 2017 The Society for Applied Microbiology.

Species diversity, community dynamics, and metabolite kinetics of spontaneous leek fermentations.

PubMed

Wouters, D; Bernaert, N; Conjaerts, W; Van Droogenbroeck, B; De Loose, M; De Vuyst, L

2013-04-01

Leek (Allium ampeloprasum var. porrum) is one of Belgium's most important vegetables. All or part of the green leek parts are often left on the fields because of their limited cooking applications compared to the white leek parts. Therefore, the possibility to perform leek fermentations in view of product valorization and diversification was investigated. This study deals with the community dynamics, species diversity, and metabolite kinetics of spontaneous leek fermentations, thereby studying the influence of added NaCl concentration, harvesting season, and duration of the fermentation. The combination of a culture-dependent and culture-independent approach revealed the prevalence of lactic acid bacteria (LAB) from the third day of fermentation onwards, which was not influenced by the fermentation conditions applied. Enterobacteriaceae, Pseudomonadaceae, and yeasts disappeared after one week of fermentation. Leuconostoc mesenteroides, Lactobacillus sakei, and Lactobacillus plantarum, Lactobacillus brevis, and Lactobacillus parabrevis were the most frequently isolated LAB species. Both added NaCl concentrations were suitable to perform successful fermentations within three weeks. By that time, glucose and fructose, the main leek carbohydrates, were metabolized into mainly lactic acid, acetic acid, ethanol, and mannitol. A sensory analysis revealed that the fermented white leek parts were generally more appreciated than the fermented green leek parts. Copyright © 2012 Elsevier Ltd. All rights reserved.

Detection of Lactobacillus, Pediococcus, Leuconostoc, and Weissella Species in Human Feces by Using Group-Specific PCR Primers and Denaturing Gradient Gel Electrophoresis

PubMed Central

Walter, Jens; Hertel, Christian; Tannock, Gerald W.; Lis, Claudia M.; Munro, Karen; Hammes, Walter P.

2001-01-01

Denaturing gradient gel electrophoresis (DGGE) of DNA fragments generated by PCR with 16S ribosomal DNA-targeted group-specific primers was used to detect lactic acid bacteria (LAB) of the genera Lactobacillus, Pediococcus, Leuconostoc, and Weissella in human feces. Analysis of fecal samples of four subjects revealed individual profiles of DNA fragments originating not only from species that have been described as intestinal inhabitants but also from characteristically food-associated bacteria such as Lactobacillus sakei, Lactobacillus curvatus, Leuconostoc mesenteroides, and Pediococcus pentosaceus. Comparison of PCR-DGGE results with those of bacteriological culture showed that the food-associated species could not be cultured from the fecal samples by plating on Rogosa agar. On the other hand, all of the LAB species cultured from feces were detected in the DGGE profile. We also detected changes in the types of LAB present in human feces during consumption of a milk product containing the probiotic strain Lactobacillus rhamnosus DR20. The analysis of fecal samples from two subjects taken before, during, and after administration of the probiotic revealed that L. rhamnosus was detectable by PCR-DGGE during the test period in the feces of both subjects, whereas it was detectable by culture in only one of the subjects. PMID:11375166

Bovine mastitis may be associated with the deprivation of gut Lactobacillus.

PubMed

Ma, C; Zhao, J; Xi, X; Ding, J; Wang, H; Zhang, H; Kwok, L Y

2016-02-01

Bovine mastitis is an economical important microbial disease in dairy industry. Some recent human clinical trials have shown that oral probiotics supplementation could effectively control clinical mastitis, suggesting that the mechanism of mastitis protection might be achieved via the host gut microbiota. We aimed to test our hypothesis that bovine mastitis was related to changes in both the mammary and gut microbial profiles. By quantitative PCR, the milk and faecal microbial profiles of cows with low (<3×10 5 cells/ml) and high (>1×10 6 cells/ml) somatic cell count (SCC) were compared. Firstly, we observed drastic differences in both the milk and faecal microbial compositions at genus and Lactobacillus-species levels between the two groups. Secondly, the pattern of faecal microbial community changes of mastitis cows was similar to that of the milk, characterised by a general increase in the mastitis pathogens (Enterococcus, Streptococcus and Staphylococcus) and deprivation of Lactobacillus and its members (L. salivarius, L. sakei, L. ruminis, L. delbrueckii, L. buchneri, and L. acidophilus). Thirdly, only the faecal lactobacilli, but not bifidobacteria correlated with the milk microbial communities and SCC. Our data together hint to a close association between bovine mastitis, the host gut and milk microbiota.

Characterization of the bacterial spoilage flora in marinated pork products.

PubMed

Schirmer, B C; Heir, E; Langsrud, S

2009-06-01

To investigate the microbiota in marinated, vacuum-packed pork and to characterize isolated bacteria with regard to their spoilage potential. Laboratory marinated pork meat and commercial products from three Norwegian producers were examined. Lactic acid bacteria dominated in all products at the expiration date. The flora in marinated products was similar only for products from the same plant. Strains of Lactobacillus algidus, Lactobacillus sakei, Lactobacillus curvatus, Carnobacterium divergens, Carnobacterium maltaromaticum, Leuconostoc mesenteroides, Leuconostoc carnosum and Leuconostoc sp. were isolated and tested for their spoilage potential. Samples inoculated with Lact. algidus or Leuc. mesenteroides were rated as most unpleasant by randomly selected people. A sensory panel scored samples with Lact. algidus highest for sour and intense odour. Lactobacillus algidus was found in products from two out of three production plants. Culture-independent DNA isolation confirmed that cultivation on Blood agar at 20 degrees C yielded a representative picture of the total flora in marinated flintsteak. Lactobacillus algidus may be an important, but underestimated, spoilage organism that needs to be focused on more when spoilage of vacuum-packed meat is considered. Routine microbial testing may have to be revised in order to detect spoilage LAB that are unable to grow under currently used conditions.

Co-culturing of Lactobacillus paracasei subsp. paracasei with a Lactobacillus delbrueckii subsp. delbrueckii mutant to make high cell density for increased lactate productivity from cassava bagasse hydrolysate.

PubMed

John, Rojan Pappy; Nampoothiri, K Madhavan

2011-03-01

To increase the productivity of lactic acid, a co-culture of lactobacilli was made by mixing 1:1 ratio of Lactobacillus paracasei subsp. paracasei and a fast growing L. delbrueckii subsp. delbrueckii mutant. The culture was embedded on to polyurethane foam (PUF) cubes as a biofilm and used for fermentation. In order to prevent the cell leakage, the PUF cubes were further entrapped in calcium cross-linked alginate. The maximum lactic acid production using a high cell density free culture was >38 g l(-1) from ~40 g l(-1) of reducing sugar within 12 h of fermentation. Using PUF biofilms, the same yield of lactic acid attained after 24 h. When the cubes were further coated with alginate it took 36 h for the maximum yield. Even though, the productivity is slightly lesser with the alginate coating, cell leakage was decreased and cubes were reused without much decrease in production in repeated batches. Using a conventional control inoculum (3%, w/v), it took 120 h to yield same amount of lactic acid.

Conditioned food aversion for control of poisoning by Ipomoea carnea subsp. fistulosa

USDA-ARS?s Scientific Manuscript database

Conditioned food aversion is a technique that can be used to train livestock to avoid ingestion of poisonous plants. This study tested the efficacy and durability of conditioned food aversion to eliminate goat’s consumption of Ipomoea carnea subsp. fistulosa. We used 14 young Moxotó goats, which wer...

Phytochemical Composition and Antinociceptive Activity of Bauhinia glauca subsp. hupehana in Rats

PubMed Central

Xu, Jinlong; Zhao, Qizhi; Wei, Lei; Yang, Yu; Xu, Rui; Yu, Nengjiang; Zhao, Yimin

2015-01-01

In traditional medicine, Bauhinia glauca subsp. hupehana has long been used as an analgesic agent in China. The aim of this study was to evaluate the antinociceptive activity of the ethanol extract of the aerial parts of B. glauca subsp. hupehana (BHE) in rats and its chemical fingerprint. The antinociceptive activity of BHE was assessed in mice using chemically and heat–induced pain models, such as the acetic acid–induced writhing, hot plate, tail–flick and glutamate tests. Naltrexone hydrochloride, a non–selective opioid receptor antagonist, was utilized to determine the involvement of the opioid system. In addition to this, the involvements of the cGMP and ATP–sensitive K+ channel pathways were also detected using methylene blue and glibenclamide. The oral administration of BHE (at doses of 50, 100 and 200 mg/kg) produced significant and dose–related inhibitions in both the chemically and heat–induced pain models. Interestingly, in the abdominal constriction test, when the dose of BHE was increased to 800 mg/kg (p.o., n = 10), the inhibition rate was 100%. The antinociceptive mechanism may involve the cGMP pathway and ATP sensitive K+ channel pathway. The central antinociceptive effect was not antagonized by naltrexone. One phenolic acid, one lignin and five flavonoids were isolated from BHE. The antinociceptive activity of BHE was most likely due to the presence of the flavonoids. The acute toxicity results showed that BHE was safe at a high dose (2 g/kg, p.o.). The current investigation demonstrates that B. glauca subsp. hupehana is a potential candidate for the development of novel, non–opioid, analgesic phytomedicines. PMID:25658740

Phytochemical composition and antinociceptive activity of Bauhinia glauca subsp. hupehana in rats.

PubMed

Xu, Jinlong; Zhao, Qizhi; Wei, Lei; Yang, Yu; Xu, Rui; Yu, Nengjiang; Zhao, Yimin

2015-01-01

In traditional medicine, Bauhinia glauca subsp. hupehana has long been used as an analgesic agent in China. The aim of this study was to evaluate the antinociceptive activity of the ethanol extract of the aerial parts of B. glauca subsp. hupehana (BHE) in rats and its chemical fingerprint. The antinociceptive activity of BHE was assessed in mice using chemically and heat-induced pain models, such as the acetic acid-induced writhing, hot plate, tail-flick and glutamate tests. Naltrexone hydrochloride, a non-selective opioid receptor antagonist, was utilized to determine the involvement of the opioid system. In addition to this, the involvements of the cGMP and ATP-sensitive K+ channel pathways were also detected using methylene blue and glibenclamide. The oral administration of BHE (at doses of 50, 100 and 200 mg/kg) produced significant and dose-related inhibitions in both the chemically and heat-induced pain models. Interestingly, in the abdominal constriction test, when the dose of BHE was increased to 800 mg/kg (p.o., n = 10), the inhibition rate was 100%. The antinociceptive mechanism may involve the cGMP pathway and ATP sensitive K+ channel pathway. The central antinociceptive effect was not antagonized by naltrexone. One phenolic acid, one lignin and five flavonoids were isolated from BHE. The antinociceptive activity of BHE was most likely due to the presence of the flavonoids. The acute toxicity results showed that BHE was safe at a high dose (2 g/kg, p.o.). The current investigation demonstrates that B. glauca subsp. hupehana is a potential candidate for the development of novel, non-opioid, analgesic phytomedicines.

A highly efficient transposon mutagenesis system for the tomato pathogen Clavibacter michiganensis subsp. michiganensis.

PubMed

Kirchner, O; Gartemann, K H; Zellermann, E M; Eichenlaub, R; Burger, A

2001-11-01

A transposon mutagenesis system for Clavibacter michiganensis subsp. michiganensis was developed based on antibiotic resistance transposons that were derived from the insertion element IS1409 from Arthrobacter sp. strain TM1 NCIB12013. As a prerequisite, the electroporation efficiency was optimized by using unmethylated DNA and treatment of the cells with glycine such that about 5 x 10(6) transformants per microg of DNA were generally obtained. Electroporation of C. michiganensis subsp. michiganensis with a suicide vector carrying transposon Tn1409C resulted in approximately 1 x 10(3) transposon mutants per pg of DNA and thus is suitable for saturation mutagenesis. Analysis of Tn1409C insertion sites suggests a random mode of transposition. Transposition of Tn1409C was also demonstrated for other subspecies of C. michiganensis.

Factors affecting isolation and identification of Mycobacterium avium subsp. paratuberculosis from fecal and tissue samples in a liquid culture system.

PubMed

Whittington, Richard J

2009-03-01

Culture of Mycobacterium avium subsp. paratuberculosis is the definitive diagnostic test for Johne's disease, a chronic granulomatous enteropathy of animals. Compared to solid media, the identification of all strains of the organism in liquid media can be more difficult because the appearance of colonies and mycobactin dependence are not observable, and the growth of other organisms needs to be distinguished, commonly by PCR. Factors affecting the isolation rate of S strains and the contamination rate in modified Middlebrook 7H9 broth (Bactec 12B) and 7H10 agar were studied using 11,598 fecal samples and 2,577 tissue samples from sheep from 1,421 farms over 10 years. Minimization of contamination in Bactec cultures required the avoidance of the carryover of fecal particles from the first sedimentation step in the double-incubation centrifugation method, and contamination was reduced significantly by incubating the sample in a solution containing vancomycin, amphotericin B, and nalidixic acid for 3 days compared to 2 days. The growth of irrelevant microorganisms confounded the identification of M. avium subsp. paratuberculosis in liquid culture by inhibiting IS900 PCR and in solid medium culture by inhibiting the growth of M. avium subsp. paratuberculosis or obscuring colonies. The contamination of samples was clustered in certain laboratory submissions and was reduced by including ampicillin in Bactec medium without affecting the odds of isolation of M. avium subsp. paratuberculosis. The long-term contamination rate for fecal cultures was about 7%, and that for tissue cultures was <0.2%. Liquid medium was more sensitive than solid medium culture for M. avium subsp. paratuberculosis. The applicability of these findings for C strains is discussed.

Increased Production of Hydrogen Peroxide by Lactobacillus delbrueckii subsp. bulgaricus upon Aeration: Involvement of an NADH Oxidase in Oxidative Stress

PubMed Central

Marty-Teysset, C.; de la Torre, F.; Garel, J.-R.

2000-01-01

The growth of Lactobacillus delbrueckii subsp. bulgaricus (L. delbrueckii subsp. bulgaricus) on lactose was altered upon aerating the cultures by agitation. Aeration caused the bacteria to enter early into stationary phase, thus reducing markedly the biomass production but without modifying the maximum growth rate. The early entry into stationary phase of aerated cultures was probably related to the accumulation of hydrogen peroxide in the medium. Indeed, the concentration of hydrogen peroxide in aerated cultures was two to three times higher than in unaerated ones. Also, a similar shift from exponential to stationary phase could be induced in unaerated cultures by adding increasing concentrations of hydrogen peroxide. A significant fraction of the hydrogen peroxide produced by L. delbrueckii subsp. bulgaricus originated from the reduction of molecular oxygen by NADH catalyzed by an NADH:H2O2 oxidase. The specific activity of this NADH oxidase was the same in aerated and unaerated cultures, suggesting that the amount of this enzyme was not directly regulated by oxygen. Aeration did not change the homolactic character of lactose fermentation by L. delbrueckii subsp. bulgaricus and most of the NADH was reoxidized by lactate dehydrogenase with pyruvate. This indicated that NADH oxidase had no (or a very small) energetic role and could be involved in eliminating oxygen. PMID:10618234

Draft Genome Sequence of Lactobacillus delbrueckii subsp. bulgaricus LBB.B5.

PubMed

Urshev, Zoltan; Hajo, Karima; Lenoci, Leonardo; Bron, Peter A; Dijkstra, Annereinou; Alkema, Wynand; Wels, Michiel; Siezen, Roland J; Minkova, Svetlana; van Hijum, Sacha A F T

2016-10-06

Lactobacillus delbrueckii subsp. bulgaricus LBB.B5 originates from homemade Bulgarian yogurt and was selected for its ability to form a strong association with Streptococcus thermophilus The genome sequence will facilitate elucidating the genetic background behind the contribution of LBB.B5 to the taste and aroma of yogurt and its exceptional protocooperation with S. thermophilus. Copyright © 2016 Urshev et al.

Chemical composition and in vitro antioxidant activity of hydro-ethanolic extracts from Bauhinia forficata subsp. pruinosa and B. variegata.

PubMed

Sayago, Carla T M; Camargo, Vanessa B; Barbosa, F; Gularte, Cláudia; Pereira, Geovana; Miotto, Silvia; Cechinel Filho, V; Luiz Puntel, R; Folmer, V; Mendez, A

2013-03-01

Bauhinia species are known to have hypoglycemiant and antioxidant activities. Here, hydro-ethanolic leaf extracts from Bauhinia forficata subsp. pruinosa and Bauhinia variegata, collected in a Pampa biome region of Brazil, were investigated to characterize their chromatographic profile, flavonoid content and in vitro antioxidant activity (TBARS and DPH assays). The extracts were obtained from dried and fresh leaves. The total flavonoid content was assessed by spectrophotometric determination, and the results ranged between 572.08 and 1,102.99 μg mL-1. Moreover, flavonoids were more predominant in B. variegata than in B. forficata subsp. pruinosa. HPLC analysis detected a complex profile of phenolic compounds, being the flavonoid kaempferitrin founded B. forficata subsp. pruinosa; in addition, other kaempferol and quercetin derivatives were present. In vitro antioxidant assays demonstrated a different behavior depending on the species, leaf treatment and extract concentration. In general, B. variegata extracts obtained from fresh material presented higher antioxidant potential, which can be attributed to the predominance of flavonoids in their chemical composition.

Rocket Immunoelectrophoresis of the Entomocidal Parasporal Crystal of Bacillus thuringiensis subsp. kurstaki†

PubMed Central

Andrews, R. E.; Iandolo, J. J.; Campbell, B. S.; Davidson, L. I.; Bulla, L. A.

1980-01-01

Rocket immunoelectrophoresis was used to quantitate the soluble parasporal crystal of Bacillus thuringiensis subsp. kurstaki. The method described is rapid, reliable, specific, and extremely accurate, and it can be used to measure crystal toxin in commercial microbial insecticides that contain a mixture of spores, vegetative cells, and carrier materials. Images PMID:16345656

Characterization of two nisin-producing Lactococcus lactis subsp. lactis strains isolated from a commercial sauerkraut fermentation.

PubMed Central

Harris, L J; Fleming, H P; Klaenhammer, T R

1992-01-01

Two Lactococcus lactis subsp. lactis strains, NCK400 and LJH80, isolated from a commercial sauerkraut fermentation were shown to produce nisin. LJH80 was morphologically unstable and gave rise to two stable, nisin-producing (Nip+) derivatives, NCK318-2 and NCK318-3. NCK400 and derivatives of LJH80 exhibited identical morphological and metabolic characteristics, but could be distinguished on the basis of plasmid profiles and genomic hybridization patterns to a DNA probe specific for the iso-ISS1 element, IS946. NCK318-2 and NCK318-3 harbored two and three plasmids, respectively, which hybridized with IS946. Plasmid DNA was not detected in NCK400, and DNA from this strain failed to hybridize with IS946. Despite the absence of detectable plasmid DNA in NCK400, nisin-negative derivatives (NCK402 and NCK403) were isolated after repeated transfer in broth at 37 degrees C. Nisin-negative derivatives concurrently lost the ability to ferment sucrose and became sensitive to nisin. A 4-kbp HindIII fragment containing the structural gene for nisin (spaN), cloned from L. lactis subsp. lactis ATCC 11454, was used to probe genomic DNA of NCK318-2, NCK318-3, NCK400, and NCK402 digested with EcoRI or HindIII. The spaN probe hybridized to an 8.8-kbp EcoRI fragment and a 10-kbp HindIII fragment in the Nip+ sauerkraut isolates, but did not hybridize to the Nip- derivative, NCK402. A different hybridization pattern was observed when the same probe was used against Nip+ L. lactis subsp. lactis ATCC 11454 and ATCC 7962. These phenotypic and genetic data confirmed that unique Nip+ L. lactis subsp. lactis strains were isolated from fermenting sauerkraut. Images PMID:1622214

Comparison of culture and a novel 5' Taq nuclease assay for direct detection of Campylobacter fetus subsp. venerealis in clinical specimens from cattle.

PubMed

McMillen, Lyle; Fordyce, Geoffry; Doogan, Vivienne J; Lew, Ala E

2006-03-01

A Campylobacter fetus subsp. venerealis-specific 5' Taq nuclease PCR assay using a 3' minor groove binder-DNA probe (TaqMan MGB) was developed based on a subspecies-specific fragment of unknown identity (S. Hum, K. Quinn, J. Brunner, and S. L. On, Aust. Vet. J. 75:827-831, 1997). The assay specifically detected four C. fetus subsp. venerealis strains with no observed cross-reaction with C. fetus subsp. fetus-related Campylobacter species or other bovine venereal microflora. The 5' Taq nuclease assay detected approximately one single cell compared to 100 and 10 cells in the conventional PCR assay and 2,500 and 25,000 cells from selective culture from inoculated smegma and mucus, respectively. The respective detection limits following the enrichments from smegma and mucus were 5,000 and 50 cells/inoculum for the conventional PCR compared to 500 and 50 cells/inoculum for the 5' Taq nuclease assay. Field sampling confirmed the sensitivity and the specificity of the 5' Taq nuclease assay by detecting an additional 40 bulls that were not detected by culture. Urine-inoculated samples demonstrated comparable detection of C. fetus subsp. venerealis by both culture and the 5' Taq nuclease assay; however, urine was found to be less effective than smegma for bull sampling. Three infected bulls were tested repetitively to compare sampling tools, and the bull rasper proved to be the most suitable, as evidenced by the improved ease of specimen collection and the consistent detection of higher levels of C. fetus subsp. venerealis. The 5' Taq nuclease assay demonstrates a statistically significant association with culture (chi2 = 29.8; P < 0.001) and significant improvements for the detection of C. fetus subsp. venerealis-infected animals from crude clinical extracts following prolonged transport.

Phylogenetic Analysis and Polyphasic Characterization of Clavibacter michiganensis Strains Isolated from Tomato Seeds Reveal that Nonpathogenic Strains Are Distinct from C. michiganensis subsp. michiganensis

PubMed Central

Durand, Karine; Orgeur, Geoffrey; Balidas, Samuel; Fricot, Céline; Bonneau, Sophie; Quillévéré, Anne; Audusseau, Corinne; Olivier, Valérie; Grimault, Valérie; Mathis, René

2012-01-01

The genus Clavibacter comprises one species and five subspecies of plant-pathogenic bacteria, four of which are classified as quarantine organisms due to the high economic threat they pose. Clavibacter michiganensis subsp. michiganensis is one of the most important pathogens of tomato, but the recommended diagnostic tools are not satisfactory due to false-negative and/or -positive results. To provide a robust analysis of the genetic relatedness among a worldwide collection of C. michiganensis subsp. michiganensis strains, relatives (strains from the four other C. michiganensis subspecies), and nonpathogenic Clavibacter-like strains isolated from tomato, we performed multilocus sequence-based analysis and typing (MLSA and MLST) based on six housekeeping genes (atpD, dnaK, gyrB, ppK, recA, and rpoB). We compared this “framework” with phenotypic and genotypic characteristics such as pathogenicity on tomato, reaction to two antisera by immunofluorescence and to five PCR identification tests, and the presence of four genes encoding the main C. michiganensis subsp. michiganensis pathogenicity determinants. We showed that C. michiganensis subsp. michiganensis is monophyletic and is distinct from its closest taxonomic neighbors. The nonpathogenic Clavibacter-like strains were identified as C. michiganensis using 16S rRNA gene sequencing. These strains, while cross-reacting with C. michiganensis subsp. michiganensis identification tools, are phylogenetically distinct from the pathogenic strains but belong to the C. michiganensis clade. C. michiganensis subsp. michiganensis clonal complexes linked strains from highly diverse geographical origins and also strains isolated over long periods of time in the same location. This illustrates the importance of seed transmission in the worldwide dispersion of this pathogen and its survival and adaptation abilities in a new environment once introduced. PMID:23001675

Phylogenetic analysis and polyphasic characterization of Clavibacter michiganensis strains isolated from tomato seeds reveal that nonpathogenic strains are distinct from C. michiganensis subsp. michiganensis.

PubMed

Jacques, Marie-Agnès; Durand, Karine; Orgeur, Geoffrey; Balidas, Samuel; Fricot, Céline; Bonneau, Sophie; Quillévéré, Anne; Audusseau, Corinne; Olivier, Valérie; Grimault, Valérie; Mathis, René

2012-12-01

The genus Clavibacter comprises one species and five subspecies of plant-pathogenic bacteria, four of which are classified as quarantine organisms due to the high economic threat they pose. Clavibacter michiganensis subsp. michiganensis is one of the most important pathogens of tomato, but the recommended diagnostic tools are not satisfactory due to false-negative and/or -positive results. To provide a robust analysis of the genetic relatedness among a worldwide collection of C. michiganensis subsp. michiganensis strains, relatives (strains from the four other C. michiganensis subspecies), and nonpathogenic Clavibacter-like strains isolated from tomato, we performed multilocus sequence-based analysis and typing (MLSA and MLST) based on six housekeeping genes (atpD, dnaK, gyrB, ppK, recA, and rpoB). We compared this "framework" with phenotypic and genotypic characteristics such as pathogenicity on tomato, reaction to two antisera by immunofluorescence and to five PCR identification tests, and the presence of four genes encoding the main C. michiganensis subsp. michiganensis pathogenicity determinants. We showed that C. michiganensis subsp. michiganensis is monophyletic and is distinct from its closest taxonomic neighbors. The nonpathogenic Clavibacter-like strains were identified as C. michiganensis using 16S rRNA gene sequencing. These strains, while cross-reacting with C. michiganensis subsp. michiganensis identification tools, are phylogenetically distinct from the pathogenic strains but belong to the C. michiganensis clade. C. michiganensis subsp. michiganensis clonal complexes linked strains from highly diverse geographical origins and also strains isolated over long periods of time in the same location. This illustrates the importance of seed transmission in the worldwide dispersion of this pathogen and its survival and adaptation abilities in a new environment once introduced.

Comparison of the acidifying activity of Lactococcus lactis subsp. lactis strains isolated from goat's milk and Valdeteja cheese.

PubMed

Alonso-Calleja, C; Carballo, J; Capita, R; Bernardo, A; García-López, M L

2002-01-01

This work was carried out to study the acid production by Lactococcus lactis subsp. lactis strains isolated from goat's milk and goat cheese (Valdeteja variety) in order to select a suitable starter culture for industrial goat cheese manufacturing. The titrable acidity of 45 Lactococcus lactis subsp. lactis strains isolated from a home-made batch of Valdeteja cheese with excellent sensory characteristics was measured over a period of 18 h. The strains were divided into two groups depending on the acid production rate: 20 fast acid producer (F) strains and 25 slow acid producer (S) strains. The kinetic parameters (lag phase, maximum acid production rate and value of upper asymptote curve) of the acid production curves for F and S strains were significantly (P < 0.001) different. Significant (P < 0.001) differences between titrable acidity of F and S strains were observed after the second hour of incubation. An F strain acetoin producer (Lactococcus lactis subsp. lactis 470Ch2) was selected as autochthonous starter culture for industrial Valdeteja goat cheese manufacturing.

Virulence of Photobacterium damselae subsp. piscicida in cultured cobia Rachycentron canadum.

PubMed

Liu, Ping-Chung; Lin, Ji-Yang; Lee, Kuo-Kau

2003-01-01

An outbreak of serious mortality among the cultured cobia Rachycentron canadum (weighing 3 kg) characterized by the presence of whitish granulomatous deposits on the kidney, liver and spleen occurred in July of 2000 in Taiwan. A non-motile strain CP1 was isolated from kidney and/or liver on tryptic soy agar and/or brain heart infusion agar plates (both supplemented with 1% NaCl, w/v). This strain was characterized and identified as Photobacterium damselae subsp. piscicida using biochemical characteristics and Bionor mono-Pp tests. The bacterium and its extracellular products (ECP) were lethal to the cobia (weighing 10 g) with LD50 values of 1.03 x 10(4) colony forming units and 1.26 microg protein/g fish body weight, respectively. All the moribund/dead fish exhibited darkness in color with no gross or internal leasions. However, the bacteria could be reisolated from kidney and liver after bacterial challenge. The present results reveal that Ph. damselae subsp. piscicida is the causative agent of fish photobacteriosis in the cobia and the bacterium isolated from sub-adult cobia (chronic form) is virulent to young cobia causing acute form of the disease.

Antimicrobial susceptibility and multilocus sequence typing of Mycoplasma capricolum subsp. capricolum

PubMed Central

Tatay-Dualde, Juan; Prats-van der Ham, Miranda; Paterna, Ana; Sánchez, Antonio; Corrales, Juan Carlos; Contreras, Antonio; Tola, Sebastiana; Gómez-Martin, Ángel

2017-01-01

Mycoplasma capricolum subsp. capricolum is one of the causative agents of contagious agalactia (CA). Nevertheless, there is still a lack of information about its antimicrobial susceptibility and genetic characteristics. Therefore, the aim of this work was to study the antimicrobial and genetic variability of different Mycoplasma capricolum subsp. capricolum field isolates. For this purpose, the growth inhibition effect of 18 antimicrobials and a multilocus sequence typing (MLST) scheme based on five housekeeping genes (fusA, glpQ, gyrB, lepA and rpoB) were performed on 32 selected field isolates from Italy and Spain.The results showed a wide range of growth inhibitory effects for almost all the antimicrobials studied. Macrolides presented lower efficacy inhibiting Mcc growth than in previous works performed on other CA-causative mycoplasmas. Erythromycin was not able to inhibit the growth of any of the studied strains, contrary to doxycycline, which inhibited the growth of all of them from low concentrations. On the other hand, the study of the concatenated genes revealed a high genetic variability among the different Mcc isolates. Hence, these genetic variations were greater than the ones reported in prior works on other mycoplasma species. PMID:28346546

Characterization of Cyt2Bc Toxin from Bacillus thuringiensis subsp. medellin

PubMed Central

Juárez-Pérez, Victor; Guerchicoff, Alejandra; Rubinstein, Clara; Delécluse, Armelle

2002-01-01

We cloned and sequenced a new cytolysin gene from Bacillus thuringiensis subsp. medellin. Three IS240-like insertion sequence elements and the previously cloned cyt1Ab and p21 genes were found in the vicinity of the cytolysin gene. The cytolysin gene encodes a protein 29.7 kDa in size that is 91.5% identical to Cyt2Ba from Bacillus thuringiensis subsp. israelensis and has been designated Cyt2Bc. Inclusions containing Cyt2Bc were purified from the crystal-negative strain SPL407 of B. thuringiensis. Cyt2Bc reacted weakly with antibodies directed against Cyt2Ba and was not recognized by an antiserum directed against the reference cytolysin Cyt1Aa. Cyt2Bc was hemolytic only upon activation with trypsin and had only one-third to one-fifth of the activity of Cyt2Ba, depending on the activation time. Cyt2Bc was also mosquitocidal against Aedes aegypti, Anopheles stephensi, and Culex quinquefasciatus, including strains resistant to the Bacillus sphaericus binary toxin. Its toxicity was half of that of Cyt2Ba on all mosquito species except resistant C. quinquefasciatus. PMID:11872472

Genome Sequence of the Cheese-Starter Strain Lactobacillus delbrueckii subsp. lactis CRL 581.

PubMed

Hebert, Elvira María; Raya, Raúl R; Brown, Lucía; Font de Valdez, Graciela; Savoy de Giori, Graciela; Taranto, María Pía

2013-08-08

We report the genome sequence of Lactobacillus delbrueckii subsp. lactis CRL 581 (1,911,137 bp, GC 49.7%), a proteolytic strain isolated from a homemade Argentinian hard cheese which has a key role in bacterial nutrition and releases bioactive health-beneficial peptides from milk proteins.

Development and Evaluation of a Novel Multicopy-Element-Targeting Triplex PCR for Detection of Mycobacterium avium subsp. paratuberculosis in Feces

PubMed Central

Garrido, Joseba M.; Molina, Elena; Geijo, María V.; Elguezabal, Natalia; Vázquez, Patricia; Juste, Ramón A.

2014-01-01

The enteropathy called paratuberculosis (PTB), which mainly affects ruminants and has a worldwide distribution, is caused by Mycobacterium avium subsp. paratuberculosis. This disease significantly reduces the cost-effectiveness of ruminant farms, and therefore, reliable and rapid detection methods are needed to control the spread of the bacterium in livestock and in the environment. The aim of this study was to identify a specific and sensitive combination of DNA extraction and amplification to detect M. avium subsp. paratuberculosis in feces. Negative bovine fecal samples were inoculated with increasing concentrations of two different bacterial strains (field and reference) to compare the performance of four extraction and five amplification protocols. The best results were obtained using the JohnePrep and MagMax extraction kits combined with an in-house triplex real-time PCR designed to detect IS900, ISMap02 (an insertion sequence of M. avium subsp. paratuberculosis present in 6 copies per genome), and an internal amplification control DNA simultaneously. These combinations detected 10 M. avium subsp. paratuberculosis cells/g of spiked feces. The triplex PCR detected 1 fg of genomic DNA extracted from the reference strain K10. The performance of the robotized version of the MagMax extraction kit combined with the IS900 and ISMap02 PCR was further evaluated using 615 archival fecal samples from the first sampling of nine Friesian cattle herds included in a PTB control program and followed up for at least 4 years. The analysis of the results obtained in this survey demonstrated that the diagnostic method was highly specific and sensitive for the detection of M. avium subsp. paratuberculosis in fecal samples from cattle and a very valuable tool to be used in PTB control programs. PMID:24727272

Terrestrial and marine Antarctic fungi extracts active against Xanthomonas citri subsp. citri.

PubMed

Vieira, G; Purić, J; Morão, L G; Dos Santos, J A; Inforsato, F J; Sette, L D; Ferreira, H; Sass, D C

2018-07-01

This study aims to obtain secondary metabolites extracts from filamentous fungi isolated from soil and marine sediments from Antarctica and assess its potential antibacterial activity on Xanthomonas citri subsp. citri, the agent of citrus canker. Metabolites production was conducted in Malt 2% broth at 15°C for 20 days after which intracellular and extracellular extracts were obtained. The extracts were evaluated by cell viability assays through Resazurin Microtitre Assay. From 158 fungal extracts, 33 hampered bacterial growth in vitro. The average inhibition of the extracts obtained from terrestrial (soil) and marine (sediments) fungi was 94 and 97% respectively. These inhibition values were close to the average of 90% cell death for the positive control. MIC90 and MBC for the bioactive extracts were established. Isolates that produced active metabolites against the phytopathogen were identified using molecular taxonomy (ITS-rRNA sequencing) as: Pseudogymnoascus, Penicillium, Cadophora, Paraconiothyrium and Toxicocladosporium. Antarctic fungal strains isolated from terrestrial and marine sediments were able to produce secondary metabolites with antimicrobial activity against X. citri subsp. citri, highlighting the importance of these microbial genetic resources. These metabolites have potential to be used as alternatives for the control of this plant pathogen. This manuscript makes an impact on the study of micro-organisms from extreme habitats and their possible contribution in discovering new active molecules against pathogens of agricultural interest. Studies on the Antarctic continent and its communities have attracted the scientific community due to the long period of isolation and low levels of disturbance that surrounds the region. Knowing the potential of fungi in this region to produce active secondary metabolites, we aim to contribute to the discovery of compounds with antibacterial action in Xanthomonas citri subsp. citri, a plant pathogen present in

Bartonella vinsonii subsp. berkhoffii and Bartonella henselae as potential causes of proliferative vascular diseases in animals.

PubMed

Beerlage, Christiane; Varanat, Mrudula; Linder, Keith; Maggi, Ricardo G; Cooley, Jim; Kempf, Volkhard A J; Breitschwerdt, Edward B

2012-08-01

Bartonella species are highly fastidious, vector borne, zoonotic bacteria that cause persistent intraerythrocytic bacteremia and endotheliotropic infection in reservoir and incidental hosts. Based upon prior in vitro research, three Bartonella sp., B. bacilliformis, B. henselae, and B. quintana can induce proliferation of endothelial cells, and each species has been associated with in vivo formation of vasoproliferative tumors in human patients. In this study, we report the molecular detection of B. vinsonii subsp. berkhoffii, B. henselae, B. koehlerae, or DNA of two of these Bartonella species simultaneously in vasoproliferative hemangiopericytomas from a dog, a horse, and a red wolf and in systemic reactive angioendotheliomatosis lesions from cats and a steer. In addition, we provide documentation that B. vinsonii subsp. berkhoffii infections induce activation of hypoxia inducible factor-1 and production of vascular endothelial growth factor, thereby providing mechanistic evidence as to how these bacteria could contribute to the development of vasoproliferative lesions. Based upon these results, we suggest that a fourth species, B. vinsonii subsp. berkhoffii, should be added to the list of bartonellae that can induce vasoproliferative lesions and that infection with one or more Bartonella sp. may contribute to the pathogenesis of systemic reactive angioendotheliomatosis and hemangiopericytomas in animals.

Complete genome sequence of salmonella enterica subsp. enterica Serovar Thompson Strain RM6836

USDA-ARS?s Scientific Manuscript database

Salmonella enterica subsp. enterica serovar Thompson (S. Thompson) strain RM6836 was isolated from lettuce in 2002. We report the complete sequence and annotation of the genome of S. Thompson strain RM6836. This is the first reported complete genome sequence for S. Thompson and will provide a point ...

Complete genomic sequence of campylobacter jejuni subsp. jejuni HS:19 penner reference strain

USDA-ARS?s Scientific Manuscript database

Campylobacter jejuni subsp. jejuni (Cjj) infections are a leading cause of foodborne gastroenteritis and the most prevalent antecedent to Guillain-Barré syndrome (GBS). Capsular type Penner HS:19 is among several capsule types shown to be markers for GBS. This study describes the genome of Cjj HS:19...

Bartonella vinsonii subsp. berkhoffii and B. henselae in dogs.

PubMed

Müller, A; Soto, F; Sepúlveda, M; Bittencourt, P; Benevenute, J L; Ikeda, P; Machado, R Z; André, M R

2018-05-06

This study aimed to molecularly survey Bartonella in dogs from Chile. Quantitative real-time PCR (qPCR) for Bartonella spp. based on nuoG gene was performed in 139 blood samples taken from dogs belonging to rural localities of the Valdivia Province, Los Ríos region, southern Chile. nuoG qPCR-positive samples were submitted to conventional PCR assays for ftsZ, gltA, rpoB and nuoG genes and sequencing for speciation and phylogenetic analysis. Based upon qPCR results, Bartonella spp. occurrence in dogs was 4.3% (6/139). Out of six nuoG qPCR-positive samples, six, three, two and none showed positive results in cPCR assays based on gltA, ftsZ, rpoB and nuoG genes, respectively. Consistent sequencing results were obtained only for the ftsZ gene from sample #1532 (GeneBank accession number: MG252491), and gltA gene from samples #1535 (MG252490) and #1532 (148 bp fragment that was not deposited in GenBank). Phylogenetic analysis of ftsZ and gltA genes allowed speciation of two nuoG-positive samples, one as Bartonella vinsonii subsp. berkhoffii and the other as B. henselae. Bartonella vinsonii subsp. berkhoffii and B. henselae are detected for the first time in dogs from Chile, highlighting the importance of the canine population as a source of zoonotic agents and potential infection risk to humans.

First isolation of Mycoplasma capricolum subsp. capricolum, one of the causal agents of caprine contagious agalactia, on the island of Lanzarote (Spain).

PubMed

De la Fe, C; Gutiérrez, A; Poveda, J B; Assunção, P; Ramírez, A S; Fabelo, F

2007-03-01

During an unusually long period of bad weather, several outbreaks of caprine contagious agalactia (CCA) were reported in a number of flocks on the island of Lanzarote (Canary Islands, Spain). Clinical and subclinical mastitis in lactating goats and some cases of arthritis and pneumonia in kids were observed in the affected flocks. Mycoplasma capricolum subsp. capricolum was isolated as the main causal agent of the outbreaks, associated with M. mycoides subsp. mycoides "large colony type" (Mmm LC) in two flocks. This is the first report of an isolation of M. capricolum subsp. capricolum on the island of Lanzarote. The finding is of epidemiological importance and could complicate plans to control the disease. The significance of this mycoplasma species in association with CCA must now be studied in detail.

DrsG from Streptococcus dysgalactiae subsp. equisimilis Inhibits the Antimicrobial Peptide LL-37

PubMed Central

Smyth, Danielle; Cameron, Ainslie; Davies, Mark R.; McNeilly, Celia; Hafner, Louise; Sriprakash, Kadaba S.

2014-01-01

SIC and DRS are related proteins present in only 4 of the >200 Streptococcus pyogenes emm types. These proteins inhibit complement-mediated lysis and/or the activity of certain antimicrobial peptides (AMPs). A gene encoding a homologue of these proteins, herein called DrsG, has been identified in the related bacterium Streptococcus dysgalactiae subsp. equisimilis. Here we show that geographically dispersed isolates representing 14 of 50 emm types examined possess variants of drsG. However, not all isolates within the drsG-positive emm types possess the gene. Sequence comparisons also revealed a high degree of conservation in different S. dysgalactiae subsp. equisimilis emm types. To examine the biological activity of DrsG, recombinant versions of two major DrsG variants, DrsGS and DrsGL, were expressed and purified. Western blot analysis using antisera raised to these proteins demonstrated both variants to be expressed and secreted into culture supernatants. Unlike SIC, but similar to DRS, DrsG does not inhibit complement-mediated lysis. However, like both SIC and DRS, DrsG is a ligand of the cathelicidin LL-37 and is inhibitory to its bactericidal activity in in vitro assays. Conservation of prolines in the C-terminal region also suggests that these residues are important in the biology of this family of proteins. This is the first report demonstrating the activity of an AMP-inhibitory protein in S. dysgalactiae subsp. equisimilis and suggests that inhibition of AMP activity is the primary function of this family of proteins. The acquisition of the complement-inhibitory activity of SIC may reflect its continuing evolution. PMID:24664506

Inferring biomarkers for Mycobacterium avium subsp. paratuberculosis infection and disease progression using experimental data

USDA-ARS?s Scientific Manuscript database

Available diagnostic assays for Mycobacterium avium subsp paratuberculosis (MAP) have poor sensitivities and cannot detect early stages of the infection, therefore, there is need to find new diagnostic markers for early infection detection and disease stages. We analyzed longitudinal IFN- gamma, ELI...

Comparison of fecal DNA extraction kits for the detection of Mycobacterium avium subsp. paratuberculosis

USDA-ARS?s Scientific Manuscript database

Fecal culture is considered the gold standard for the diagnostics of paratuberculosis, however, PCR for the detection of Mycobacterium avium subsp. paratuberculosis (MAP) in fecal material is widely used today, having demonstrated great sensitivity and specificity. To insure the most efficient and r...

Comparison of fecal DNA extraction kits for the detection of Mycobacterium avium subsp. paratuberculosis

USDA-ARS?s Scientific Manuscript database

Fecal culture is considered the gold standard for the diagnosis of paratuberculosis, however, PCR for the detection of Mycobacterium avium subsp. paratuberculosis (MAP) in fecal material is widely used today, having demonstrated great sensitivity and specificity. To insure the most efficient and rep...

Isolation of Bartonella henselae, Bartonella koehlerae subsp. koehlerae, Bartonella koehlerae subsp. bothieri and a new subspecies of B. koehlerae from free-ranging lions (Panthera leo) from South Africa, cheetahs (Acinonyx jubatus) from Namibia and captive cheetahs from California.

PubMed

Molia, S; Kasten, R W; Stuckey, M J; Boulouis, H J; Allen, J; Borgo, G M; Koehler, J E; Chang, C C; Chomel, B B

2016-11-01

Bartonellae are blood- and vector-borne Gram-negative bacteria, recognized as emerging pathogens. Whole-blood samples were collected from 58 free-ranging lions (Panthera leo) in South Africa and 17 cheetahs (Acinonyx jubatus) from Namibia. Blood samples were also collected from 11 cheetahs (more than once for some of them) at the San Diego Wildlife Safari Park. Bacteria were isolated from the blood of three (5%) lions, one (6%) Namibian cheetah and eight (73%) cheetahs from California. The lion Bartonella isolates were identified as B. henselae (two isolates) and B. koehlerae subsp. koehlerae. The Namibian cheetah strain was close but distinct from isolates from North American wild felids and clustered between B. henselae and B. koehlerae. It should be considered as a new subspecies of B. koehlerae. All the Californian semi-captive cheetah isolates were different from B. henselae or B. koehlerae subsp. koehlerae and from the Namibian cheetah isolate. They were also distinct from the strains isolated from Californian mountain lions (Felis concolor) and clustered with strains of B. koehlerae subsp. bothieri isolated from free-ranging bobcats (Lynx rufus) in California. Therefore, it is likely that these captive cheetahs became infected by an indigenous strain for which bobcats are the natural reservoir.

Phylogeography and seed dispersal in islands: the case of Rumex bucephalophorus subsp. canariensis (Polygonaceae).

PubMed

Talavera, María; Navarro-Sampedro, Laura; Ortiz, Pedro L; Arista, Montserrat

2013-02-01

Rumex bucephalophorus subsp. canariensis is an endemic taxon to Macaronesia with diaspore polymorphism. The origin and colonizing route of this taxon in Macaronesia was studied using molecular data and information on diaspore types. Amplified fragment length polymorphism (AFLP) was used in 260 plants from 22 populations of R. bucephalophorus subsp. canariensis, four from the Madeiran archipelago and 18 from the Canary archipelago. Diaspore production was analysed in 9-50 plants from each population used for AFLP analysis. One hundred and one plants from the Madeiran archipelago and 375 plants from the Canary Islands were studied. For each plant the type of diaspore produced was recorded. Overall populations had low genetic diversity but they showed a geographical pattern of genetic diversity that was higher in the older eastern islands than in the younger western ones. Two types of dispersible diaspores were found: in the eastern Canary islands (Lanzarote, Fuerteventura and Gran Canaria), plants produced exclusively long-dispersible diaspores, whereas in the western Canary islands (Tenerife, La Gomera, El Hierro) and the Madeiran archipelago plants produced exclusively short-dispersible diaspores. Genetically, the studied populations fell into four main island groups: Lanzarote-Fuerteventura, Gran Canaria, Tenerife-El Hierro and La Gomera-Madeira archipelago. A Moroccan origin of R. bucephalophorus subsp. canariensis is hypothesized with a colonization route from the eastern to the western islands. In addition, at least one gene flow event from La Gomera to the Madeiran archipelago has taken place. During the colonization process the type of dispersible diaspore changed so that dispersability decreased in populations of the westernmost islands.

Detection of Mycobacterium avium subsp. paratuberculosis in Drinking Water and Biofilms Using Quantitative PCR

EPA Science Inventory

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s disease in domestic animals and has been implicated in Crohn’s disease in humans. Cows infected with Johne’s disease shed large quantities of MAP into soil. Further, MAP has been isolated from surface water, is resi...

Two-component regulators involved in the global control of virulence in Erwinia carotovora subsp. carotovora.

PubMed

Eriksson, A R; Andersson, R A; Pirhonen, M; Palva, E T

1998-08-01

Production of extracellular, plant cell wall degrading enzymes, the main virulence determinants of the plant pathogen Erwinia carotovora subsp. carotovora, is coordinately controlled by a complex regulatory network. Insertion mutants in the exp (extracellular enzyme production) loci exhibit pleiotropic defects in virulence and the growth-phase-dependent transcriptional activation of genes encoding extracellular enzymes. Two new exp mutations, designated expA and expS, were characterized. Introduction of the corresponding wild-type alleles to the mutants complemented both the lack of virulence and the impaired production of plant cell wall degrading enzymes. The expA gene was shown to encode a 24-kDa polypeptide that is structurally and functionally related to the uvrY gene product of Escherichia coli and the GacA response regulator of Pseudomonas fluorescens. Functional similarity of expA and uvrY was demonstrated by genetic complementation. The expA gene is organized in an operon together with a uvrC-like gene, identical to the organization of uvrY and uvrC in E. coli. The unlinked expS gene encodes a putative sensor kinase that shows 92% identity to the recently described rpfA gene product from another E. carotovora subsp. carotovora strain. Our data suggest that ExpS and ExpA are members of two-component sensor kinase and response regulator families, respectively. These two proteins might interact in controlling virulence gene expression in E. carotovora subsp. carotovora.

Genome of the Actinomycete Plant Pathogen Clavibacter michiganensis subsp. sepedonicus Suggests Recent Niche Adaptation▿ †

PubMed Central

Bentley, Stephen D.; Corton, Craig; Brown, Susan E.; Barron, Andrew; Clark, Louise; Doggett, Jon; Harris, Barbara; Ormond, Doug; Quail, Michael A.; May, Georgiana; Francis, David; Knudson, Dennis; Parkhill, Julian; Ishimaru, Carol A.

2008-01-01

Clavibacter michiganensis subsp. sepedonicus is a plant-pathogenic bacterium and the causative agent of bacterial ring rot, a devastating agricultural disease under strict quarantine control and zero tolerance in the seed potato industry. This organism appears to be largely restricted to an endophytic lifestyle, proliferating within plant tissues and unable to persist in the absence of plant material. Analysis of the genome sequence of C. michiganensis subsp. sepedonicus and comparison with the genome sequences of related plant pathogens revealed a dramatic recent evolutionary history. The genome contains 106 insertion sequence elements, which appear to have been active in extensive rearrangement of the chromosome compared to that of Clavibacter michiganensis subsp. michiganensis. There are 110 pseudogenes with overrepresentation in functions associated with carbohydrate metabolism, transcriptional regulation, and pathogenicity. Genome comparisons also indicated that there is substantial gene content diversity within the species, probably due to differential gene acquisition and loss. These genomic features and evolutionary dating suggest that there was recent adaptation for life in a restricted niche where nutrient diversity and perhaps competition are low, correlated with a reduced ability to exploit previously occupied complex niches outside the plant. Toleration of factors such as multiplication and integration of insertion sequence elements, genome rearrangements, and functional disruption of many genes and operons seems to indicate that there has been general relaxation of selective pressure on a large proportion of the genome. PMID:18192393

Co-isolation of Bartonella henselae and Bartonella vinsonii subsp. berkhoffii from blood, joint and subcutaneous seroma fluids from two naturally infected dogs.

PubMed

Diniz, Pedro Paulo Vissotto de Paiva; Wood, Michael; Maggi, Ricardo G; Sontakke, Sushama; Stepnik, Matt; Breitschwerdt, Edward B

2009-09-18

This report describes the clinical presentation, isolation and treatment of two dogs naturally infected with Bartonella henselae and Bartonella vinsonii subsp. berkhoffii. Chronic and progressive polyarthritis was the primary complaint for dog #1, from which B. henselae and B. vinsonii subsp. berkhoffii were cultured on three independent occasions from blood and joint fluid samples, despite administration of nearly 4 months of non-consecutive antibiotic therapy. A clinically atypical and progressively severe trauma-associated seroma was the primary complaint for dog #2, from which B. henselae and B. vinsonii subsp. berkhoffii were isolated from serum, blood and seroma fluid. Dogs can be co-infected with two Bartonella spp. and infection with these organisms should not be ruled out if specific antibodies are not detected. Specialized culture techniques should be used for isolation and to assess antibiotic efficacy.

Lymphoproliferative and gamma interferon responses to stress-regulated Mycobacterium avium subsp. paratuberculosis recombinant proteins

USDA-ARS?s Scientific Manuscript database

Johne’s disease in ruminants is a chronic infection of the intestines caused by Mycobacterium avium subsp. paratuberculosis. Economic losses associated with Johne’s disease arise due to premature culling, reduced production of milk and wool and mortalities. The disease is characterised by a long inc...

Bacteriocinogenic Lactococcus lactis subsp. lactis DF04Mi isolated from goat milk: Characterization of the bacteriocin

PubMed Central

Furtado, Danielle N.; Todorov, Svetoslav D.; Landgraf, Mariza; Destro, Maria T.; Franco, Bernadette D.G.M.

2014-01-01

Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its antimicrobial activity. The bacteriocin presented a broad spectrum of activity, was sensitive to proteolytic enzymes, resistant to heat and pH extremes, and not affected by the presence of SDS, Tween 20, Tween 80, EDTA or NaCl. Bacteriocin production was dependent on the components of the culture media, especially nitrogen source and salts. When tested by PCR, the bacteriocin gene presented 100% homology to nisin Z gene. These properties indicate that this L. lactis subsp. lactis DF4Mi can be used for enhancement of dairy foods safety and quality. PMID:25763065

Complete Genome Sequence of the Quality Control Strain Staphylococcus aureus subsp. aureus ATCC 25923

PubMed Central

Treangen, Todd J.; Maybank, Rosslyn A.; Enke, Sana; Friss, Mary Beth; Diviak, Lynn F.; Karaolis, David K. R.; Koren, Sergey; Ondov, Brian; Phillippy, Adam M.; Bergman, Nicholas H.

2014-01-01

Staphylococcus aureus subsp. aureus ATCC 25923 is commonly used as a control strain for susceptibility testing to antibiotics and as a quality control strain for commercial products. We present the completed genome sequence for the strain, consisting of the chromosome and a 27.5-kb plasmid. PMID:25377701

Development of a Multilocus Sequence Typing (MLST) scheme for Treponema pallidum subsp. pertenue: Application to yaws in Lihir Island, Papua New Guinea

PubMed Central

Godornes, Charmie; Giacani, Lorenzo; Barry, Alyssa E.; Mitja, Oriol

2017-01-01

Background Yaws is a neglected tropical disease, caused by Treponema pallidum subsp. pertenue. The disease causes chronic lesions, primarily in young children living in remote villages in tropical climates. As part of a global yaws eradication campaign initiated by the World Health Organization, we sought to develop and evaluate a molecular typing method to distinguish different strains of T. pallidum subsp. pertenue for disease control and epidemiological purposes. Methods and principal findings Published genome sequences of strains of T. pallidum subsp. pertenue and pallidum were compared to identify polymorphic genetic loci among the strains. DNA from a number of existing historical Treponema isolates, as well as a subset of samples from yaws patients collected in Lihir Island, Papua New Guinea, were analyzed using these targets. From these data, three genes (tp0548, tp0136 and tp0326) were ultimately selected to give a high discriminating capability among the T. pallidum subsp. pertenue samples tested. Intragenic regions of these three target genes were then selected to enhance the discriminating capability of the typing scheme using short readily amplifiable loci. This 3-gene multilocus sequence typing (MLST) method was applied to existing historical human yaws strains, the Fribourg-Blanc simian isolate, and DNA from 194 lesion swabs from yaws patients on Lihir Island, Papua New Guinea. Among all samples tested, fourteen molecular types were identified, seven of which were found in patient samples and seven among historical isolates or DNA. Three types (JG8, TD6, and SE7) were predominant on Lihir Island. Conclusions This MLST approach allows molecular typing and differentiation of yaws strains. This method could be a useful tool to complement epidemiological studies in regions where T. pallidum subsp. pertenue is prevalent with the overall goals of improving our understanding of yaws transmission dynamics and helping the yaws eradication campaign to succeed

Differential effects of the essential oils of Lavandula luisieri and Eryngium duriaei subsp. juresianum in cell models of two chronic inflammatory diseases.

PubMed

Rufino, Ana T; Ferreira, Isabel; Judas, Fernando; Salgueiro, Lígia; Lopes, M Celeste; Cavaleiro, Carlos; Mendes, Alexandrina F

2015-08-01

Effective drugs to treat osteoarthritis (OA) and inflammatory bowel disease (IBD) are needed. To identify essential oils (EOs) with anti-inflammatory activity in cell models of OA and IBD. EOs from Eryngium duriaei subsp. juresianum (M. Laínz) M. Laínz (Apiaceae), Laserpitium eliasii subsp. thalictrifolium Sennen & Pau (Apiaceae), Lavandula luisieri (Rozeira) Rivas-Martínez (Lamiaceae), Othantus maritimus (L.) Hoff. & Link (Asteraceae), and Thapsia villosa L. (Apiaceae) were analyzed by GC and GC/MS. The anti-inflammatory activity of EOs (5-200 μg/mL) was evaluated by measuring inducible nitric oxide synthase (iNOS) and nuclear factor-κB (NF-κB) activation (total and phosphorylated IκB-α), in primary human chondrocytes and the intestinal cell line, C2BBe1, stimulated with interleukin-1β (IL-1β) or interferon-γ (IFN-γ), IL-1β and tumor necrosis factor-α (TNF-α), respectively. The EO of L. luisieri significantly reduced iNOS (by 54.9 and 81.0%, respectively) and phosphorylated IκB-α (by 87.4% and 62.3%, respectively) in both cell models. The EO of E. duriaei subsp. juresianum caused similar effects in human chondrocytes, but was inactive in intestinal cells, even at higher concentrations. The EOs of L. eliasii subsp. thalictrifolium and O. maritimus decreased iNOS expression by 45.2 ± 8.7% and 45.2 ± 6.2%, respectively, in C2BBe1 cells and were inactive in chondrocytes. The EO of T. villosa was inactive in both cell types. This is the first study showing anti-inflammatory effects of the EOs of L. luisieri and E. duriaei subsp. juresianum. These effects are specific of the cell type and may be valuable to develop new therapies or as sources of active compounds with improved efficacy and selectivity towards OA and IBD.

Phylogeography and seed dispersal in islands: the case of Rumex bucephalophorus subsp. canariensis (Polygonaceae)

PubMed Central

Talavera, María; Navarro-Sampedro, Laura; Ortiz, Pedro L.; Arista, Montserrat

2013-01-01

Background and Aims Rumex bucephalophorus subsp. canariensis is an endemic taxon to Macaronesia with diaspore polymorphism. The origin and colonizing route of this taxon in Macaronesia was studied using molecular data and information on diaspore types. Methods Amplified fragment length polymorphism (AFLP) was used in 260 plants from 22 populations of R. bucephalophorus subsp. canariensis, four from the Madeiran archipelago and 18 from the Canary archipelago. Diaspore production was analysed in 9–50 plants from each population used for AFLP analysis. One hundred and one plants from the Madeiran archipelago and 375 plants from the Canary Islands were studied. For each plant the type of diaspore produced was recorded. Key Results Overall populations had low genetic diversity but they showed a geographical pattern of genetic diversity that was higher in the older eastern islands than in the younger western ones. Two types of dispersible diaspores were found: in the eastern Canary islands (Lanzarote, Fuerteventura and Gran Canaria), plants produced exclusively long-dispersible diaspores, whereas in the western Canary islands (Tenerife, La Gomera, El Hierro) and the Madeiran archipelago plants produced exclusively short-dispersible diaspores. Genetically, the studied populations fell into four main island groups: Lanzarote–Fuerteventura, Gran Canaria, Tenerife–El Hierro and La Gomera–Madeira archipelago. Conclusions A Moroccan origin of R. bucephalophorus subsp. canariensis is hypothesized with a colonization route from the eastern to the western islands. In addition, at least one gene flow event from La Gomera to the Madeiran archipelago has taken place. During the colonization process the type of dispersible diaspore changed so that dispersability decreased in populations of the westernmost islands. PMID:23267005

Oral supplementation with Lactobacillus delbrueckii subsp. bulgaricus 8481 enhances systemic immunity in elderly subjects.

PubMed

Moro-García, Marco Antonio; Alonso-Arias, Rebeca; Baltadjieva, Maria; Fernández Benítez, Carlos; Fernández Barrial, Manuel Amadeo; Díaz Ruisánchez, Enrique; Alonso Santos, Ricardo; Alvarez Sánchez, Magdalena; Saavedra Miján, Juan; López-Larrea, Carlos

2013-08-01

Throughout life, there is an aging of the immune system that causes impairment of its defense capability. Prevention or delay of this deterioration is considered crucial to maintain general health and increase longevity. We evaluated whether dietary supplementation with Lactobacillus delbrueckii subsp. bulgaricus 8481 could enhance the immune response in the elderly. This multi-center, double-blind, and placebo controlled study enrolled 61 elderly volunteers who were randomly assigned to receive either placebo or probiotics. Each capsule of probiotics contained at least 3 × 10(7)  L. delbrueckii subsp. bulgaricus 8481. Individuals in the study were administered three capsules per day for 6 months. Blood samples were obtained at baseline (time 0), end of month 3, and month 6. We characterized cell subpopulations, measured cytokines by flow cytometry, quantified T cell receptor excision circle (TREC) by real-time PCR (RT-PCR), and determined human β-defensin-2 (hBD-2) concentrations and human cytomegalovirus (CMV) titers by enzyme-linked immunosorbent assay (ELISA). Elderly responded to the intake of probiotic with an increase in the percentage of NK cells, an improvement in the parameters defining the immune risk profile (IRP), and an increase in the T cell subsets that are less differentiated. The probiotic group also showed decreased concentrations of the pro-inflammatory cytokine IL-8 but increased antimicrobial peptide hBD-2. These effects disappeared within 6 months of stopping the probiotic intake. Immunomodulation induced by L. delbrueckii subsp. bulgaricus 8481 could favor the maintenance of an adequate immune response, mainly by slowing the aging of the T cell subpopulations and increasing the number of immature T cells which are potential responders to new antigens.

Draft Genome Sequence of Staphylococcus cohnii subsp. urealyticus Isolated from a Healthy Dog

PubMed Central

Wigmore, Sarah M.; Wareham, David W.

2017-01-01

ABSTRACT   Staphylococcus cohnii subsp. urealyticus strain SW120 was isolated from the ear swab of a healthy dog. The isolate is resistant to methicillin and fusidic acid. The SW120 draft genome is 2,805,064 bp and contains 2,667 coding sequences, including 58 tRNAs and nine complete rRNA coding regions. PMID:28209829

Complete Whole-Genome Sequence of Salmonella enterica subsp. enterica Serovar Java NCTC5706.

PubMed

Fazal, Mohammed-Abbas; Alexander, Sarah; Burnett, Edward; Deheer-Graham, Ana; Oliver, Karen; Holroyd, Nancy; Parkhill, Julian; Russell, Julie E

2016-11-03

Salmonellae are a significant cause of morbidity and mortality globally. Here, we report the first complete genome sequence for Salmonella enterica subsp. enterica serovar Java strain NCTC5706. This strain is of historical significance, having been isolated in the pre-antibiotic era and was deposited into the National Collection of Type Cultures in 1939. © Crown copyright 2016.

Essential oil composition and enantiomeric distribution of fenchone and camphor of Lavandula cariensis and L. stoechas subsp. stoechas grown in Greece.

PubMed

Tzakou, Olga; Bazos, Ioannis; Yannitsaros, Artemios

2009-08-01

The essential oils from leaves and inflorescences of L. cariensis Boiss. and L. stoechas L. subsp. stoechas collected in Greece were analyzed by GC and GC/MS. In the inflorescences and leaves essential oils of L. cariensis the most abundant metabolite was camphor (51.8, 48.8% respectively), whereas in the essential oils of L. stoechas subsp. stoechas, the main constituents were fenchone (39.9, 21.0% respectively) and camphor (24.2, 26.3% respectively). Both enantiomers of camphor were present, whereas only (+) fenchone was detected.

Development of loop-mediated isothermal amplification for detection of Leifsonia xyli subsp. xyli in sugarcane

USDA-ARS?s Scientific Manuscript database

Ratoon stunt, caused by the xylem-limited coryneform bacterium Leifsonia xyli subsp. xyli (Lxx), is prevalent in most sugarcane-producing countries. Because the disease does not cause characteristic external symptoms, a laboratory-based technique is needed for accurate diagnosis. We developed a diag...

Development of loop-mediated isothermal amplification for detection of Leifsonia xyli subsp. xyli in sugarcane

USDA-ARS?s Scientific Manuscript database

Ratoon stunt, caused by the xylem-limited coryneform bacterium Leifsonia xyli subsp. xyli (Lxx), is prevalent in most sugarcane-planting countries. Because the disease does not cause characteristic external symptoms, a laboratory-based technique is needed for accurate diagnosis. Based on loop-mediat...

Geography of genetic differentiation in the barley wild relative Hordeum vulgare subsp. spontaneum in Jordan

USDA-ARS?s Scientific Manuscript database

Informed collecting, conservation, monitoring and utilization of genetic diversity require knowledge of the distribution and structure of genetic variation occurring in a species. Hordeum vulgare subsp. spontaneum (K. Koch) Thell., a primary wild relative of barley, is an important source of genetic...

Draft Genome Sequence of the Putrescine-Producing Strain Lactococcus lactis subsp. lactis 1AA59

PubMed Central

del Rio, Beatriz; Linares, Daniel M.; Fernandez, María; Mayo, Baltasar; Martín, M. Cruz

2015-01-01

We report here the 2,576,542-bp genome annotated draft assembly sequence of Lactococcus lactis subsp. lactis 1AA59. This strain—isolated from a traditional cheese—produces putrescine, one of the most frequently biogenic amines found in dairy products. PMID:26089428

Continuous D-lactic acid production by a novel thermotolerant Lactobacillus delbrueckii subsp. lactis QU 41.

PubMed

Tashiro, Yukihiro; Kaneko, Wataru; Sun, Yanqi; Shibata, Keisuke; Inokuma, Kentaro; Zendo, Takeshi; Sonomoto, Kenji

2011-03-01

We isolated and characterized a D-lactic acid-producing lactic acid bacterium (D-LAB), identified as Lactobacillus delbrueckii subsp. lactis QU 41. When compared to Lactobacillus coryniformis subsp. torquens JCM 1166 (T) and L. delbrueckii subsp. lactis JCM 1248 (T), which are also known as D-LAB, the QU 41 strain exhibited a high thermotolerance and produced D-lactic acid at temperatures of 50 °C and higher. In order to optimize the culture conditions of the QU 41 strain, we examined the effects of pH control, temperature, neutralizing reagent, and initial glucose concentration on D-lactic acid production in batch cultures. It was found that the optimal production of 20.1 g/l D-lactic acid was acquired with high optical purity (>99.9% of D-lactic acid) in a pH 6.0-controlled batch culture, by adding ammonium hydroxide as a neutralizing reagent, at 43 °C in MRS medium containing 20 g/l glucose. As a result of product inhibition and low cell density, continuous cultures were investigated using a microfiltration membrane module to recycle flow-through cells in order to improve D-lactic acid productivity. At a dilution rate of 0.87 h(-1), the high cell density continuous culture exhibited the highest D-lactic acid productivity of 18.0 g/l/h with a high yield (ca. 1.0 g/g consumed glucose) and a low residual glucose (<0.1 g/l) in comparison with systems published to date.

Bacillus amyloliquefaciens subsp. plantarum GR53, a potent biocontrol agent resists Rhizoctonia disease on Chinese cabbage through hormonal and antioxidants regulation.

PubMed

Kang, Sang-Mo; Radhakrishnan, Ramalingam; Lee, In-Jung

2015-10-01

The fungus Rhizoctonia solani is one of the causal agents of numerous diseases that affect crop growth and yield. The aim of this present investigation was to identify a biocontrol agent that acts against R. solani and to determine the agent's protective effect through phytohormones and antioxidant regulation in experimentally infected Chinese cabbage plants. Four rhizospheric soil bacterial isolates GR53, GR169, GR786, and GR320 were tested for their antagonistic activity against R. solani. Among these isolates, GR53 significantly suppressed fungal growth. GR53 was identified as Bacillus amyloliquefaciens subsp. plantarum by phylogenetic analysis of the 16S rDNA sequence. The biocontrol activity of B. amyloliquefaciens subsp. plantarum GR53 was tested in Chinese cabbage plants under controlled conditions. Results showed that R. solani inhibited plant growth (length, width, fresh and dry weight of leaves) by reducing chlorophyll and total phenolic content, as well as by increasing the levels of salicylic acid, jasmonic acid, abscisic acid, and DPPH scavenging activity. By regulating the levels of these compounds, the co-inoculation of B. amyloliquefaciens subsp. plantarum GR53 heightened induced systemic resistance in infected Chinese cabbage, effectively mitigating R. solani-induced damaging effects and improving plant growth. The results obtained from this study suggest that B. amyloliquefaciens subsp. plantarum GR53 is an effective biocontrol agent to prevent the damage caused by R. solani in Chinese cabbage plants.

Whole-genome sequencing of Salmonella enterica subsp. enterica serovar Cubana strains isolated from agricultural sources

USDA-ARS?s Scientific Manuscript database

We report draft genomes of Salmonella enterica subsp. enterica Serovar Cubana strain CVM42234 isolated from chick feed in 2012 and Salmonella Cubana strain 76814 isolated from swine in 2004. The genome sizes are 4,975,046 and 4,936,251 base pairs, respectively....

Identification and characterization of a NBS–LRR class resistance gene analog in Pistacia atlantica subsp. Kurdica

PubMed Central

Bahramnejad, Bahman

2014-01-01

P. atlantica subsp. Kurdica, with the local name of Baneh, is a wild medicinal plant which grows in Kurdistan, Iran. The identification of resistance gene analogs holds great promise for the development of resistant cultivars. A PCR approach with degenerate primers designed according to conserved NBS-LRR (nucleotide binding site-leucine rich repeat) regions of known disease-resistance (R) genes was used to amplify and clone homologous sequences from P. atlantica subsp. Kurdica. A DNA fragment of the expected 500-bp size was amplified. The nucleotide sequence of this amplicon was obtained through sequencing and the predicted amino acid sequence compared to the amino acid sequences of known R-genes revealed significant sequence similarity. Alignment of the deduced amino acid sequence of P. atlantica subsp. Kurdica resistance gene analog (RGA) showed strong identity, ranging from 68% to 77%, to the non-toll interleukin receptor (non-TIR) R-gene subfamily from other plants. A P-loop motif (GMMGGEGKTT), a conserved and hydrophobic motif GLPLAL, a kinase-2a motif (LLVLDDV), when replaced by IAVFDDI in PAKRGA1 and a kinase-3a (FGPGSRIII) were presented in all RGA. A phylogenetic tree, based on the deduced amino-acid sequences of PAKRGA1 and RGAs from different species indicated that they were separated in two clusters, PAKRGA1 being on cluster II. The isolated NBS analogs can be eventually used as guidelines to isolate numerous R-genes in Pistachio. PMID:27843981

A Mycobacterium avium subsp. paratuberculosis predicted serine protease is associated with acid stress and intraphagosomal survival

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) is an intracellular pathogen that persists inside host macrophages despite severe oxidative stress and nutrient deprivation. Intrabacterial pH homeostasis is vital to pathogenic mycobacteria to preserve cellular biological processes and stability of ...

Characterization of starter kimchi fermented with Leuconostoc kimchii GJ2 and its cholesterol-lowering effects in rats fed a high-fat and high-cholesterol diet.

PubMed

Jo, Se Yeon; Choi, Eun A; Lee, Jae Joon; Chang, Hae Choon

2015-10-01

The hypocholesterolemic effects of lactic acid bacteria and kimchi have been demonstrated previously. However, the kimchi fermentation process still relies on naturally present microorganisms. To obtain functional kimchi with consistent quality, we validated the capacity of Leuconostoc kimchii GJ2 as a starter culture to control kimchi fermentation. Moreover, cholesterol-lowering effects of starter kimchi as a health-promoting product were explored. Bacteriocin production by Lc. kimchii GJ2 was highly enhanced in the presence of 5% Lactobacillus sakei NJ1 cell fractions. When kimchi was fermented with bacteriocin-enhanced Lc. kimchii GJ2, Lc. kimchii GJ2 became overwhelmingly predominant (98.3%) at the end of fermentation and maintained its dominance (up to 82%) for 84 days. Growing as well as dead cells of Lc. kimchii GJ2 showed high cholesterol assimilation (in vitro). Rats were fed a high-fat and high-cholesterol diet supplemented with starter kimchi. The results showed that feeding of starter kimchi significantly reduced serum total cholesterol, triglyceride and low-density lipoprotein cholesterol levels. Additionally, atherogenic index, cardiac risk factor and triglyceride and total cholesterol levels in liver and epididymal adipose tissue decreased significantly in rats fed starter kimchi. Kimchi fermented with Lc. kimchii GJ2 as a starter culture has efficient cholesterol-lowering effects. © 2014 Society of Chemical Industry.

The Cyclic Antibacterial Peptide Enterocin AS-48: Isolation, Mode of Action, and Possible Food Applications.

PubMed

Grande Burgos, María José; Pulido, Rubén Pérez; Del Carmen López Aguayo, María; Gálvez, Antonio; Lucas, Rosario

2014-12-08

Enterocin AS-48 is a circular bacteriocin produced by Enterococcus. It contains a 70 amino acid-residue chain circularized by a head-to-tail peptide bond. The conformation of enterocin AS-48 is arranged into five alpha-helices with a compact globular structure. Enterocin AS-48 has a wide inhibitory spectrum on Gram-positive bacteria. Sensitivity of Gram-negative bacteria increases in combination with outer-membrane permeabilizing treatments. Eukaryotic cells are bacteriocin-resistant. This cationic peptide inserts into bacterial membranes and causes membrane permeabilization, leading ultimately to cell death. Microarray analysis revealed sets of up-regulated and down-regulated genes in Bacillus cereus cells treated with sublethal bacteriocin concentration. Enterocin AS-48 can be purified in two steps or prepared as lyophilized powder from cultures in whey-based substrates. The potential applications of enterocin AS-48 as a food biopreservative have been corroborated against foodborne pathogens and/or toxigenic bacteria (Listeria monocytogenes, Bacillus cereus, Staphylococcus aureus, Escherichia coli, Salmonella enterica) and spoilage bacteria (Alicyclobacillus acidoterrestris, Bacillus spp., Paenibacillus spp., Geobacillus stearothermophilus, Brochothrix thermosphacta, Staphylococcus carnosus, Lactobacillus sakei and other spoilage lactic acid bacteria). The efficacy of enterocin AS-48 in food systems increases greatly in combination with chemical preservatives, essential oils, phenolic compounds, and physico-chemical treatments such as sublethal heat, high-intensity pulsed-electric fields or high hydrostatic pressure.

The Cyclic Antibacterial Peptide Enterocin AS-48: Isolation, Mode of Action, and Possible Food Applications

PubMed Central

Grande Burgos, María José; Pérez Pulido, Rubén; López Aguayo, María del Carmen; Gálvez, Antonio; Lucas, Rosario

2014-01-01

Enterocin AS-48 is a circular bacteriocin produced by Enterococcus. It contains a 70 amino acid-residue chain circularized by a head-to-tail peptide bond. The conformation of enterocin AS-48 is arranged into five alpha-helices with a compact globular structure. Enterocin AS-48 has a wide inhibitory spectrum on Gram-positive bacteria. Sensitivity of Gram-negative bacteria increases in combination with outer-membrane permeabilizing treatments. Eukaryotic cells are bacteriocin-resistant. This cationic peptide inserts into bacterial membranes and causes membrane permeabilization, leading ultimately to cell death. Microarray analysis revealed sets of up-regulated and down-regulated genes in Bacillus cereus cells treated with sublethal bacteriocin concentration. Enterocin AS-48 can be purified in two steps or prepared as lyophilized powder from cultures in whey-based substrates. The potential applications of enterocin AS-48 as a food biopreservative have been corroborated against foodborne pathogens and/or toxigenic bacteria (Listeria monocytogenes, Bacillus cereus, Staphylococcus aureus, Escherichia coli, Salmonella enterica) and spoilage bacteria (Alicyclobacillus acidoterrestris, Bacillus spp., Paenibacillus spp., Geobacillus stearothermophilus, Brochothrix thermosphacta, Staphylococcus carnosus, Lactobacillus sakei and other spoilage lactic acid bacteria). The efficacy of enterocin AS-48 in food systems increases greatly in combination with chemical preservatives, essential oils, phenolic compounds, and physico-chemical treatments such as sublethal heat, high-intensity pulsed-electric fields or high hydrostatic pressure. PMID:25493478

Influence of Artisan Bakery- or Laboratory-Propagated Sourdoughs on the Diversity of Lactic Acid Bacterium and Yeast Microbiotas

PubMed Central

Minervini, Fabio; Lattanzi, Anna; De Angelis, Maria; Gobbetti, Marco

2012-01-01

Seven mature type I sourdoughs were comparatively back-slopped (80 days) at artisan bakery and laboratory levels under constant technology parameters. The cell density of presumptive lactic acid bacteria and related biochemical features were not affected by the environment of propagation. On the contrary, the number of yeasts markedly decreased from artisan bakery to laboratory propagation. During late laboratory propagation, denaturing gradient gel electrophoresis (DGGE) showed that the DNA band corresponding to Saccharomyces cerevisiae was no longer detectable in several sourdoughs. Twelve species of lactic acid bacteria were variously identified through a culture-dependent approach. All sourdoughs harbored a certain number of species and strains, which were dominant throughout time and, in several cases, varied depending on the environment of propagation. As shown by statistical permutation analysis, the lactic acid bacterium populations differed among sourdoughs propagated at artisan bakery and laboratory levels. Lactobacillus plantarum, Lactobacillus sakei, and Weissella cibaria dominated in only some sourdoughs back-slopped at artisan bakeries, and Leuconostoc citreum seemed to be more persistent under laboratory conditions. Strains of Lactobacillus sanfranciscensis were indifferently found in some sourdoughs. Together with the other stable species and strains, other lactic acid bacteria temporarily contaminated the sourdoughs and largely differed between artisan bakery and laboratory levels. The environment of propagation has an undoubted influence on the composition of sourdough yeast and lactic acid bacterium microbiotas. PMID:22635989

Antibiotic resistance and molecular characterization of probiotic and clinical Lactobacillus strains in relation to safety aspects of probiotics.

PubMed

Klein, Günter

2011-02-01

The evaluation of the safety of probiotic strains includes the exclusion of antibiotic resistance of clinical importance. Ninety-two strains from the genus Lactobacillus isolated from probiotics, food, and clinical sources were included in the investigation. Species tested were the L. acidophilus group, L. casei group, L. reuteri/fermentum group, and L. sakei/curvatus group. Cell and colony morphology, fermentation patterns, and growth characteristics as well as soluble whole cell proteins were analyzed. Antibiotic resistance against clinically important agents was determined by broth dilution tests. The vanA and tet genes were confirmed. Resistances occurred mainly against gentamicin, ciprofloxacin, clindamycin, sulfonamides, and, in some cases, glycopeptides. The natural glycopeptide resistance within the L. casei group and L. reuteri appears to be not of clinical relevance, as there was no vanA gene present. Therefore, the transfer of this resistance is very unlikely. Tet-(A), -(B), -(C), -(M), or -(O) gene could not be detected. The protein fingerprinting within the L. casei group proved that L. rhamnosus strains of clinical origin clustered together with probiotic strains. For safety evaluations resistance patterns of a broad range of strains are a useful criterion together with the exclusion of known resistance genes (like the vanA gene) and can be used for decision making on the safety of probiotics, both by authorization bodies and manufacturers.

Complete Genome Sequence of the Quality Control Strain Staphylococcus aureus subsp. aureus ATCC 25923.

PubMed

Treangen, Todd J; Maybank, Rosslyn A; Enke, Sana; Friss, Mary Beth; Diviak, Lynn F; Karaolis, David K R; Koren, Sergey; Ondov, Brian; Phillippy, Adam M; Bergman, Nicholas H; Rosovitz, M J

2014-11-06

Staphylococcus aureus subsp. aureus ATCC 25923 is commonly used as a control strain for susceptibility testing to antibiotics and as a quality control strain for commercial products. We present the completed genome sequence for the strain, consisting of the chromosome and a 27.5-kb plasmid. Copyright © 2014 Treangen et al.

Genome Sequence of the Rice-Pathogenic Bacterium Acidovorax avenae subsp. avenae RS-1 ▿

PubMed Central

Xie, Guan-Lin; Zhang, Guo-Qing; Liu, He; Lou, Miao-Miao; Tian, Wen-Xiao; Li, Bin; Zhou, Xue-Ping; Zhu, Bo; Jin, Gu-Lei

2011-01-01

Acidovorax avenae subsp. avenae is a phytobacterium which is the causative agent of several plant diseases with economic significance. Here, we present the draft genome sequence of strain RS-1, which was isolated from rice shoots in a rice field in China. This strain can cause bacterial stripe of rice. PMID:21742879

Splenic Abscess Caused by Streptococcus gallolyticus subsp. pasteurianus as Presentation of a Pancreatic Cancer

PubMed Central

Su, Yanli; Miao, Bin; Wang, Hong; Wang, Chao

2013-01-01

Splenic abscesses caused by Streptococcus bovis are rarely reported in the literature and are mainly seen in patients with endocarditis and associated colonic neoplasia/carcinoma. We report the first case of splenic abscess caused by Streptococcus gallolyticus subsp. pasteurianus (Streptococcus bovis biotype II/2) as presentation of a pancreatic cancer. PMID:24025909

An outbreak of fatal hemorrhagic pneumonia caused by Streptococcus equi subsp. zooepidemicus in shelter dogs.

PubMed

Byun, Jae Won; Yoon, Soon Seek; Woo, Gye-Hyeong; Jung, Byeong Yeal; Joo, Yi-Seok

2009-09-01

An outbreak of fatal hemorrhagic pneumonia with 70-90% morbidity and 50% mortality occurred in an animal shelter in Yangju, Gyeonggi Province, Korea. Clinically, the affected dogs showed severe respiratory distress within 48 h after arriving in the shelter. The dead were found mainly with nasal bleeding and hematemesis. At necropsy, hemothorax and hemorrhagic pneumonia along with severe pulmonary consolidation was observed, though histopathological analysis showed mainly hemorrhagic bronchopneumonia. Lymphoid depletion was inconsistently seen in the spleen, tonsil and bronchial lymph node. Gram-positive colonies were shown in blood vessels or parenchyma of cerebrum, lung, liver, spleen, and kidney. Also, Streptococcus (S.) equi subsp. zooepidemicus was isolated from the various organs in which the bacterium was microscopically and histologically detected. In addition, approximately 0.9 Kb specific amplicon, antiphagocytic factor H binding protein, was amplified in the bacterial isolates. In this study, we reported an outbreak of canine hemorrhagic bronchopneumonia caused by S. equi subsp. zooepidemicus in an animal shelter in Yangju, Korea.

An outbreak of fatal hemorrhagic pneumonia caused by Streptococcus equi subsp. zooepidemicus in shelter dogs

PubMed Central

Yoon, Soon-Seek; Woo, Gye-Hyeong; Jung, Byeong Yeal; Joo, Yi-Seok

2009-01-01

An outbreak of fatal hemorrhagic pneumonia with 70~90% morbidity and 50% mortality occurred in an animal shelter in Yangju, Gyeonggi Province, Korea. Clinically, the affected dogs showed severe respiratory distress within 48 h after arriving in the shelter. The dead were found mainly with nasal bleeding and hematemesis. At necropsy, hemothorax and hemorrhagic pneumonia along with severe pulmonary consolidation was observed, though histopathological analysis showed mainly hemorrhagic bronchopneumonia. Lymphoid depletion was inconsistently seen in the spleen, tonsil and bronchial lymph node. Gram-positive colonies were shown in blood vessels or parenchyma of cerebrum, lung, liver, spleen, and kidney. Also, Streptococcus (S.) equi subsp. zooepidemicus was isolated from the various organs in which the bacterium was microscopically and histologically detected. In addition, approximately 0.9 Kb specific amplicon, antiphagocytic factor H binding protein, was amplified in the bacterial isolates. In this study, we reported an outbreak of canine hemorrhagic bronchopneumonia caused by S. equi subsp. zooepidemicus in an animal shelter in Yangju, Korea. PMID:19687630

A circular genetic map of Erwinia carotovora subsp. atroseptica 3-2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nikolaichik, E.A.; Pesnyakevich, A.G.

1995-08-01

A circular genetic map of Erwinia carotovora subsp. atroseptica 3-2 was constructed on the basis of the R471a plasmid and Tn5 and Tn9 using Hfr-like donors. Forty-six genes, including phytopathogenicity genes, were located on the basis of interrupted mating experiment results and analysis of coinheritance of markers on a map of 183 min in length. The similarity and differences of chromosomal genetic maps of Erwinia genus bacteria are discussed. 23 refs., 2 figs., 4 tabs.

Streptococcus gallolyticus subsp. pasteurianus meningitis complicated by venous sinus thrombosis: A case report.

PubMed

Wardle, Martin; Mu, Andre; Tong, Steven Y C

2018-06-01

A case of Streptococcus gallolyticus subsp. pasteurianus meningitis, unusually occurring in a splenectomized patient and complicated by cerebral venous thrombosis, is described. Following presentation with meningism and diagnosis and management of S. gallolyticus meningitis, the patient presented again with a further 4days of fevers and subsequently developed left-sided paresthesias. Cerebral imaging revealed a venous thrombus in the right frontal cortical veins and left sigmoid sinus. The patient recovered following 4 weeks of intravenous ceftriaxone and anticoagulation with enoxaparin and then warfarin. Apart from the splenectomy, no underlying cause was found. The patient was commenced on life-long prophylactic amoxicillin, given appropriate vaccinations, and anticoagulated with warfarin. After initial difficulties, identification of the causative organism to the subspecies level was confirmed by analysis of short-read whole genome sequencing data. This case demonstrates two features that have not previously been reported for S. gallolyticus subsp. pasteurianus infections: splenectomy as a potential risk factor and that infection may be complicated by cerebral venous thrombosis. The resolution provided by whole genome sequencing was valuable in accurately identifying the bacterial subspecies. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Cell-Wall-Bound Proteinase of Lactobacillus delbrueckii subsp. lactis ACA-DC 178: Characterization and Specificity for β-Casein

PubMed Central

Tsakalidou, E.; Anastasiou, R.; Vandenberghe, I.; van Beeumen, J.; Kalantzopoulos, G.

1999-01-01

Lactobacillus delbrueckii subsp. lactis ACA-DC 178, which was isolated from Greek Kasseri cheese, produces a cell-wall-bound proteinase. The proteinase was removed from the cell envelope by washing the cells with a Ca2+-free buffer. The crude proteinase extract shows its highest activity at pH 6.0 and 40°C. It is inhibited by phenylmethylsulfonyl fluoride, showing that the enzyme is a serine-type proteinase. Considering the substrate specificity, the enzyme is similar to the lactococcal PI-type proteinases, since it hydrolyzes β-casein mainly and α- and κ-caseins to a much lesser extent. The cell-wall-bound proteinase from L. delbrueckii subsp. lactis ACA-DC 178 liberates four main peptides from β-casein, which have been identified. PMID:10223997

Six-Month Multicenter Study on Invasive Infections Due to Streptococcus pyogenes and Streptococcus dysgalactiae subsp. equisimilis in Argentina

PubMed Central

Lopardo, Horacio A.; Vidal, Patricia; Sparo, Monica; Jeric, Paola; Centron, Daniela; Facklam, Richard R.; Paganini, Hugo; Pagniez, N. Gaston; Lovgren, Marguerite; Beall, Bernard

2005-01-01

During a 6-month period, 95 invasive infections due to Streptococcus pyogenes and group C or group G Streptococcus dysgalactiae subsp. equisimilis were recorded from 40 centers of 16 cities in Argentina. We describe here epidemiologic data available for 55 and 19 patients, respectively, associated with invasive infections due to S. pyogenes and S. dysgalactiae subsp. equisimilis. The associated isolates and 58 additional pharyngeal isolates were genotyped and subjected to serologic and/or antibiotic susceptibility testing. Group A streptococcal emm type distribution and strain association with toxic shock appeared to differ somewhat from results found within the United States; however, serologic characterization and sof sequence typing suggested that emm types found in both countries are reflective of shared clonal types. PMID:15695683

Comparison of nine PCR primer sets designed to detect Pantoea stewartii subsp. stewartii in maize

USDA-ARS?s Scientific Manuscript database

Pantoea stewartii subsp. stewartii, the causal agent of Stewart's bacterial wilt of maize, is a major quarantine pest in maize seed. Verifying freedom from P. stewartii remains a significant hurdle in exporting corn seed from the U.S. Several PCR primer sets have been developed and suggested as bein...

Genetic analysis and antimicrobial susceptibility of Francisella noatunensis subsp. orientalis (sun. F. asiatica) isolates from fish

USDA-ARS?s Scientific Manuscript database

Francisella noatunensis subsp. orientalis (syn. F. asiatica) (Fno) is an emergent fish pathogen that causes acute to chronic disease in a wide variety of freshwater, brackish and marine fish. Due to the emergent nature of this bacterium, established protocols to measure antimicrobial susceptibility ...

Mapping of the chromosome of bacteria Erwinia carotovora subsp. atroseptica 3-2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nikolaichik, E.A.; Pesnyakevich, A.G.

1995-07-01

Two Hfr-like donor strains of bacteria Erwinia carotovora subsp. atroseptica (Eca) 3-2 were developed by integration into the chromosome of the conjugative plasmid R471a via homology with transposon Tn9. Using these and two donor strains created earlier, we constructed the genetic map of a fragment of the chromosome of strain Eca 3-2. The location of 14 loci is shown in this map. 15 refs., 3 figs., 1 tab.

Use of biochemical kinetic data to determine strain relatedness among Salmonella enterica subsp. enterica isolates.

PubMed

de la Torre, E; Tello, M; Mateu, E M; Torre, E

2005-11-01

Classical biotyping characterizes strains by creating biotype profiles that consider only positive and negative results for a predefined set of biochemical tests. This method allows Salmonella subspecies to be distinguished but does not allow serotypes and phage types to be distinguished. The objective of this study was to determine the relatedness of isolates belonging to distinct Salmonella enterica subsp. enterica serotypes by using a refined biotyping process that considers the kinetics at which biochemical reactions take place. Using a Vitek GNI+ card for the identification of gram-negative organisms, we determined the biochemical kinetic reactions (28 biochemical tests) of 135 Salmonella enterica subsp. enterica strains of pig origin collected in Spain from 1997 to 2002 (59 Salmonella serotype Typhimurium strains, 25 Salmonella serotype Typhimurium monophasic variant strains, 25 Salmonella serotype Anatum strains, 12 Salmonella serotype Tilburg strains, 7 Salmonella serotype Virchow strains, 6 Salmonella serotype Choleraesuis strains, and 1 Salmonella enterica serotype 4,5,12:-:- strain). The results were expressed as the colorimetric and turbidimetric changes (in percent) and were used to enhance the classical biotype profile by adding kinetic categories. A hierarchical cluster analysis was performed by using the enhanced profiles and resulted in 14 clusters. Six major clusters grouped 94% of all isolates with a similarity of > or =95% within any given cluster, and eight clusters contained a single isolate. The six major clusters grouped not only serotypes of the same type but also phenotypic serotype variations into individual clusters. This suggests that metabolic kinetic reaction data from the biochemical tests commonly used for classic Salmonella enterica subsp. enterica biotyping can possibly be used to determine the relatedness between isolates in an easy and timely manner.

Microsatellites for Oenothera gayleana and O. hartwegii subsp. filifolia (Onagraceae), and their utility in section Calylophus.

PubMed

Lewis, Emily M; Fant, Jeremie B; Moore, Michael J; Hastings, Amy P; Larson, Erica L; Agrawal, Anurag A; Skogen, Krissa A

2016-02-01

Eleven nuclear and four plastid microsatellite markers were screened for two gypsum endemic species, Oenothera gayleana and O. hartwegii subsp. filifolia, and tested for cross-amplification in the remaining 11 taxa within Oenothera sect. Calylophus (Onagraceae). Microsatellite markers were tested in two to three populations spanning the ranges of both O. gayleana and O. hartwegii subsp. filifolia. The nuclear microsatellite loci consisted of both di- and trinucleotide repeats with one to 17 alleles per population. Several loci showed significant deviation from Hardy-Weinberg equilibrium, which may be evidence of chromosomal rings. The plastid microsatellite markers identified one to seven haplotypes per population. The transferability of these markers was confirmed in all 11 taxa within Oenothera sect. Calylophus. The microsatellite loci characterized here are the first developed and tested in Oenothera sect. Calylophus. These markers will be used to assess whether pollinator foraging distance influences population genetic parameters in predictable ways.

Fermented milk containing Lactobacillus paracasei subsp. paracasei CNCM I-1518 reduces bacterial translocation in rats treated with carbon tetrachloride

PubMed Central

Sánchez, Elisabet; Nieto, Juan C.; Vidal, Silvia; Santiago, Alba; Martinez, Xavier; Sancho, Francesc J.; Sancho-Bru, Pau; Mirelis, Beatriz; Corominola, Helena; Juárez, Candido; Manichanh, Chaysavanh; Guarner, Carlos; Soriano, German

2017-01-01

Probiotics can prevent pathological bacterial translocation by modulating intestinal microbiota and improving the gut barrier. The aim was to evaluate the effect of a fermented milk containing Lactobacillus paracasei subsp. paracasei CNCM I-1518 on bacterial translocation in rats with carbon tetrachloride (CCl4)-induced cirrhosis. Sprague-Dawley rats treated with CCl4 were randomized into a probiotic group that received fermented milk containing Lactobacillus paracasei subsp. paracasei CNCM I-1518 in drinking water or a water group that received water only. Laparotomy was performed one week after ascites development. We evaluated bacterial translocation, intestinal microbiota, the intestinal barrier and cytokines in mesenteric lymph nodes and serum. Bacterial translocation decreased and gut dysbiosis improved in the probiotic group compared to the water group. The ileal β-defensin-1 concentration was higher and ileal malondialdehyde levels were lower in the probiotic group than in water group. There were no differences between groups in serum cytokines but TNF-α levels in mesenteric lymph nodes were lower in the probiotic group than in the water group. Fermented milk containing Lactobacillus paracasei subsp. paracasei CNCM I-1518 decreases bacterial translocation, gut dysbiosis and ileal oxidative damage and increases ileal β-defensin-1 expression in rats treated with CCl4, suggesting an improvement in the intestinal barrier integrity. PMID:28368023

Live Attenuated Tularemia Vaccines for Protection Against Respiratory Challenge With Virulent F. tularensis subsp. tularensis

PubMed Central

Jia, Qingmei; Horwitz, Marcus A.

2018-01-01

Francisella tularensis is the causative agent of tularemia and a Tier I bioterrorism agent. In the 1900s, several vaccines were developed against tularemia including the killed “Foshay” vaccine, subunit vaccines comprising F. tularensis protein(s) or lipoproteins(s) in an adjuvant formulation, and the F. tularensis Live Vaccine Strain (LVS); none were licensed in the U.S.A. or European Union. The LVS vaccine retains toxicity in humans and animals—especially mice—but has demonstrated efficacy in humans, and thus serves as the current gold standard for vaccine efficacy studies. The U.S.A. 2001 anthrax bioterrorism attack spawned renewed interest in vaccines against potential biowarfare agents including F. tularensis. Since live attenuated—but not killed or subunit—vaccines have shown promising efficacy and since vaccine efficacy against respiratory challenge with less virulent subspecies holarctica or F. novicida, or against non-respiratory challenge with virulent subsp. tularensis (Type A) does not reliably predict vaccine efficacy against respiratory challenge with virulent subsp. tularensis, the route of transmission and species of greatest concern in a bioterrorist attack, in this review, we focus on live attenuated tularemia vaccine candidates tested against respiratory challenge with virulent Type A strains, including homologous vaccines derived from mutants of subsp. holarctica, F. novicida, and subsp. tularensis, and heterologous vaccines developed using viral or bacterial vectors to express F. tularensis immunoprotective antigens. We compare the virulence and efficacy of these vaccine candidates with that of LVS and discuss factors that can significantly impact the development and evaluation of live attenuated tularemia vaccines. Several vaccines meet what we would consider the minimum criteria for vaccines to go forward into clinical development—safety greater than LVS and efficacy at least as great as LVS, and of these, several meet the

Functional cream cheese supplemented with Bifidobacterium animalis subsp. lactis DSM 10140 and Lactobacillus reuteri DSM 20016 and prebiotics.

PubMed

Speranza, Barbara; Campaniello, Daniela; Monacis, Noemi; Bevilacqua, Antonio; Sinigaglia, Milena; Corbo, Maria Rosaria

2018-06-01

The aim of this study was to develop a functional fresh cream cheese with Bifidobacterium animalis subsp. lactis DSM 10140 or Lactobacillus reuteri DSM 20016 and prebiotics (inulin, FOS and lactulose). The research was divided into two steps: in vitro evaluation of the effects of prebiotic compounds; validation at laboratory level with production of functional cream mini-cheeses. Prebiotics showed a protective effect: B. animalis subsp. lactis DSM 10140 cultivability on Petri dishes was positively influenced by lactulose, whereas fructooligosaccharides (FOS) were the prebiotic compounds able to prolong Lb. reuteri DSM 20016 cultivability. At 30 °C, a prolongation of the death time (more than 300 days) was observed, while the controls showed death time values about 100 days. At 45 °C, death time values increased from 32.2 (control) to 33, 35, and 38 days in the samples added with FOS, inulin and lactulose, respectively. Lactulose and FOS were chosen to be added to cream mini-cheeses inoculated with B. animalis subsp. lactis DSM 10140 and Lb. reuteri DSM 20016, respectively; the proposed functional cream cheese resulted in a product with favourable conditions for the viability of both probiotics which maintained cultivable cells above the recommended level during 28 days of storage at 4 °C with good sensory characteristics. Copyright © 2017 Elsevier Ltd. All rights reserved.

Fluoroquinolone Resistance in Streptococcus dysgalactiae subsp. equisimilis and Evidence for a Shared Global Gene Pool with Streptococcus pyogenes▿

PubMed Central

Pinho, M. D.; Melo-Cristino, J.; Ramirez, M.

2010-01-01

Quinolone resistance is an emerging problem in Streptococcus pyogenes, and recombination with Streptococcus dysgalactiae DNA has been implicated as a frequent mechanism leading to resistance. We have characterized a collection of S. dysgalactiae subsp. equisimilis isolates responsible for infections in humans (n = 314) and found a high proportion of levofloxacin-resistant isolates (12%). Resistance was associated with multiple emm types and genetic lineages, as determined by pulsed-field gel electrophoretic profiling. Since we could not find evidence for a role of efflux pumps in resistance, we sequenced the quinolone resistance-determining regions of the gyrA and parC genes of representative resistant and susceptible isolates. We found much greater diversity among the parC genes (19 alleles) than among the gyrA genes (5 alleles). While single mutations in either GyrA or ParC were sufficient to raise the MIC so that the strains were classified as intermediately resistant, higher-level resistance was associated with mutations in both GyrA and ParC. Evidence for recombination with S. pyogenes DNA was found in some parC alleles, but this was not exclusively associated with resistance. Our data support the existence of a common reservoir of genes conferring quinolone resistance shared between S. dysgalactiae subsp. equisimilis and S. pyogenes, while no recombination with the animal pathogen S. dysgalactiae subsp. dysgalactiae could be found. PMID:20145082

Escherichia coli, Salmonella, and Mycobacterium avium subsp. paratuberculosis in wild European starlings at a Kansas cattle feedlot.

PubMed

Gaukler, Shannon M; Linz, George M; Sherwood, Julie S; Dyer, Neil W; Bleier, William J; Wannemuehler, Yvonne M; Nolan, Lisa K; Logue, Catherine M

2009-12-01

The prevalence of Escherichia coli, Salmonella spp., and Mycobacterium avium subsp. paratuberculosis isolated from the feces of wild European starlings (Sturnus vulgaris) humanely trapped at a feedlot in central Kansas was assessed. All E. coli and Salmonella isolates recovered were tested for antimicrobial susceptibility using National Antimicrobial Resistance Monitoring System panels and the E. coli isolates were classified as to their content of genes associated with pathogenic E. coli of birds and cattle, including cvaC, iroN2, ompTp, hlyF2, eitC, iss, iutA, ireA, papC, stxI, stxII, sta, K99, F41, and eae. Escherichia coli O157:H7 and Mycobacterium avium subsp. paratuberculosis were not detected and Salmonella was isolated from only three samples, two of which displayed antimicrobial resistance. Approximately half of the E. coli isolates were resistant to antimicrobial agents with 96% showing resistance to tetracycline. Only one isolate was positive for a single gene associated with bovine pathogenic E. coli. An interesting finding of this study was that 5% of the E. coli isolates tested met the criteria established for identification as avian pathogenic E. coli (APEC). Thus these findings suggest that starlings are not a significant source of Salmonella spp., Mycobacterium avium subsp. paratuberculosis, E. coli O157, or other shiga toxin-producing E. coli in this feedlot. However, they may have the potential to spread APEC, an important pathogen of poultry and a potential pathogen to human beings.

Selection of single chain variable fragments (scFv) against Xylella fastidiosa subsp. pauca by phage display

USDA-ARS?s Scientific Manuscript database

Xylella fastidiosa is a gram-negative member of the gamma proteobacteria. Xylella fastidiosa subsp pauca causes citrus variegated chlorosis in Brazil and enjoys ‘select agent’ status in the United States. Antibody based detection assays are commercially available for Xylella fastidiosa, and are ef...

Lactobacillus delbrueckii subsp lactis CIDCA 133 modulates response of human epithelial and dendritic cells infected with Bacillus cereus.

PubMed

Rolny, I S; Tiscornia, I; Racedo, S M; Pérez, P F; Bollati-Fogolín, M

2016-11-30

It is known that probiotic microorganisms are able to modulate pathogen virulence. This ability is strain dependent and involves multiple interactions between microorganisms and relevant host's cell populations. In the present work we focus on the effect of a potentially probiotic lactobacillus strain (Lactobacillus delbrueckii subsp. lactis CIDCA 133) in an in vitro model of Bacillus cereus infection. Our results showed that infection of intestinal epithelial HT-29 cells by B. cereus induces nuclear factor kappa B (NF-κB) pathway. Noteworthy, the presence of strain L. delbrueckii subsp.lactis CIDCA 133 increases stimulation. However, B. cereus-induced interleukin (IL)-8 production by epithelial cells is partially abrogated by L. delbrueckii subsp. lactis CIDCA 133. These findings suggest that signalling pathways other than that of NF-κB are involved. In a co-culture system (HT-29 and monocyte-derived dendritic cells), B. cereus was able to translocate from the epithelial (upper) to the dendritic cell compartment (lower). This translocation was partially abrogated by the presence of lactobacilli in the upper compartment. In addition, infection of epithelial cells in the co-culture model, led to an increase in the expression of CD86 by dendritic cells. This effect could not be modified in the presence of lactobacilli. Interestingly, infection of enterocytes with B. cereus triggers production of proinflammatory cytokines by dendritic cells (IL-8, IL-6 and tumour necrosis factor alpha (TNF-α)). The production of TNF-α (a protective cytokine in B. cereus infections) by dendritic cells was increased in the presence of lactobacilli. The present work demonstrates for the first time the effect of L. delbrueckii subsp. lactis CIDCA 133, a potentially probiotic strain, in an in vitro model of B. cereus infection. The presence of the probiotic strain modulates cell response both in infected epithelial and dendritic cells thus suggesting a possible beneficial effect of

Production of Buttery-Odor Compounds and Transcriptome Response in Leuconostoc gelidum subsp. gasicomitatum LMG18811T during Growth on Various Carbon Sources

PubMed Central

Vesterinen, Sanna; Parshintsev, Jevgeni; Johansson, Per; Riekkola, Marja-Liisa; Björkroth, Johanna

2014-01-01

Leuconostoc gelidum subsp. gasicomitatum is a common spoilage bacterium in meat products packaged under oxygen-containing modified atmospheres. Buttery off-odors related to diacetyl/acetoin formation are frequently associated with the spoilage of these products. A whole-genome microarray study, together with gas chromatography (GC)-mass spectrometry (MS) analyses of the pathway end products, was performed to investigate the transcriptome response of L. gelidum subsp. gasicomitatum LMG18811T growing on semidefined media containing glucose, ribose, or inosine, which are essential carbon sources in meat. Generally, the gene expression patterns with ribose and inosine were quite similar, indicating that catabolism of ribose and nucleosides is closely linked. Diacetyl/acetoin concentrations as high as 110 or 470 μM were measured when growth was based on inosine or ribose, respectively. The gene expression results for pyruvate metabolism (upregulation of α-acetolactate synthase, downregulation of l-lactate dehydrogenase and pyruvate dehydrogenase) were as expected when diacetyl and acetoin were the end products. No diacetyl production (<7.5 μM) was detected with the glucose-containing medium, even though the cell counts of LMG18811T was 6 or 10 times higher than that on inosine or ribose, respectively. Although glucose was the most effective carbon source for the growth of L. gelidum subsp. gasicomitatum, utilization of inosine and ribose resulted in the production of the unwanted buttery-odor compounds. These results increase our understanding of which compounds are likely to enhance the formation of buttery odors during meat spoilage caused by L. gelidum subsp. gasicomitatum. PMID:25548057

Phenotypic, Genotypic, and Antimicrobial Characteristics of Streptococcus halichoeri Isolates from Humans, Proposal To Rename Streptococcus halichoeri as Streptococcus halichoeri subsp. halichoeri, and Description of Streptococcus halichoeri subsp. hominis subsp. nov., a Bacterium Associated with Human Clinical Infections.

PubMed

Shewmaker, P L; Whitney, A M; Humrighouse, B W

2016-03-01

Phenotypic, genotypic, and antimicrobial characteristics of six phenotypically distinct human clinical isolates that most closely resembled the type strain of Streptococcus halichoeri isolated from a seal are presented. Sequencing of the 16S rRNA, rpoB, sodA, and recN genes; comparative whole-genome analysis; conventional biochemical and Rapid ID 32 Strep identification methods; and antimicrobial susceptibility testing were performed on the human isolates, the type strain of S. halichoeri, and type strains of closely related species. The six human clinical isolates were biochemically indistinguishable from each other and showed 100% 16S rRNA, rpoB, sodA, and recN gene sequence similarity. Comparative 16S rRNA gene sequencing analysis revealed 98.6% similarity to S. halichoeri CCUG 48324(T), 97.9% similarity to S. canis ATCC 43496(T), and 97.8% similarity to S. ictaluri ATCC BAA-1300(T). A 3,530-bp fragment of the rpoB gene was 98.8% similar to the S. halichoeri type strain, 84.6% to the S. canis type strain, and 83.8% to the S. ictaluri type strain. The S. halichoeri type strain and the human clinical isolates were susceptible to the antimicrobials tested based on CLSI guidelines for Streptococcus species viridans group with the exception of tetracycline and erythromycin. The human isolates were phenotypically distinct from the type strain isolated from a seal; comparative whole-genome sequence analysis confirmed that the human isolates were S. halichoeri. On the basis of these results, a novel subspecies, Streptococcus halichoeri subsp. hominis, is proposed for the human isolates and Streptococcus halichoeri subsp. halichoeri is proposed for the gray seal isolates. The type strain of the novel subspecies is SS1844(T) = CCUG 67100(T) = LMG 28801(T). Copyright © 2016, American Society for Microbiology. All Rights Reserved.

The complete and fully assembled genome sequence of Aeromonas salmonicida subsp. pectinolytica and its comparative analysis with other Aeromonas species: investigation of the mobilome in environmental and pathogenic strains.

PubMed

Pfeiffer, Friedhelm; Zamora-Lagos, Maria-Antonia; Blettinger, Martin; Yeroslaviz, Assa; Dahl, Andreas; Gruber, Stephan; Habermann, Bianca H

2018-01-05

Due to the predominant usage of short-read sequencing to date, most bacterial genome sequences reported in the last years remain at the draft level. This precludes certain types of analyses, such as the in-depth analysis of genome plasticity. Here we report the finalized genome sequence of the environmental strain Aeromonas salmonicida subsp. pectinolytica 34mel, for which only a draft genome with 253 contigs is currently available. Successful completion of the transposon-rich genome critically depended on the PacBio long read sequencing technology. Using finalized genome sequences of A. salmonicida subsp. pectinolytica and other Aeromonads, we report the detailed analysis of the transposon composition of these bacterial species. Mobilome evolution is exemplified by a complex transposon, which has shifted from pathogenicity-related to environmental-related gene content in A. salmonicida subsp. pectinolytica 34mel. Obtaining the complete, circular genome of A. salmonicida subsp. pectinolytica allowed us to perform an in-depth analysis of its mobilome. We demonstrate the mobilome-dependent evolution of this strain's genetic profile from pathogenic to environmental.

Genetic diversity of clinical Mycobacterium avium subsp. hominissuis and Mycobacterium intracellulare isolates causing pulmonary diseases recovered from different geographical regions.

PubMed

Ichikawa, Kazuya; van Ingen, Jakko; Koh, Won-Jung; Wagner, Dirk; Salfinger, Max; Inagaki, Takayuki; Uchiya, Kei-Ichi; Nakagawa, Taku; Ogawa, Kenji; Yamada, Kiyofumi; Yagi, Tetsuya

2015-12-01

Mycobacterium avium complex (MAC) infections are increasing annually in many countries. MAC strains are the most common nontuberculous mycobacterial pathogens isolated from respiratory samples and predominantly consist of two species, Mycobacterium avium and Mycobacterium intracellulare. The aim of this study was to analyze the molecular epidemiology and genetic backgrounds of clinical MAC isolates collected from The Netherlands, Germany, United States, Korea and Japan. Variable numbers of tandem repeats (VNTR) analysis was used to examine the genetic relatedness of clinical isolates of M. avium subsp. hominissuis (n=261) and M. intracellulare (n=116). Minimum spanning tree and unweighted pair group method using arithmetic averages analyses based on the VNTR data indicated that M. avium subsp. hominissuis isolates from Japan shared a high degree of genetic relatedness with Korean isolates, but not with isolates from Europe or the United States, whereas M. intracellulare isolates did not show any specific clustering by geographic origin. The findings from the present study indicate that strains of M. avium subsp. hominissuis, but not M. intracellulare, exhibit geographical differences in genetic diversity and imply that MAC strains may have different sources, routes of transmission and perhaps clinical manifestations. Copyright © 2015 Elsevier B.V. All rights reserved.

Mycobacterium avium subsp. hominissuis infection in swine associated with peat used for bedding.

PubMed

Johansen, Tone Bjordal; Agdestein, Angelika; Lium, Bjørn; Jørgensen, Anne; Djønne, Berit

2014-01-01

Mycobacterium avium subsp. hominissuis is an environmental bacterium causing opportunistic infections in swine, resulting in economic losses. Additionally, the zoonotic aspect of such infections is of concern. In the southeastern region of Norway in 2009 and 2010, an increase in condemnation of pig carcasses with tuberculous lesions was seen at the meat inspection. The use of peat as bedding in the herds was suspected to be a common factor, and a project examining pigs and environmental samples from the herds was initiated. Lesions detected at meat inspection in pigs originating from 15 herds were sampled. Environmental samples including peat from six of the herds and from three peat production facilities were additionally collected. Samples were analysed by culture and isolates genotyped by MLVA analysis. Mycobacterium avium subsp. hominissuis was detected in 35 out of 46 pigs, in 16 out of 20 samples of peat, and in one sample of sawdust. MLVA analysis demonstrated identical isolates from peat and pigs within the same farms. Polyclonal infection was demonstrated by analysis of multiple isolates from the same pig. To conclude, the increase in condemnation of porcine carcasses at slaughter due to mycobacteriosis seemed to be related to untreated peat used as bedding.

Mycobacterium avium subsp. hominissuis Infection in Swine Associated with Peat Used for Bedding

PubMed Central

Johansen, Tone Bjordal; Lium, Bjørn; Jørgensen, Anne; Djønne, Berit

2014-01-01

Mycobacterium avium subsp. hominissuis is an environmental bacterium causing opportunistic infections in swine, resulting in economic losses. Additionally, the zoonotic aspect of such infections is of concern. In the southeastern region of Norway in 2009 and 2010, an increase in condemnation of pig carcasses with tuberculous lesions was seen at the meat inspection. The use of peat as bedding in the herds was suspected to be a common factor, and a project examining pigs and environmental samples from the herds was initiated. Lesions detected at meat inspection in pigs originating from 15 herds were sampled. Environmental samples including peat from six of the herds and from three peat production facilities were additionally collected. Samples were analysed by culture and isolates genotyped by MLVA analysis. Mycobacterium avium subsp. hominissuis was detected in 35 out of 46 pigs, in 16 out of 20 samples of peat, and in one sample of sawdust. MLVA analysis demonstrated identical isolates from peat and pigs within the same farms. Polyclonal infection was demonstrated by analysis of multiple isolates from the same pig. To conclude, the increase in condemnation of porcine carcasses at slaughter due to mycobacteriosis seemed to be related to untreated peat used as bedding. PMID:25431762

Effect of Turbulent-Flow Pasteurization on Survival of Mycobacterium avium subsp. paratuberculosis Added to Raw Milk

PubMed Central

Pearce, Lindsay E.; Truong, H. Tuan; Crawford, Robert A.; Yates, Gary F.; Cavaignac, Sonia; de Lisle, Geoffrey W.

2001-01-01

A pilot-scale pasteurizer operating under validated turbulent flow (Reynolds number, 11,050) was used to study the heat sensitivity of Mycobacterium avium subsp. paratuberculosis added to raw milk. The ATCC 19698 type strain, ATCC 43015 (Linda, human isolate), and three bovine isolates were heated in raw whole milk for 15 s at 63, 66, 69, and 72°C in duplicate trials. No strains survived at 72°C for 15 s; and only one strain survived at 69°C. Means of pooled D values (decimal reduction times) at 63 and 66°C were 15.0 ± 2.8 s (95% confidence interval) and 5.9 ± 0.7 s (95% confidence interval), respectively. The mean extrapolated D72°C was <2.03 s. This was equivalent to a >7 log10 kill at 72°C for 15 s (95% confidence interval). The mean Z value (degrees required for the decimal reduction time to traverse one log cycle) was 8.6°C. These five strains showed similar survival whether recovery was on Herrold's egg yolk medium containing mycobactin or by a radiometric culture method (BACTEC). Milk was inoculated with fresh fecal material from a high-level fecal shedder with clinical Johne's disease. After heating at 72°C for 15 s, the minimum M. avium subsp. paratuberculosis kill was >4 log10. Properly maintained and operated equipment should ensure the absence of viable M. avium subsp. paratuberculosis in retail milk and other pasteurized dairy products. An additional safeguard is the widespread commercial practice of pasteurizing 1.5 to 2° above 72°C. PMID:11525992

Draft Genome Sequence of Staphylococcus carnosus subsp. utilis LTH 7013, Isolated from South Tyrolean Ham.

PubMed

Müller, Anne; Huptas, Christopher; Wenning, Mareike; Schmidt, Herbert; Weiss, Agnes

2015-05-14

Staphylococcus carnosus is used as a starter culture in meat fermentation, where it contributes to color formation and produces aromatic compounds. Here, we report the first draft genome sequence of an S. carnosus subsp. utilis strain, LTH 7013, isolated from South Tyrolean ham, with potential application as a starter culture. Copyright © 2015 Müller et al.

Re-evaluation of the taxonomy of the Mitis group of the genus Streptococcus based on whole genome phylogenetic analyses, and proposed reclassification of Streptococcus dentisani as Streptococcus oralis subsp. dentisani comb. nov., Streptococcus tigurinus as Streptococcus oralis subsp. tigurinus comb. nov., and Streptococcus oligofermentans as a later synonym of Streptococcus cristatus.

PubMed

Jensen, Anders; Scholz, Christian F P; Kilian, Mogens

2016-11-01

The Mitis group of the genus Streptococcus currently comprises 20 species with validly published names, including the pathogen S. pneumoniae. They have been the subject of much taxonomic confusion, due to phenotypic overlap and genetic heterogeneity, which has hampered a full appreciation of their clinical significance. The purpose of this study was to critically re-examine the taxonomy of the Mitis group using 195 publicly available genomes, including designated type strains for phylogenetic analyses based on core genomes, multilocus sequences and 16S rRNA gene sequences, combined with estimates of average nucleotide identity (ANI) and in silico and in vitro analyses of specific phenotypic characteristics. Our core genomic phylogenetic analyses revealed distinct clades that, to some extent, and from the clustering of type strains represent known species. However, many of the genomes have been incorrectly identified adding to the current confusion. Furthermore, our data show that 16S rRNA gene sequences and ANI are unsuitable for identifying and circumscribing new species of the Mitis group of the genus Streptococci. Based on the clustering patterns resulting from core genome phylogenetic analysis, we conclude that S. oligofermentans is a later synonym of S. cristatus. The recently described strains of the species Streptococcus dentisani includes one previously referred to as 'S. mitis biovar 2'. Together with S. oralis, S. dentisani and S. tigurinus form subclusters within a coherent phylogenetic clade. We propose that the species S. oralis consists of three subspecies: S. oralis subsp. oralis subsp. nov., S. oralis subsp. tigurinus comb. nov., and S. oralis subsp. dentisani comb. nov.

Genome sequences of Salmonella enterica subsp. Kentucky ST152 isolated from dairy cows in the United States

USDA-ARS?s Scientific Manuscript database

Salmonella enterica subsp. enterica serovar Kentucky is frequently isolated from dairy cows in the United States, but is an infrequent cause of human salmonellosis. To investigate the genomic features of S. Kentucky strains isolated from these animals, genomes of eight isolates were sequenced and ad...

Development of single chain variable fragment (scFv) antibodies against Xylella fastidiosa subsp. pauca by phage display

USDA-ARS?s Scientific Manuscript database

Xylella fastidiosa is a member of the gamma proteobacteria. It is fastidious, insect-vectored and xylem-limited and causes a variety of diseases, some severe, on a wide range of economically important perennial crops, including grape and citrus. Xylella fastidiosa subsp pauca causes citrus variegat...

Enhancing the Sweetness of Yoghurt through Metabolic Remodeling of Carbohydrate Metabolism in Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus.

PubMed

Sørensen, Kim I; Curic-Bawden, Mirjana; Junge, Mette P; Janzen, Thomas; Johansen, Eric

2016-06-15

Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus are used in the fermentation of milk to produce yoghurt. These species normally metabolize only the glucose moiety of lactose, secreting galactose and producing lactic acid as the main metabolic end product. We used multiple serial selection steps to isolate spontaneous mutants of industrial strains of S. thermophilus and L. delbrueckii subsp. bulgaricus that secreted glucose rather than galactose when utilizing lactose as a carbon source. Sequencing revealed that the S. thermophilus strains had mutations in the galKTEM promoter, the glucokinase gene, and genes encoding elements of the glucose/mannose phosphotransferase system (PTS). These strains metabolize galactose but are unable to phosphorylate glucose internally or via the PTS. The L. delbrueckii subsp. bulgaricus mutants had mutations in genes of the glucose/mannose PTS and in the pyruvate kinase gene. These strains cannot grow on exogenous glucose but are proficient at metabolizing internal glucose released from lactose by β-galactosidase. The resulting strains can be combined to ferment milk, producing yoghurt with no detectable lactose, moderate levels of galactose, and high levels of glucose. Since glucose tastes considerably sweeter than either lactose or galactose, the sweetness of the yoghurt is perceptibly enhanced. These strains were produced without the use of recombinant DNA technology and can be used for the industrial production of yoghurt with enhanced intrinsic sweetness and low residual levels of lactose. Based on a good understanding of the physiology of the lactic acid bacteria Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, we were able, by selecting spontaneously occurring mutants, to change dramatically the metabolic products secreted into the growth medium. These mutants consume substantially more of the lactose, metabolize some of the galactose, and secrete the remaining galactose

Enhancing the Sweetness of Yoghurt through Metabolic Remodeling of Carbohydrate Metabolism in Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus

PubMed Central

Sørensen, Kim I.; Curic-Bawden, Mirjana; Junge, Mette P.; Janzen, Thomas

2016-01-01

ABSTRACT Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus are used in the fermentation of milk to produce yoghurt. These species normally metabolize only the glucose moiety of lactose, secreting galactose and producing lactic acid as the main metabolic end product. We used multiple serial selection steps to isolate spontaneous mutants of industrial strains of S. thermophilus and L. delbrueckii subsp. bulgaricus that secreted glucose rather than galactose when utilizing lactose as a carbon source. Sequencing revealed that the S. thermophilus strains had mutations in the galKTEM promoter, the glucokinase gene, and genes encoding elements of the glucose/mannose phosphotransferase system (PTS). These strains metabolize galactose but are unable to phosphorylate glucose internally or via the PTS. The L. delbrueckii subsp. bulgaricus mutants had mutations in genes of the glucose/mannose PTS and in the pyruvate kinase gene. These strains cannot grow on exogenous glucose but are proficient at metabolizing internal glucose released from lactose by β-galactosidase. The resulting strains can be combined to ferment milk, producing yoghurt with no detectable lactose, moderate levels of galactose, and high levels of glucose. Since glucose tastes considerably sweeter than either lactose or galactose, the sweetness of the yoghurt is perceptibly enhanced. These strains were produced without the use of recombinant DNA technology and can be used for the industrial production of yoghurt with enhanced intrinsic sweetness and low residual levels of lactose. IMPORTANCE Based on a good understanding of the physiology of the lactic acid bacteria Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, we were able, by selecting spontaneously occurring mutants, to change dramatically the metabolic products secreted into the growth medium. These mutants consume substantially more of the lactose, metabolize some of the galactose, and secrete the

Proposal to reclassify Roseivirga ehrenbergii (Nedashkovskaya et al., 2008) as Roseivirga seohaensis comb. nov., description of Roseivirga seohaensis subsp. aquiponti subsp. nov. and emendation of the genus Roseivirga.

PubMed

Selvaratnam, Chitra; Thevarajoo, Suganthi; Goh, Kian Mau; Chan, Kok-Gan; Chong, Chun Shiong

2016-12-01

The genus Roseivirga currently includes five species: Roseivirga ehrenbergii, R. echinicomitans, R. spongicola, R. marina and R. maritima. Marinicola seohaensis SW-152T was renamed as Roseivirgaseohaensis SW-152T and then reclassified again as a later heterotypic synonym of R. ehrenbergii KMM 6017T. In this study, based on average nucleotide identity and digital DNA-DNA hybridization values obtained from in silico methods, together with fatty acid analyses and biochemical tests, we propose to reclassify R. ehrenbergii SW-152 as Roseivirga seohaensis comb. nov. (type strain SW-152T=KCTC 1231T=JCM 12600T). In this work, a Gram-negative, rod-shaped, aerobic and pink-pigmented strain designated as D-25T was isolated from seawater (Desaru Beach, Johor, Malaysia). The 16S rRNA gene analysis revealed that strain D-25T was related to the genus Roseivirga. Strain D-25T was found most closely related to R. seohaensis SW-152T based on average nucleotide identity and digital DNA-DNA hybridization values, phenotypic and chemotaxonomic analyses, indicating that these strains belong to the same species. Thus, it is proposed to split the species R.oseivirga seohaensis into two novel subspecies, Roseivirga seohaensissubsp. seohaensis subsp. nov. (type strain SW-152T=KCTC 12312T=JCM 12600T) and Roseivirga seohaensissubsp. aquiponti subsp. nov. (type strain D-25T=KCTC 42709T=DSM 101709T) and to emend the description of the genus Roseivirga.

Marked Differences in Mucosal Immune Responses Induced in Ileal versus Jejunal Peyer’s Patches to Mycobacterium avium subsp. paratuberculosis Secreted Proteins following Targeted Enteric Infection in Young Calves

PubMed Central

Facciuolo, Antonio; Gonzalez-Cano, Patricia; Napper, Scott; Griebel, Philip J.

2016-01-01

In cattle, Mycobacterium avium subsp. paratuberculosis infection is primarily mediated through M cells overlying Peyer’s patches (PP) in the ileum. The capacity of M. avium subsp. paratuberculosis to invade ileal PP (IPP) versus discrete PP in the jejunum (JPP) and subsequent differences in mucosal immune responses were investigated. Intestinal segments were surgically prepared in both mid-jejunum, containing two JPPs, and in terminal small intestine containing continuous IPP. M. avium subsp. paratuberculosis (109 CFU) was injected into the lumen of half of each intestinal segment when calves were 10–14 days-old and infection confirmed 1–2 months later by PCR and immunohistochemistry. Thirteen recombinant M. avium subsp. paratuberculosis proteins, previously identified as immunogenic, were used to analyze pathogen-specific B- and T-cell responses in PP and mesenteric lymph nodes. IgA plasma cell responses to 9 of 13 recombinant proteins were detected in JPP but not in IPP. Secretory IgA reacting in ELISA with 9 of the 13 recombinant proteins was detected in luminal contents from both jejunal and ileal segments. These observations support the conclusion that pathogen-specific IgA B cells were induced in JPP but not IPP early after a primary infection. The presence of secretory IgA in intestinal contents is consistent with dissemination of IgA plasma cells from the identified mucosa-associated immune induction sites. This is the first direct evidence for M. avium subsp. paratuberculosis uptake by bovine JPP and for local induction of pathogen-specific IgA plasma cell responses after enteric infection. We also provide evidence that bacterial invasion of IPP, a primary B lymphoid tissue, provides a novel strategy to evade induction of mucosal immune responses. Over 60% of PPs in the newborn calf small intestine is primary lymphoid tissue, which has significant implications when designing oral vaccines or diagnostic tests to detect early M. avium subsp

Draft Genome Sequence of Bifidobacterium animalis subsp. lactis Strain CECT 8145, Able To Improve Metabolic Syndrome In Vivo.

PubMed

Chenoll, E; Codoñer, F M; Silva, A; Martinez-Blanch, J F; Martorell, P; Ramón, D; Genovés, S

2014-03-27

Bifidobacterium animalis subsp. lactis strain CECT 8145 is able to reduce body fat content and improve metabolic syndrome biomarkers. Here, we report the draft genome sequence of this strain, which may provide insights into its safety status and functional role.

Draft Genome Sequences of 18 Salmonella enterica subsp. enterica Serovar Oranienburg Strains Isolated from Rivers in Northwestern Mexico

PubMed Central

Casteñeda-Ruelas, Gloria M.; Carreón-Gaxiola, César; Castelán-Sánchez, Hugo G.; Acatzi-Silva, Abraham; Romero-Martínez, Salvador; García-Molina, Alejandra

2017-01-01

ABSTRACT Salmonella enterica subsp. enterica serovar Oranienburg is recognized as a foodborne pathogen widely distributed in the environment. Here, we report 18 draft genomes of S. Oranienburg strains isolated from rivers in the northwestern region of Mexico. PMID:28280020

Microsatellites for Oenothera gayleana and O. hartwegii subsp. filifolia (Onagraceae), and their utility in section Calylophus1

PubMed Central

Lewis, Emily M.; Fant, Jeremie B.; Moore, Michael J.; Hastings, Amy P.; Larson, Erica L.; Agrawal, Anurag A.; Skogen, Krissa A.

2016-01-01

Premise of the study: Eleven nuclear and four plastid microsatellite markers were screened for two gypsum endemic species, Oenothera gayleana and O. hartwegii subsp. filifolia, and tested for cross-amplification in the remaining 11 taxa within Oenothera sect. Calylophus (Onagraceae). Methods and Results: Microsatellite markers were tested in two to three populations spanning the ranges of both O. gayleana and O. hartwegii subsp. filifolia. The nuclear microsatellite loci consisted of both di- and trinucleotide repeats with one to 17 alleles per population. Several loci showed significant deviation from Hardy–Weinberg equilibrium, which may be evidence of chromosomal rings. The plastid microsatellite markers identified one to seven haplotypes per population. The transferability of these markers was confirmed in all 11 taxa within Oenothera sect. Calylophus. Conclusions: The microsatellite loci characterized here are the first developed and tested in Oenothera sect. Calylophus. These markers will be used to assess whether pollinator foraging distance influences population genetic parameters in predictable ways. PMID:26949578

[A comparison of the properties of bacteriocins formed by Lactococcus lactis subsp. lactis strains of diverse origin].

PubMed

Stoianova, L G; Egorov, N S; Fedorova, G B; Katrukha, G S; Netrusov, A I

2007-01-01

Bacteriocins formed by four strains of Lactococcus lactis subsp. lactis have been studied and compared: 729 (a natural strain isolated from milk), 1605 (a mutant of strain 729), F-116 (a recombinant obtained by fusing of protoplasts of the two related strain 729 and 1605), and a nisin-forming strain obtained by adaptive selection at Moscow State University. Antimicrobial activity studies revealed differences between the strains in the effects on individual groups of microorganisms; the activities of the strains were also distinct from that of Nisaplin (a commercial preparation of the bacteriocin nisin). Methods for isolation and purification of bacteriocins have been developed, making it possible to obtain individual components of antibiotic complexes as chromatographically pure preparations. Bacteriocins formed by the strains of Lactococcus lactis subsp. lactis have been identified and differences in their biological and physicochemical properties, established. A novel potent broad-spectrum antibiotic substance distinct from nisin has been isolated from the recombinant strain F-116.

Efficacy of florfenicol for control of mortality with Francisella noatunensis subsp. orientalis in Nile tilapia, oreochromis niloticus (L.)

USDA-ARS?s Scientific Manuscript database

Francisella noatunensis subsp. orientalis (Fno) (syn. F. asiatica) is an emergent Gram-negative facultative intracellular bacterium. Although it is considered one of the most pathogenic bacteria in fish, there are no commercially available treatments of vaccines. The objective of this project was ...

Comparison of fecal DNA extraction kits for the detection of Mycobacterium avium subsp. paratuberculosis by polymerase chain reaction

USDA-ARS?s Scientific Manuscript database

Culture of Mycobacterium avium subsp. paratuberculosis (MAP) from feces has been considered the gold standard for the diagnosis of paratuberculosis for many years. However, direct fecal PCR is becoming more widely used today, demonstrating similar sensitivity and specificity to culture. To ensure ef...

Antigenicity of recombinant maltose binding protein-Mycobacterium avium subsp. paratuberculosis fusion proteins with and without factor Xa cleaving

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp paratuberculosis (MAP) causes Johne’s disease (JD) in ruminants. Proteomic studies have shown that MAP expresses certain proteins when exposed to in vitro physiological stress conditions similar to the conditions experienced within a host during natural infection. Such prot...

Identification of quantitative trait loci for Goss’s wilt of maize caused by Clavibacter michiganensis subsp. nebraskensis

USDA-ARS?s Scientific Manuscript database

Since its discovery in 1969, Goss’s wilt, a vascular disease caused by the gram-positive bacterium Clavibacter michiganensis subsp. nebraskensis (Cmn), has emerged as one of the top four diseases of maize in the United States and Ontario, Canada. No source of complete resistance has been described f...

Stimulation of indigenous lactobacilli by fermented milk prepared with probiotic bacterium, Lactobacillus delbrueckii subsp. bulgaricus strain 2038, in the pigs.

PubMed

Ohashi, Yuji; Tokunaga, Makoto; Taketomo, Naoki; Ushida, Kazunari

2007-02-01

The aim of this study was to evaluate the effect of feeding yoghurt, prepared with Lactobacillus delbrueckii subsp. bulgaricus strain 2038, on indigenous lactobacilli in the pig cecum. Three female pigs fistulated at the cecum were fed 250 g of this yoghurt that contained over 10(11) colony-forming units of L. delbrueckii subsp. bulgaricus strain 2038 with their daily meal for 2 wk. The relative abundance and the composition of cecal lactobacilli was monitored by analysis of bacterial 16S rDNA with real time PCR and amplified bacterial rDNA restriction analysis using Lactobacillus-group specific primers, respectively, for 2 wk prior to, at the end of 2 wk of and 2 wk after the administration of this yoghurt. The relative abundance of lactobacilli was significantly increased by feeding yoghurt (p<0.01), although the bacterial 16S rDNA matching L. delbrueckii subsp. bulgaricus strain 2038 was not detected by amplified bacterial rDNA restriction analysis during this study. The number of operational taxonomic units (OTUs) detected was increased with feeding of the yoghurt in all pigs. At the same time, the estimated cell number of each OTU was increased with feeding of the yoghurt. It is demonstrated that continuous consumption of the probiotic lactobacilli will stimulate the growth of some indigenous lactobacilli and alter the composition of the lactobacilli.

A Multidirectional Perspective for Novel Functional Products: In vitro Pharmacological Activities and In silico Studies on Ononis natrix subsp. hispanica

PubMed Central

Yerlikaya, Serife; Zengin, Gokhan; Mollica, Adriano; Baloglu, Mehmet C.; Celik Altunoglu, Yasemin; Aktumsek, Abdurrahman

2017-01-01

The genus Ononis has important value as traditional drugs and foods. In the present work, we aimed to assess the chemical profiles and biological effects of Ononis natrix subsp. hispanica extracts (ethyl acetate, methanol, and water). For chemical profile, total and individual phenolic components were detected. For biological effects, antioxidant (DPPH, ABTS, CUPRAC, FRAP, phosphomolybdenum, and metal chelating assays), enzyme inhibitory (against cholinesterase, tyrosinase, α-amylase and α-glucosidase), antimicrobial, DNA protection and cytotoxic abilities were tested. The predominant phenolics were apigenin, luteolin, and quercetin in the tested extracts. Generally, the ethyl acetate and methanol extracts were noted as the most active in the antioxidant and enzyme inhibitory assays. Water extract with different concentrations indicated high level of DNA protection activity. Methanol and ethyl acetate extracts showed antibacterial effect against to Staphylococcus aureus and Staphylococcus epidermidis strains. The cytotoxic effects of O. natrix subsp. hispanica extracts on the survival of HeLa and PC3 cells were determined by MTT cell viability assay. Water and methanol extracts caused initiation of apoptosis for PC3 cell line. Furthermore, molecular docking was performed to better understand interactions between dominant phenolic compounds and selected enzymes. Our results clearly indicate that O. natrix subsp. hispanica could be considered a potential candidate for designing novel pharmaceuticals, cosmeceuticals and nutraceuticals. PMID:28919860

Essential oil of Juniperus communis subsp. alpina (Suter) Čelak needles: chemical composition, antifungal activity and cytotoxicity.

PubMed

Cabral, C; Francisco, V; Cavaleiro, C; Gonçalves, M J; Cruz, M T; Sales, F; Batista, M T; Salgueiro, L

2012-09-01

Essential oils are known to possess antimicrobial activity against a wide spectrum of bacteria and fungi. In the present work the composition and the antifungal activity of the oils of Juniperus communis subsp. alpina (Suter) Čelak were evaluated. Moreover, the skin cytotoxicity, at concentrations showing significant antifungal activity, was also evaluated. The oils were isolated by hydrodistillation and analysed by gas chromatography and gas chromatography-mass spectrometry. Minimal inhibitory concentration (MIC) and minimal lethal concentration (MLC) were used to evaluate the antifungal activity of the oil against dermatophytes (Epidermophyton floccosum, Microsporum canis, M. gypseum, Trichophyton mentagrophytes, T. mentagrophytes var. interdigitale, T. rubrum, T. verrucosum), yeasts (Candida albicans, C. guillermondii, C. krusei, C. parapsilosis, C. tropicalis, Cryptococcus neoformans) and Aspergillus species (Aspergillus flavus, A. fumigatus, A. niger). Cytotoxicity was tested in HaCaT keratinocytes through the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Essential oil of J. communis subsp. alpina needles was predominantly composed of monoterpene hydrocarbons (78.4%), with the main compounds being sabinene (26.2%), α-pinene (12-9%) and limonene (10.4%). Results concerning the antifungal activity demonstrated the potential of needle oil against dermatophytes, particularly for Microsporum canis and Trichophyton rubrum with MIC and MLC of 0.32 μL/mL. Furthermore, evaluation of cell viability showed no significant cytotoxicity in HaCaT keratinocytes at concentrations between 0.32 and 0.64 μL/mL. These results show that it is possible to find appropriate doses of J. communis subsp. alpina oil with both antifungal activity and a very low detrimental effect on keratinocytes. Copyright © 2012 John Wiley & Sons, Ltd.

Rapid Real-Time PCR Assay for Detection and Quantitation of Mycobacterium avium subsp. paratuberculosis DNA in Artificially Contaminated Milk

PubMed Central

O'Mahony, Jim; Hill, Colin

2004-01-01

Using fluorescence resonance energy transfer technology and Lightcycler analysis, we developed a real-time PCR assay with primers and probes designed by using IS900 which allowed rapid detection of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk. Initially, the PCR parameters (including primer and probe levels, assay volume, Mg2+ concentration, and annealing temperature) were optimized. Subsequently, the quantitative ability of the assay was tested and was found to be accurate over a broad linear range (3 × 106 to 3 × 101 copies). The assay sensitivity when purified DNA was used was determined to be as low as five copies, with excellent reproducibility. A range of DNA isolation strategies was developed for isolating M. avium subsp. paratuberculosis DNA from spiked milk, the most effective of which involved the use of 50 mM Tris HCl, 10 mM EDTA, 2% Triton X-100, 4 M guanidinium isothiocyante, and 0.3 M sodium acetate combined with boiling, physical grinding, and nucleic acid spin columns. When this technique was used in conjunction with the real-time PCR assay, it was possible to consistently detect <100 organisms per ml of milk (equivalent to 2,000 organisms per 25 ml). Furthermore, the entire procedure (extraction and PCR) was performed in less than 3 h and was successfully adapted to quantify M. avium subsp. paratuberculosis in spiked milk from heavily and mildly contaminated samples. PMID:15294786

The Host Genotype and Environment Affect Strain Types of Bifidobacterium longum subsp. longum Inhabiting the Intestinal Tracts of Twins.

PubMed

Zhang, Min; Hang, Xiaomin; Tan, Jing; Yang, Hong

2015-07-01

To investigate the influences of host genotype and environment on Bifidobacterium longum subsp. longum inhabiting human intestines at the strain level, six pairs of twins, divided into two groups (children and adults), were recruited. Each group consisted of two monozygotic (MZ) twin pairs and one dizygotic (DZ) twin pair. Child twins had been living together from birth, while adult twins had been living separately for 5 to 10 years. A total of 345 B. longum subsp. longum isolates obtained from 60 fecal samples from these twins were analyzed by multilocus sequence typing (MLST), and 35 sequence types (STs) were finally acquired. Comparison of strains within and between the twin pairs showed that no strains with identical STs were observed between unrelated individuals or within adult DZ twin pairs. Eight STs were found to be monophyletic, existing within MZ twins and child DZ twins. The similarity of strain types within child cotwins was significantly higher than that within adult cotwins, which indicated that environment was one of the important determinants in B. longum subsp. longum strain types inhabiting human intestines. However, although these differences between MZ and DZ twins were observed, it is still difficult to reach an exact conclusion about the impact of host genotype. This is mainly because of the limited number of subjects tested in the present study and the lack of strain types tracing in the same twin pairs from birth until adulthood. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Environmental Survival of Mycobacterium avium subsp. paratuberculosis in Different Climatic Zones of Eastern Australia

PubMed Central

Begg, Douglas J.; Dhand, Navneet K.; Watt, Bruce; Whittington, Richard J.

2014-01-01

The duration of survival of both the S and C strains of Mycobacterium avium subsp. paratuberculosis in feces was quantified in contrasting climatic zones of New South Wales, Australia, and detailed environmental temperature data were collected. Known concentrations of S and C strains in feces placed on soil in polystyrene boxes were exposed to the environment with or without the provision of shade (70%) at Bathurst, Armidale, Condobolin, and Broken Hill, and subsamples taken every 2 weeks were cultured for the presence of M. avium subsp. paratuberculosis. The duration of survival ranged from a minimum of 1 week to a maximum of 16 weeks, and the provision of 70% shade was the most important factor in extending the survival time. The hazard of death for exposed compared to shaded samples was 20 and 9 times higher for the S and C strains, respectively. Site did not affect the survival of the C strain, but for the S strain, the hazard of death was 2.3 times higher at the two arid zone sites (Broken Hill and Condobolin) than at the two temperate zone sites (Bathurst and Armidale). Temperature measurements revealed maximum temperatures exceeding 60°C and large daily temperature ranges at the soil surface, particularly in exposed boxes. PMID:24463974

Causal Relationship between Microbial Ecology Dynamics and Proteolysis during Manufacture and Ripening of Protected Designation of Origin (PDO) Cheese Canestrato Pugliese

PubMed Central

De Pasquale, Ilaria; Calasso, Maria; Mancini, Leonardo; Ercolini, Danilo; La Storia, Antonietta; De Angelis, Maria; Gobbetti, Marco

2014-01-01

Pyrosequencing of the 16S rRNA gene, community-level physiological profiles determined by the use of Biolog EcoPlates, and proteolysis analyses were used to characterize Canestrato Pugliese Protected Designation of Origin (PDO) cheese. The number of presumptive mesophilic lactococci in raw ewes' milk was higher than that of presumptive mesophilic lactobacilli. The numbers of these microbial groups increased during ripening, showing temporal and numerical differences. Urea-PAGE showed limited primary proteolysis, whereas the analysis of the pH 4.6-soluble fraction of the cheese revealed that secondary proteolysis increased mainly from 45 to 75 days of ripening. This agreed with the concentration of free amino acids. Raw ewes' milk was contaminated by several bacterial phyla: Proteobacteria (68%; mainly Pseudomonas), Firmicutes (30%; mainly Carnobacterium and Lactococcus), Bacteroidetes (0.05%), and Actinobacteria (0.02%). Almost the same microbial composition persisted in the curd after molding. From day 1 of ripening onwards, the phylum Firmicutes dominated. Lactococcus dominated throughout ripening, and most of the Lactobacillus species appeared only at 7 or 15 days. At 90 days, Lactococcus (87.2%), Lactobacillus (4.8%; mainly Lactobacillus plantarum and Lactobacillus sakei), and Leuconostoc (3.9%) dominated. The relative utilization of carbon sources by the bacterial community reflected the succession. This study identified strategic phases that characterized the manufacture and ripening of Canestrato Pugliese cheese and established a causal relationship between mesophilic lactobacilli and proteolysis. PMID:24771032

Causal relationship between microbial ecology dynamics and proteolysis during manufacture and ripening of protected designation of origin (PDO) cheese Canestrato Pugliese.

PubMed

De Pasquale, Ilaria; Calasso, Maria; Mancini, Leonardo; Ercolini, Danilo; La Storia, Antonietta; De Angelis, Maria; Di Cagno, Raffaella; Gobbetti, Marco

2014-07-01

Pyrosequencing of the 16S rRNA gene, community-level physiological profiles determined by the use of Biolog EcoPlates, and proteolysis analyses were used to characterize Canestrato Pugliese Protected Designation of Origin (PDO) cheese. The number of presumptive mesophilic lactococci in raw ewes' milk was higher than that of presumptive mesophilic lactobacilli. The numbers of these microbial groups increased during ripening, showing temporal and numerical differences. Urea-PAGE showed limited primary proteolysis, whereas the analysis of the pH 4.6-soluble fraction of the cheese revealed that secondary proteolysis increased mainly from 45 to 75 days of ripening. This agreed with the concentration of free amino acids. Raw ewes' milk was contaminated by several bacterial phyla: Proteobacteria (68%; mainly Pseudomonas), Firmicutes (30%; mainly Carnobacterium and Lactococcus), Bacteroidetes (0.05%), and Actinobacteria (0.02%). Almost the same microbial composition persisted in the curd after molding. From day 1 of ripening onwards, the phylum Firmicutes dominated. Lactococcus dominated throughout ripening, and most of the Lactobacillus species appeared only at 7 or 15 days. At 90 days, Lactococcus (87.2%), Lactobacillus (4.8%; mainly Lactobacillus plantarum and Lactobacillus sakei), and Leuconostoc (3.9%) dominated. The relative utilization of carbon sources by the bacterial community reflected the succession. This study identified strategic phases that characterized the manufacture and ripening of Canestrato Pugliese cheese and established a causal relationship between mesophilic lactobacilli and proteolysis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Characterization by culture-dependent and culture-independent methods of the bacterial population of suckling-lamb packaged in different atmospheres.

PubMed

Osés, Sandra M; Diez, Ana M; Melero, Beatriz; Luning, Pieternel A; Jaime, Isabel; Rovira, Jordi

2013-12-01

This study offers insight into the dynamics of bacterial populations in fresh cuts of suckling lamb under four different atmospheric conditions: air (A), and three Modified Atmosphere Packaging (MAP) environments, 15%O2/30%CO2/55%N2 (C, commercial), 70%O2/30%CO2 (O), and 15%O2/85%CO2 (H) for 18 days. Microbial analyses by both conventional methods and PCR-DGGE were performed. Controversial and surprising results emerged from comparing both methods in relation to the genus Pseudomonas. Thus, conventional methods detected the presence of high numbers of Pseudomonas colonies, although PCR-DGGE only detected this genus in air-packaged samples. PCR-DGGE detected higher microbial diversity in the control samples (A) than in the modified atmospheres (C, O, H), having atmosphere H the fewest number of species. Brochothrix thermosphacta, LAB (Carnobacterium divergens and Lactobacillus sakei), and Escherichia spp. were detected in all the atmospheres throughout storage. Moreover, previously undescribed bacteria from lamb meat such as Enterobacter hormaechei, Staphylococcus equorum and Jeotgalicoccus spp. were also isolated in this study by DGGE. Additionally, qPCR analysis was used to detect and characterize strains of Escherichia coli. Virulence genes (stx1, stx2 and eae) were detected throughout storage in 97% of the samples. A high CO2 atmosphere was the most effective packaging combination doubling storage time in comparison with commercial atmosphere. Copyright © 2013 Elsevier Ltd. All rights reserved.

Lipoteichoic Acid of Probiotic Lactobacillus plantarum Attenuates Poly I:C-Induced IL-8 Production in Porcine Intestinal Epithelial Cells

PubMed Central

Kim, Kyoung Whun; Kang, Seok-Seong; Woo, Sun-Je; Park, Ok-Jin; Ahn, Ki Bum; Song, Ki-Duk; Lee, Hak-Kyo; Yun, Cheol-Heui; Han, Seung Hyun

2017-01-01

Probiotics in livestock feed supplements are considered a replacement for antibiotics that enhance gastrointestinal immunity. Although bacterial cell wall components have been proposed to be associated with probiotic function, little evidence demonstrates that they are responsible for probiotic functions in livestock. The present study demonstrated that lipoteichoic acid (LTA) of Lactobacillus plantarum (Lp.LTA) confers anti-inflammatory responses in porcine intestinal epithelial cell line, IPEC-J2. A synthetic analog of viral double-stranded RNA, poly I:C, dose-dependently induced IL-8 production at the mRNA and protein levels in IPEC-J2 cells. Lp.LTA, but not lipoprotein or peptidoglycan from L. plantarum, exclusively suppressed poly I:C-induced IL-8 production. Compared with LTAs from other probiotic Lactobacillus strains including L. delbrueckii, L. sakei, and L. rhamnosus GG, Lp.LTA had higher potential to suppress poly I:C-induced IL-8 production. Dealanylated or deacylated Lp.LTA did not suppress poly I:C-induced IL-8 production, suggesting that D-alanine and lipid moieties in the Lp.LTA structure were responsible for the inhibition. Furthermore, Lp.LTA attenuated the phosphorylation of ERK and p38 kinase as well as the activation of NF-κB, resulting in decreased IL-8 production. Taken together, these results suggest that Lp.LTA acts as an effector molecule to inhibit viral pathogen-induced inflammatory responses in porcine intestinal epithelial cells. PMID:28983294

The predominance of Lactobacillus sanfranciscensis in French organic sourdoughs and its impact on related bread characteristics.

PubMed

Lhomme, Emilie; Orain, Servane; Courcoux, Philippe; Onno, Bernard; Dousset, Xavier

2015-11-20

Fourteen bakeries located in different regions of France were selected. These bakers use natural sourdough and organic ingredients. Consequently, different organic sourdoughs used for the manufacture of French bread were studied by the enumeration of lactic acid bacteria (LAB) and 16S rRNA sequencing of the isolates. In addition, after DNA extraction the bacterial diversity was assessed by pyrosequencing of the 16S rDNA V1-V3 region. Although LAB counts showed significant variations (7.6-9.5log10CFU/g) depending on the sourdough studied, their identification through a polyphasic approach revealed a large predominance of Lactobacillus sanfranciscensis in all samples. In ten sourdoughs, both culture and independent methods identified L. sanfranciscensis as the dominant LAB species identified. In the remaining sourdoughs, culture methods identified 30-80% of the LAB as L. sanfranciscensis whereas more than 95% of the reads obtained by pyrosequencing belonged to L. sanfranciscensis. Other sub-dominant species, such as Lactobacillus curvatus, Lactobacillus hammesii, Lactobacillus paralimentarius, Lactobacillus plantarum, Lactobacillus pentosus, and Lactobacillus sakei, were also identified. Quantification of L. sanfranciscensis by real-time PCR confirmed the predominance of this species ranging from 8.24 to 10.38log10CFU/g. Regarding the acidification characteristics, sourdough and related bread physico-chemical characteristics varied, questioning the involvement of sub-dominant species or L. sanfranciscensis intra-species diversity and/or the role of the baker's practices. Copyright © 2015 Elsevier B.V. All rights reserved.

Reduction of acrylamide in whole-wheat bread by combining lactobacilli and yeast fermentation.

PubMed

Nasiri Esfahani, Behnaz; Kadivar, Mahdi; Shahedi, Mohammad; Soleimanian-Zad, Sabihe

2017-11-01

This study mainly focuses on a strategy for reducing acrylamide content in whole-wheat bread by combining lactobacilli and yeast in sourdough breadmaking. Combinations of sourdough (fermented dough using different Lactobacillus strains including Lactobacillus plantarum PTCC 1896 [probiotic], L. sakei DSM 20,017, L. rhamnosus DSM 20,021, and L. delbrueckii DSM 20,081) and yeast, in comparison with yeast alone, were used for breadmaking. The results showed that acrylamide levels in breads fermented using sourdough+yeast were in all cases much lower (6.9-20 μg/kg on a dry weight basis [d.b.]) than those in the yeast-only fermented bread (47.6 μg/kg d.b.). Significant (p < 0.05) correlations were also found between pH, total titratable acids (TTA) and lactic acid, and acrylamide content. Furthermore, the obtained results showed that the moisture content of dough directly influenced the formation of acrylamide in bread (r = 0.925, p < 0.0001). In addition, no significant correlations were observed between acrylamide content in breads and either the reducing sugar or free amino acid contents in dough samples. According to the different effects of Lactobacillus strains, it could be concluded that the acrylamide reducing potential of lactobacilli was strain-specific, with L. rhamnosus being the most effective. This suggests that sourdough fermentation with appropriate Lactobacillus strains can be used as an advantageous technology to reduce the acrylamide content of whole-wheat breads.

Catabolic flexibility of mammalian-associated lactobacilli

PubMed Central

2013-01-01

Metabolic flexibility may be generally defined as “the capacity for the organism to adapt fuel oxidation to fuel availability”. The metabolic diversification strategies used by individual bacteria vary greatly from the use of novel or acquired enzymes to the use of plasmid-localised genes and transporters. In this review, we describe the ability of lactobacilli to utilise a variety of carbon sources from their current or new environments in order to grow and survive. The genus Lactobacillus now includes more than 150 species, many with adaptive capabilities, broad metabolic capacity and species/strain variance. They are therefore, an informative example of a cell factory capable of adapting to new niches with differing nutritional landscapes. Indeed, lactobacilli naturally colonise and grow in a wide variety of environmental niches which include the roots and foliage of plants, silage, various fermented foods and beverages, the human vagina and the mammalian gastrointestinal tract (GIT; including the mouth, stomach, small intestine and large intestine). Here we primarily describe the metabolic flexibility of some lactobacilli isolated from the mammalian gastrointestinal tract, and we also describe some of the food-associated species with a proven ability to adapt to the GIT. As examples this review concentrates on the following species - Lb. plantarum, Lb. acidophilus, Lb. ruminis, Lb. salivarius, Lb. reuteri and Lb. sakei, to highlight the diversity and inter-relationships between the catabolic nature of species within the genus. PMID:23680304

Characterisation of IS153, an IS3-family insertion sequence isolated from Lactobacillus sanfranciscensis and its use for strain differentiation.

PubMed

Ehrmann, M A; Vogel, R E

2001-11-01

An insertion sequence has been identified in the genome of Lactobacillus sanfranciscensis DSM 20451T as segment of 1351 nucleotides containing 37-bp imperfect terminal inverted repeats. The sequence of this element encodes two out of phase, overlapping open reading frames, orfA and orfB, from which three putative proteins are produced. OrfAB is a transframe protein produced by -1 translational frame shifting between orf A and orf B that is presumed to be the transposase. The large orfAB of this element encodes a 342 amino acid protein that displays similarities with transposases encoded by bacterial insertion sequences belonging to the IS3 family. In L. sanfranciscensis type strain DSM 20451T multiple truncated IS elements were identified. Inverse PCR was used to analyze target sites of four of these elements, but except of their highly AT rich character not any sequence specificity was identified so far. Moreover, no flanking direct repeats were identified. Multiple copies of IS153 were detected by hybridization in other strains of L. sanfranciscensis. Resulting hybridization patterns were shown to differentiate between organisms at strain level rather than a probe targeted against the 16S rDNA. With a PCR based approach IS153 or highly similar sequences were detected in L. acidophilus, L. casei, L. malefermentans, L. plantarum, L. hilgardii, L. collinoides L. farciminis L. sakei and L. salivarius, L. reuteri as well as in Enterococcus faecium, Pediococcus acidilactici and P. pentosaceus.

Immunogenicity and protective efficacy of the Mycobacterium avium subsp. paratuberculosis attenuated mutants against challenge in a mouse model

USDA-ARS?s Scientific Manuscript database

Johne’s disease (JD), caused by Mycobacterium avium subsp. paratuberculosis (MAP), results in serious economic losses worldwide especially in cattle, sheep and goats. To control the impact of JD on the animal industry, an effective vaccine with minimal adverse effects is urgently required. In order ...

Islandinium minutum subsp. barbatum subsp. nov. (Dinoflagellata), a New Organic-Walled Dinoflagellate Cyst from the Western Arctic: Morphology, Phylogenetic Position Based on SSU rDNA and LSU rDNA, and Distribution.

PubMed

Potvin, Éric; Kim, So-Young; Yang, Eun Jin; Head, Martin J; Kim, Hyun-Cheol; Nam, Seung-Il; Yim, Joung Han; Kang, Sung-Ho

2018-03-25

A study of modern sediment from the Western Arctic has revealed the presence of a distinctive brown-colored cyst with a spherical central body bearing unbranched processes that are usually solid with a small basal pericoel. Distinctive barbs project from some processes, and process tips are usually minutely expanded into conjoined barbs. The archeopyle is apical and saphopylic. This cyst corresponds to Islandinium? cezare morphotype 2 of Head et al. (2001, J. Quat. Sci., 16:621). Phylogenetic analyses based on the small and large subunit rRNA genes infer close relationship with Islandinium minutum, the type of which is that of the genus. Re-examination of specimens of I. minutum reveals the presence of minute barbs on its processes, but differences with Islandinium? cezare morphotype 2 remain based on size, process distribution, and barb development. Furthermore, the internal transcribed spacer shows I. minutum to be distinct from this morphotype. On the basis of these small but discrete differences, we propose the new subspecies Islandinium minutum subsp. barbatum subsp. nov. Molecular sequencing of other cysts encountered, namely Echinidinium karaense, an unidentified flattened cyst, and "Polykrikos quadratus", places them in the Monovela clade, the latter showing greater morphological variability than previously thought. © 2018 The Author(s) Journal of Eukaryotic Microbiology © 2018 International Society of Protistologists.

Detection of Olea europaea subsp. cuspidata and Juniperus procera in the dry Afromontane forest of northern Ethiopia using subpixel analysis of Landsat imagery

NASA Astrophysics Data System (ADS)

Hishe, Hadgu; Giday, Kidane; Neka, Mulugeta; Soromessa, Teshome; Van Orshoven, Jos; Muys, Bart

2015-01-01

Comprehensive and less costly forest inventory approaches are required to monitor the spatiotemporal dynamics of key species in forest ecosystems. Subpixel analysis using the earth resources data analysis system imagine subpixel classification procedure was tested to extract Olea europaea subsp. cuspidata and Juniperus procera canopies from Landsat 7 enhanced thematic mapper plus imagery. Control points with various canopy area fractions of the target species were collected to develop signatures for each of the species. With these signatures, the imagine subpixel classification procedure was run for each species independently. The subpixel process enabled the detection of O. europaea subsp. cuspidata and J. procera trees in pure and mixed pixels. Total of 100 pixels each were field verified for both species. An overall accuracy of 85% was achieved for O. europaea subsp. cuspidata and 89% for J. procera. A high overall accuracy level of detecting species at a natural forest was achieved, which encourages using the algorithm for future species monitoring activities. We recommend that the algorithm has to be validated in similar environment to enrich the knowledge on its capability to ensure its wider usage.

Novel Phytases from Bifidobacterium pseudocatenulatum ATCC 27919 and Bifidobacterium longum subsp. infantis ATCC 15697

PubMed Central

Tamayo-Ramos, Juan Antonio; Sanz-Penella, Juan Mario; Yebra, María J.

2012-01-01

Two novel phytases have been characterized from Bifidobacterium pseudocatenulatum and Bifidobacterium longum subsp. infantis. The enzymes belong to a new subclass within the histidine acid phytases, are highly specific for the hydrolysis of phytate, and render myo-inositol triphosphate as the final hydrolysis product. They represent the first phytases characterized from this group of probiotic microorganisms, opening the possibilities for their use in the processing of high-phytate-content foods. PMID:22582052

Molecular Characterization of Three Lactobacillus delbrueckii subsp. bulgaricus Phages

PubMed Central

Casey, Eoghan; Mahony, Jennifer; O'Connell-Motherway, Mary; Bottacini, Francesca; Cornelissen, Anneleen; Neve, Horst; Heller, Knut J.; Noben, Jean-Paul; Dal Bello, Fabio

2014-01-01

In this study, three phages infecting Lactobacillus delbrueckii subsp. bulgaricus, named Ld3, Ld17, and Ld25A, were isolated from whey samples obtained from various industrial fermentations. These phages were further characterized in a multifaceted approach: (i) biological and physical characterization through host range analysis and electron microscopy; (ii) genetic assessment through genome analysis; (iii) mass spectrometry analysis of the structural components of the phages; and (iv), for one phage, transcriptional analysis by Northern hybridization, reverse transcription-PCR, and primer extension. The three obtained phage genomes display high levels of sequence identity to each other and to genomes of the so-called group b L. delbrueckii phages c5, LL-Ku, and phiLdb, where some of the observed differences are believed to be responsible for host range variations. PMID:25002431

Characterization of nitrite degradation by Lactobacillus casei subsp. rhamnosus LCR 6013.

PubMed

Liu, Dong-mei; Wang, Pan; Zhang, Xin-yue; Xu, Xi-lin; Wu, Hui; Li, Li

2014-01-01

Nitrites are potential carcinogens. Therefore, limiting nitrites in food is critically important for food safety. The nitrite degradation capacity of Lactobacillus casei subsp. rhamnosus LCR 6013 was investigated in pickle fermentation. After LCR 6013 fermentation for 120 h at 37°C, the nitrite concentration in the fermentation system was significantly lower than that in the control sample without the LCR 6013 strain. The effects of NaCl and Vc on nitrite degradation by LCR 6013 in the De Man, Rogosa and Sharpe (MRS) medium were also investigated. The highest nitrite degradations, 9.29 mg/L and 9.89 mg/L, were observed when NaCl and Vc concentrations were 0.75% and 0.02%, respectively in the MRS medium, which was significantly higher than the control group (p ≤ 0.01). Electron capture/gas chromatography and indophenol blue staining were used to study the nitrite degradation pathway of LCR 6013. The nitrite degradation products contained N2O, but no NH4(+). The LCR 6013 strain completely degraded all NaNO2 (50.00 mg/L) after 16 h of fermentation. The enzyme activity of NiR in the periplasmic space was 2.5 times of that in the cytoplasm. Our results demonstrated that L. casei subsp. rhamnosus LCR 6013 can effectively degrade nitrites in both the pickle fermentation system and in MRS medium by NiR. Nitrites are degraded by the LCR 6013 strain, likely via the nitrate respiration pathway (NO2(-)>NO->N2O->N2), rather than the aammonium formation pathway (dissimilatory nitrate reduction to ammonium, DNRA), because the degradation products contain N2O, but not NH4(+).

Draft Genome Sequences of 18 Salmonella enterica subsp. enterica Serovar Oranienburg Strains Isolated from Rivers in Northwestern Mexico.

PubMed

Casteñeda-Ruelas, Gloria M; Carreón-Gaxiola, César; Castelán-Sánchez, Hugo G; Acatzi-Silva, Abraham; Romero-Martínez, Salvador; García-Molina, Alejandra; Jiménez-Edeza, Maribel

2017-03-09

Salmonella enterica subsp. enterica serovar Oranienburg is recognized as a foodborne pathogen widely distributed in the environment. Here, we report 18 draft genomes of S  Oranienburg strains isolated from rivers in the northwestern region of Mexico. Copyright © 2017 Casteñeda-Ruelas et al.

Genome Sequence of Streptococcus phocae subsp. salmonis Strain C-4T, Isolated from Atlantic Salmon (Salmo salar)

PubMed Central

Suarez, Rudy; Lazo, Eduardo; Bravo, Diego; Llegues, Katerina O.; Romalde, Jesús L.; Godoy, Marcos G.

2014-01-01

Streptococcus phocae subsp. salmonis is a fish pathogen that has an important impact on the Chilean salmon industry. Here, we report the genome sequence of the type strain C-4T isolated from Atlantic salmon (Salmo salar), showing a number of interesting features and genes related to its possible virulence factors. PMID:25502668

Complete genome sequence of Clavibacter michiganensis subsp. insidiosus R1-1 using PacBio single-molecule real-time technology

USDA-ARS?s Scientific Manuscript database

We report the complete genome sequence of Clavibacter michiganensis subsp. insidiosus R1-1 isolated in Minnesota, USA. The R1-1 genome, generated by de novo assembly of PacBio sequencing data, is the first complete genome sequence available for this subspecies....

Mediation of host immune responses after immunization of neonatal calves with a heat-killed Mycobacterium avium subsp. paratuberculosis vaccine

USDA-ARS?s Scientific Manuscript database

A major drawback of current whole-cell vaccines for Mycobacterium avium subsp. paratuberculosis(MAP) is the interference with diagnostic tests for bovine tuberculosis and paratuberculosis. The current study was designed to explore effects of immunization with a heat-killed whole cell vaccine (Mycop...

Evaluation of Control Points in Youngstock and Adult Dairy Cow Management to Control Transmission of Mycobacterium avium subsp. paratuberculosis

USDA-ARS?s Scientific Manuscript database

Goal Complete a series of controlled on-farm trials to critically evaluate the efficacy and cost-benefit of commonly recommended management practices for reducing the transmission of Mycobacterium avium subsp. paratuberculosis (Map) in infected herds. Objective 1. Evaluate the effect of maternity...

Genome Sequence of Lactobacillus delbrueckii subsp. lactis CNRZ327, a Dairy Bacterium with Anti-Inflammatory Properties.

PubMed

El Kafsi, Hela; Binesse, Johan; Loux, Valentin; Buratti, Julien; Boudebbouze, Samira; Dervyn, Rozenn; Hammani, Amal; Maguin, Emmanuelle; van de Guchte, Maarten

2014-07-17

Lactobacillus delbrueckii subsp. lactis CNRZ327 is a dairy bacterium with anti-inflammatory properties both in vitro and in vivo. Here, we report the genome sequence of this bacterium, which appears to contain no less than 215 insertion sequence (IS) elements, an exceptionally high number regarding the small genome size of the strain. Copyright © 2014 El Kafsi et al.

Genetic diversity of Streptococcus equi subsp. zooepidemicus and doxycycline resistance in kennelled dogs.

PubMed

Chalker, Victoria J; Waller, Andrew; Webb, Katy; Spearing, Emma; Crosse, Patricia; Brownlie, Joe; Erles, Kerstin

2012-06-01

The genetic diversity and antibiotic resistance profiles of 38 Streptococcus equi subsp. zooepidemicus isolates were determined from a kennelled canine population during two outbreaks of hemorrhagic pneumonia (1999 to 2002 and 2007 to 2010). Analysis of the szp gene hypervariable region and the 16S-23S rRNA intergenic spacer region and multilocus sequence typing (MLST) indicated a predominant tetO-positive, doxycycline-resistant ST-10 strain during 1999 to 2002 and a predominant tetM-positive doxycycline-resistant ST-62 strain during 2007 to 2010.

Streptococcus equi subsp zooepidemicus pleuropneumonia and peritonitis in a dromedary camel (Camelus dromedarius) calf in North America.

PubMed

Stoughton, William B; Gold, Jenifer

2015-08-01

A 12-week-old female dromedary camel (Camelus dromedarius) calf was evaluated because of acute (< 24 hours) inappetence and lethargy. The calf was being bottle-fed because of maternal rejection. Physical examination revealed decreased bronchovesicular sounds and absent borborygmi. The rectal temperature was 38.9°C (102.0°F). A CBC indicated leukopenia with a degenerative left shift suggestive of a systemic infection. Results of abdominal and thoracic ultrasonography showed severe bicavitary effusion, peripheral lung consolidation, and intestinal hypomotility. Pleural and peritoneal fluid analysis confirmed a diagnosis of septic pleuritis and peritonitis. Results of aerobic bacterial culture of venous blood, peritoneal fluid, and pleural fluid samples indicated Streptococcus equi subsp zooepidemicus septicemia as the etiology for the polyserositis (ie, alpaca fever). Treatment with IV broad-spectrum antimicrobials, an NSAID, and pleural drainage was initiated. Clinical signs of pleuropneumonia, peritonitis, and systemic infection improved rapidly 24 hours after initiation of medical treatment. The calf was discharged from the hospital after 11 days, and antimicrobial treatment continued for 2 weeks after discharge. At follow-up approximately 4 weeks after hospital discharge (6 weeks after the initial examination), there were no clinical signs suggestive of relapse or any reported complications. S equi subsp zooepidemicus may cause polyserositis in Old World camelids (eg, dromedary camels) with signs similar to those seen in New World camelids (eg, alpaca and llama). The rapid response to medical treatment for the patient described suggested that S equi subsp zooepidemicus-induced polyserositis (alpaca fever) in dromedary camels may respond favorably to appropriate treatment. Reducing stress, reducing overcrowding, and separate housing of equids and camelids are suggested. Further studies are needed to better assess the epidemiology of alpaca fever in dromedary

The cellodextrinase from Pseudomonas fluorescens subsp. cellulosa consists of multiple functional domains.

PubMed Central

Ferreira, L M; Hazlewood, G P; Barker, P J; Gilbert, H J

1991-01-01

A genomic library of Pseudomonas fluorescens subsp. cellulosa DNA was constructed in pUC18 and Escherichia coli recombinants expressing 4-methylumbelliferyl beta-D-cellobioside-hydrolysing activity (MUCase) were isolated. Enzyme produced by MUCase-positive clones did not hydrolyse either cellobiose or cellotriose but converted cellotetraose into cellobiose and cleaved cellopentaose and cellohexaose, producing a mixture of cellobiose and cellotriose. There was no activity against CM-cellulose, insoluble cellulose or xylan. On this basis, the enzyme is identified as an endo-acting cellodextrinase and is designated cellodextrinase C (CELC). Nucleotide sequencing of the gene (celC) which directs the synthesis of CELC revealed an open reading frame of 2153 bp, encoding a protein of Mr 80,189. The deduced primary sequence of CELC was confirmed by the Mr of purified CELC (77,000) and by the experimentally determined N-terminus of the enzyme which was identical with residues 38-47 of the translated sequence. The N-terminal region of CELC showed strong homology with endoglucanase, xylanases and an arabinofuranosidase of Ps. fluorescens subsp. cellulosa; homologous sequences included highly conserved serine-rich regions. Full-length CELC bound tightly to crystalline cellulose. Truncated forms of celC from which the DNA sequence encoding the conserved domain had been deleted, directed the synthesis of a functional cellodextrinase that did not bind to crystalline cellulose. This is consistent with the N-terminal region of CELC comprising a non-catalytic cellulose-binding domain which is distinct from the catalytic domain. The role of the cellulose-binding region is discussed. Images Fig. 2. Fig. 6. PMID:1953673

Cyto-adherence of Mycoplasma mycoides subsp. mycoides to bovine lung epithelial cells.

PubMed

Aye, Racheal; Mwirigi, Martin Kiogora; Frey, Joachim; Pilo, Paola; Jores, Joerg; Naessens, Jan

2015-02-07

Mycoplasma mycoides subsp. mycoides (Mmm) is the causative agent of contagious bovine pleuropneumonia (CBPP), a respiratory disease of cattle, whereas the closely related Mycoplasma mycoides subsp. capri (Mmc) is a goat pathogen. Cyto-adherence is a crucial step in host colonization by mycoplasmas and subsequent pathogenesis. The aim of this study was to investigate the interactions between Mmm and mammalian host cells by establishing a cyto-adherence flow cytometric assay and comparing tissue and species specificity of Mmm and Mmc strains. There were little significant differences in the adherence patterns of eight different Mmm strains to adult bovine lung epithelial cells. However, there was statistically significant variation in binding to different host cells types. Highest binding was observed with lung epithelial cells, intermediate binding with endothelial cells and very low binding with fibroblasts, suggesting the presence of effective adherence of Mmm on cells lining the airways of the lung, which is the target organ for this pathogen, possibly by high expression of a specific receptor. However, binding to bovine fetal lung epithelial cells was comparably low; suggesting that the lack of severe pulmonary disease seen in many infected young calves can be explained by reduced expression of a specific receptor. Mmm bound with high efficiency to adult bovine lung cells and less efficiently to calves or goat lung cells. The data show that cyto-adherence of Mmm is species- and tissue- specific confirming its role in colonization of the target host and subsequent infection and development of CBPP.

Hypericum perforatum L. subsp. perforatum induces inhibition of free radicals and enhanced phototoxicity in human melanoma cells under ultraviolet light.

PubMed

Menichini, G; Alfano, C; Marrelli, M; Toniolo, C; Provenzano, E; Statti, G A; Nicoletti, M; Menichini, F; Conforti, F

2013-04-01

Our interest continues in discovering phytocomplexes from medicinal plants with phototoxic activity against human melanoma cells; thus the aim of the present study was to assess antioxidant, anti-inflammatory and phototoxic activity of Hypericum perforatum L. subsp. perforatum, and relate these properties to the plant's chemical composition. Components of H. perforatum subsp. perforatum were extracted by hydroalcoholic solution and chemical profiles of preparations (HyTE-3) performed by HPTLC. Linoleic acid peroxidation and DPPH tests were used to assess antioxidant activity, while MTT assay allowed evaluation of anti-proliferative activity with respect to A375 human melanoma cells after irradiation with UVA dose, 1.8 J/cm(2) . Inhibition of nitric oxide production of macrophages was also investigated. HyTE-3 indicated better antioxidant activity with β-carotene bleaching test in comparison to DPPH assay (IC50 = 0.89 μg/ml); significant phototoxicity in A375 cells at 78 μg/ml concentration resulted in cell destruction of 50%. HyTE-3 caused significant dose-related inhibition of nitric oxide production in murine monocytic macrophage cell line RAW 264.7 with IC50 value of 342 μg/ml. The H. perforatum subsp. perforatum-derived product was able to suppress proliferation of human malignant melanoma A375 cells; extract together with UVA irradiation enhanced phototoxicity. This biological activity of antioxidant effects was combined with inhibition of nitric oxide production. © 2013 Blackwell Publishing Ltd.

Clavibacter michiganensis subsp. michiganensis: first steps in the understanding of virulence of a Gram-positive phytopathogenic bacterium.

PubMed

Gartemann, Karl-Heinz; Kirchner, Oliver; Engemann, Jutta; Gräfen, Ines; Eichenlaub, Rudolf; Burger, Annette

2003-12-19

Clavibacter michiganensis subsp. michiganensis is a plant-pathogenic actinomycete. It infects tomato, spreads through the xylem and causes bacterial wilt and canker. The wild-type strain NCPPB382 carries two plasmids, pCM1 and pCM2. The cured plasmid-free derivative CMM100 is still able to colonize tomato, but no disease symptoms develop indicating that all genes required for successful infection, establishment and growth in the plant reside on the chromosome. Both plasmids carry one virulence factor, a gene encoding a cellulase, CelA in case of pCM1 and a putative serine protease Pat-1 on pCM2. These genes can independently convert the non-virulent strain CMM100 into a pathogen causing wilt on tomatoes. Currently, genome projects for Cmm and the closely related potato-pathogen C. michiganensis subsp. sepedonicus have been initiated. The data from the genome project shall give clues on further genes involved in plant-microbe interaction that can be tested experimentally. Especially, identification of genes related to host-specificity through genome comparison of the two subspecies might be possible.

Effect of growth conditions on production of rhamnose-containing cell wall and capsular polysaccharides by strains of Lactobacillus casei subsp. rhamnosus.

PubMed

Wicken, A J; Ayres, A; Campbell, L K; Knox, K W

1983-01-01

Strains of Lactobacillus casei subsp. rhamnosus possessing two cell wall polysaccharides, a hexosamine-containing H-polysaccharide and a rhamnose-containing R-polysaccharide, were examined for the effect of growth conditions on the production of these two components. In strain NCTC 6375, R- and H-polysaccharides accounted for an estimated 44 and 20%, respectively, of the cell wall for organisms grown in batch culture with glucose as the carbohydrate source. Growth on fructose-containing media reduced the amount of R-polysaccharide by approximately 50% without affecting the amount of H-polysaccharide. Subculture of fructose-grown organisms in glucose restored the original proportions of the two polysaccharides. Galactose- and sucrose-grown cells behaved similarly to glucose-grown cells with respect to polysaccharide production, whereas growth in rhamnose or ribose showed values close to those for fructose-grown cells. Continuous culture of strain NCTC 6375 for more than 100 generations showed a gradual and irreversible reduction of the R-polysaccharide to less than 5% of the cell wall and an increase of the H-polysaccharide to 40% of the cell wall. Other type culture strains of L. casei subsp. rhamnosus, NCIB 7473 and ATCC 7469, behaved similarly in batch and continuous culture. In contrast, strains of L. casei subsp. rhamnosus isolated at the Institute of Dental Research showed phenotypic stability with respect to the relative proportions of R- and H-polysaccharides in both batch and continuous culture. Changes in polysaccharide composition of type culture strains were also mirrored in changes in the immunogenicity of the two components and resistance to the rate of enzymic lysis of whole organisms. For L. casei subsp. rhamnosus strain NCTC 10302 the R-polysaccharide is present entirely as capsular material. The amount of R-polysaccharide produced was also markedly dependent on the carbohydrate component of the medium in batch culture and both dilution rate and

Effect of growth conditions on production of rhamnose-containing cell wall and capsular polysaccharides by strains of Lactobacillus casei subsp. rhamnosus.

PubMed Central

Wicken, A J; Ayres, A; Campbell, L K; Knox, K W

1983-01-01

Strains of Lactobacillus casei subsp. rhamnosus possessing two cell wall polysaccharides, a hexosamine-containing H-polysaccharide and a rhamnose-containing R-polysaccharide, were examined for the effect of growth conditions on the production of these two components. In strain NCTC 6375, R- and H-polysaccharides accounted for an estimated 44 and 20%, respectively, of the cell wall for organisms grown in batch culture with glucose as the carbohydrate source. Growth on fructose-containing media reduced the amount of R-polysaccharide by approximately 50% without affecting the amount of H-polysaccharide. Subculture of fructose-grown organisms in glucose restored the original proportions of the two polysaccharides. Galactose- and sucrose-grown cells behaved similarly to glucose-grown cells with respect to polysaccharide production, whereas growth in rhamnose or ribose showed values close to those for fructose-grown cells. Continuous culture of strain NCTC 6375 for more than 100 generations showed a gradual and irreversible reduction of the R-polysaccharide to less than 5% of the cell wall and an increase of the H-polysaccharide to 40% of the cell wall. Other type culture strains of L. casei subsp. rhamnosus, NCIB 7473 and ATCC 7469, behaved similarly in batch and continuous culture. In contrast, strains of L. casei subsp. rhamnosus isolated at the Institute of Dental Research showed phenotypic stability with respect to the relative proportions of R- and H-polysaccharides in both batch and continuous culture. Changes in polysaccharide composition of type culture strains were also mirrored in changes in the immunogenicity of the two components and resistance to the rate of enzymic lysis of whole organisms. For L. casei subsp. rhamnosus strain NCTC 10302 the R-polysaccharide is present entirely as capsular material. The amount of R-polysaccharide produced was also markedly dependent on the carbohydrate component of the medium in batch culture and both dilution rate and

Contribution of type IV pili to the virulence of Aeromonas salmonicida subsp. salmonicida in Atlantic salmon (Salmo salar L.).

PubMed

Boyd, Jessica M; Dacanay, Andrew; Knickle, Leah C; Touhami, Ahmed; Brown, Laura L; Jericho, Manfred H; Johnson, Stewart C; Reith, Michael

2008-04-01

Aeromonas salmonicida subsp. salmonicida, a bacterial pathogen of Atlantic salmon, has no visible pili, yet its genome contains genes for three type IV pilus systems. One system, Tap, is similar to the Pseudomonas aeruginosa Pil system, and a second, Flp, resembles the Actinobacillus actinomycetemcomitans Flp pilus, while the third has homology to the mannose-sensitive hemagglutinin pilus of Vibrio cholerae. The latter system is likely nonfunctional since eight genes, including the gene encoding the main pilin subunit, are deleted compared with the orthologous V. cholerae locus. The first two systems were characterized to investigate their expression and role in pathogenesis. The pili of A. salmonicida subsp. salmonicida were imaged using atomic force microscopy and Tap- and Flp-overexpressing strains. The Tap pili appeared to be polar, while the Flp pili appeared to be peritrichous. Strains deficient in tap and/or flp were used in live bacterial challenges of Atlantic salmon, which showed that the Tap pilus made a moderate contribution to virulence, while the Flp pilus made little or no contribution. Delivery of the tap mutant by immersion resulted in reduced cumulative morbidity compared with the cumulative morbidity observed with the wild-type strain; however, delivery by intraperitoneal injection resulted in cumulative morbidity similar to that of the wild type. Unlike the pili of other piliated bacterial pathogens, A. salmonicida subsp. salmonicida type IV pili are not absolutely required for virulence in Atlantic salmon. Significant differences in the behavior of the two mutant strains indicated that the two pilus systems are not redundant.

Antimicrobial phenolics and unusual glycerides from Helichrysum italicum subsp. microphyllum.

PubMed

Taglialatela-Scafati, Orazio; Pollastro, Federica; Chianese, Giuseppina; Minassi, Alberto; Gibbons, Simon; Arunotayanun, Warunya; Mabebie, Blessing; Ballero, Mauro; Appendino, Giovanni

2013-03-22

During a large-scale isolation campaign for the heterodimeric phloroglucinyl pyrone arzanol (1a) from Helichrysum italicum subsp. microphyllum, several new phenolics as well as an unusual class of lipids named santinols (5a-c, 6-8) have been characterized. Santinols are angeloylated glycerides characterized by the presence of branched acyl- or keto-acyl chains and represent a hitherto unreported class of plant lipids. The antibacterial activity of arzanol and of a selection of Helichrysum phenolics that includes coumarates, benzofurans, pyrones, and heterodimeric phloroglucinols was evaluated, showing that only the heterodimers showed potent antibacterial action against multidrug-resistant Staphylococcus aureus isolates. These observations validate the topical use of Helichrysum extracts to prevent wound infections, a practice firmly established in the traditional medicine of the Mediterranean area.

Genotypic and phenotypic changes in wild barley (Hordeum vulgare subsp. spontaneum) during a period of climate change in Jordan

USDA-ARS?s Scientific Manuscript database

Climate change and other anthropogenic disturbances can lead to the loss of genetic variation and thereby affect evolutionary potential and survival of plant populations in the wild. We examined these predictions in the primary wild relative of barley, Hordeum vulgare L. subsp. spontaneum (K. Koch) ...

Occurrence of transgenic feral alfalfa (Medicago sativa subsp. sativa L.) in alfalfa seed production areas in the United States

USDA-ARS?s Scientific Manuscript database

Genetically-engineered glyphosate-resistant alfalfa (Medicago sativa subsp. sativa) was commercialized in 2011. The potential risk of transgene dispersal into the environment is not clearly understood for alfalfa, a perennial crop that is cross-pollinated by insects. We gathered data on feral and tr...

Erwinia carotovora subsp. carotovora extracellular protease: characterization and nucleotide sequence of the gene.

PubMed Central

Kyöstiö, S R; Cramer, C L; Lacy, G H

1991-01-01

The prt1 gene encoding extracellular protease from Erwinia carotovora subsp. carotovora EC14 in cosmid pCA7 was subcloned to create plasmid pSK1. The partial nucleotide sequence of the insert in pSK1 (1,878 bp) revealed a 1,041-bp open reading frame (ORF1) that correlated with protease activity in deletion mutants. ORF1 encodes a polypeptide of 347 amino acids with a calculated molecular mass of 38,826 Da. Escherichia coli transformed with pSK1 or pSK23, a subclone of pSK1, produces a protease (Prt1) intracellularly with a molecular mass of 38 kDa and a pI of 4.8. Prt1 activity was inhibited by phenanthroline, suggesting that it is a metalloprotease. The prt1 promoter was localized between 173 and 1,173 bp upstream of ORF1 by constructing transcriptional lacZ fusions. Primer extension identified the prt1 transcription start site 205 bp upstream of ORF1. The deduced amino acid sequence of ORF1 showed significant sequence identity to metalloproteases from Bacillus thermoproteolyticus (thermolysin), B. subtilis (neutral protease), Legionella pneumophila (metalloprotease), and Pseudomonas aeruginosa (elastase). It has less sequence similarity to metalloproteases from Serratia marcescens and Erwinia chrysanthemi. Locations for three zinc ligands and the active site for E. carotovora subsp. carotovora protease were predicted from thermolysin. Images FIG. 2 FIG. 5 FIG. 6 FIG. 8 FIG. 9 PMID:1917878

Accumulation of petroleum hydrocarbons in intracellular lipid bodies of the freshwater diatom Synedra acus subsp. radians.

PubMed

Shishlyannikov, Sergey M; Nikonova, Alyona A; Klimenkov, Igor V; Gorshkov, Alexander G

2017-01-01

The accumulation of hydrophobic compounds by phytoplankton plays a crucial role in the biogeochemical cycle of persistent organic pollutants (POPs) in aquatic environments. We studied the accumulation of polycyclic aromatic hydrocarbons (PAHs) in the freshwater diatom Synedra acus subsp. radians during its cultivation with crude oil hydrocarbons, using epifluorescent and laser confocal microscopy as well as gas chromatography-mass spectrometry (GC/MS) analysis. Our results revealed that in the presence of crude oil or an extract of a crude oil/n-hexane solution (light oil), S. acus subsp. radians accumulated PAHs in its lipid bodies. During cultivation in the presence of a crude oil/n-hexane solution, the cells selectively accumulated C12-C18 alkanes, with a preference for C15 and C16 homologues. The length of n-alkane hydrocarbon chains accumulated in cells was similar to the acyl chains of fatty acids of the diatom. We therefore suggest that the insertion of n-alkanes into the membrane lipid bilayer promotes the transmembrane transport of PAH in diatoms. Our results confirm the hypothesis that diatoms play a role in the elimination of hydrophobic hydrocarbons from aquatic systems.

Molecular characterization of three Lactobacillus delbrueckii subsp. bulgaricus phages.

PubMed

Casey, Eoghan; Mahony, Jennifer; O'Connell-Motherway, Mary; Bottacini, Francesca; Cornelissen, Anneleen; Neve, Horst; Heller, Knut J; Noben, Jean-Paul; Dal Bello, Fabio; van Sinderen, Douwe

2014-09-01

In this study, three phages infecting Lactobacillus delbrueckii subsp. bulgaricus, named Ld3, Ld17, and Ld25A, were isolated from whey samples obtained from various industrial fermentations. These phages were further characterized in a multifaceted approach: (i) biological and physical characterization through host range analysis and electron microscopy; (ii) genetic assessment through genome analysis; (iii) mass spectrometry analysis of the structural components of the phages; and (iv), for one phage, transcriptional analysis by Northern hybridization, reverse transcription-PCR, and primer extension. The three obtained phage genomes display high levels of sequence identity to each other and to genomes of the so-called group b L. delbrueckii phages c5, LL-Ku, and phiLdb, where some of the observed differences are believed to be responsible for host range variations. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Characterization of the mature cell surface proteinase of Lactobacillus delbrueckii subsp. lactis CRL 581.

PubMed

Villegas, Josefina M; Brown, Lucía; Savoy de Giori, Graciela; Hebert, Elvira M

2015-05-01

The cell envelope-associated proteinase (CEP) of Lactobacillus delbrueckii subsp. lactis CRL 581 (PrtL) has an essential role in bacterial growth, contributes to the flavor and texture development of fermented products, and can release bioactive health-beneficial peptides during milk fermentation. The genome of L. delbrueckii subsp. lactis CRL 581 possesses only one gene that encodes PrtL, which consists of 1924 amino acids and is a multidomain protein anchored to the cell via its W domain. PrtL was extracted from the cell under high ionic strength conditions using NaCl, suggesting an electrostatic interaction between the proteinase and the cell envelope. The released PrtL was purified and biochemically characterized; its activity was maximal at temperatures between 37 and 40 °C and at pH between 7 and 8. Under optimal conditions, PrtL exhibited higher affinity for succinyl-alanyl-alanyl-prolyl-phenylalanine-p-nitroanilide than for succinyl-alanyl-glutamyl-prolyl-phenylalanine-p-nitroanilide, while methoxy-succinyl-arginyl-prolyl-tyrosyl-p-nitroanilide was not degraded. A similar α- and β-casein degradation pattern was observed with the purified and the cell envelope-bound proteinase. Finally, on the basis of its specificity towards caseins and the unique combination of amino acids at residues thought to be involved in substrate specificity, PrtL can be classified as a representative of a new group of CEP.

Lactobacillus delbrueckii subsp. lactis (strain CIDCA 133) stimulates murine macrophages infected with Citrobacter rodentium.

PubMed

Hugo, Ayelén A; Rolny, Ivanna S; Romanin, David; Pérez, Pablo F

2017-03-01

Citrobacter rodentium is a specific murine enteropathogen which causes diarrheal disease characterized by colonic hyperplasia and intestinal inflammation. Recruitment of neutrophils and macrophages constitute a key step to control the infection. Since modulation of the activity of professional phagocytic cells could contribute to improve host´s defences against C. rodentium, we investigated the effect of Lactobacillus delbrueckii subsp. lactis (strain CIDCA 133) on the interaction between murine macrophages (RAW 264.7) and C. rodentium. Phagocytosis, surface molecules and inducible nitric oxide synthase (iNOs) expression were determined by flow cytometry. Reactive oxygen species (ROS) were assessed by fluorescence microscopy. The presence of lactobacilli increased phagocytosis of C. rodentium whereas C. rodentium had no effect on lactobacilli internalization. Survival of internalized C. rodentium diminished when strain CIDCA 133 was present. CD-86, MHCII, iNOs expression and nitrite production were increased when C. rodentium and lactobacilli were present even though strain CIDCA 133 alone had no effect. Strain CIDCA 133 led to a strong induction of ROS activity which was not modified by C. rodentium. Lactobacillus delbrueckii subsp. lactis (strain CIDCA 133) is able to increase the activation of murine macrophages infected with C. rodentium. The sole presence of lactobacilli is enough to modify some stimulation markers (e.g. ROS induction) whereas other markers require the presence of both bacteria; thus, indicating a synergistic effect.

Multilocus Sex Determination Revealed in Two Populations of Gynodioecious Wild Strawberry, Fragaria vesca subsp. bracteata

PubMed Central

Ashman, Tia-Lynn; Tennessen, Jacob A.; Dalton, Rebecca M.; Govindarajulu, Rajanikanth; Koski, Matthew H.; Liston, Aaron

2015-01-01

Gynodioecy, the coexistence of females and hermaphrodites, occurs in 20% of angiosperm families and often enables transitions between hermaphroditism and dioecy. Clarifying mechanisms of sex determination in gynodioecious species can thus illuminate sexual system evolution. Genetic determination of gynodioecy, however, can be complex and is not fully characterized in any wild species. We used targeted sequence capture to genetically map a novel nuclear contributor to male sterility in a self-pollinated hermaphrodite of Fragaria vesca subsp. bracteata from the southern portion of its range. To understand its interaction with another identified locus and possibly additional loci, we performed crosses within and between two populations separated by 2000 km, phenotyped the progeny and sequenced candidate markers at both sex-determining loci. The newly mapped locus contains a high density of pentatricopeptide repeat genes, a class commonly involved in restoration of fertility caused by cytoplasmic male sterility. Examination of all crosses revealed three unlinked epistatically interacting loci that determine sexual phenotype and vary in frequency between populations. Fragaria vesca subsp. bracteata represents the first wild gynodioecious species with genomic evidence of both cytoplasmic and nuclear genes in sex determination. We propose a model for the interactions between these loci and new hypotheses for the evolution of sex determining chromosomes in the subdioecious and dioecious Fragaria. PMID:26483011

Application of Raman spectroscopy for direct analysis of Carlina acanthifolia subsp. utzka root essential oil.

PubMed

Strzemski, Maciej; Wójciak-Kosior, Magdalena; Sowa, Ireneusz; Agacka-Mołdoch, Monika; Drączkowski, Piotr; Matosiuk, Dariusz; Kurach, Łukasz; Kocjan, Ryszard; Dresler, Sławomir

2017-11-01

Carlina genus plants e.g. Carlina acanthifolia subsp. utzka have been still used in folk medicine of many European countries and its biological activity is mostly associated with root essential oils. In the present paper, Raman spectroscopy (RS) was applied for the first time for evaluation of essential oil distribution in root of C. acnthifolia subsp. utzka and identification of root structures containing the essential oil. Furthermore, RS technique was applied to assess chemical stability of oil during drying of plant material or distillation process. Gas chromatography-mass spectrometry was used for qualitative and quantitative analysis of the essential oil. The identity of compounds was confirmed using Raman, ATR-IR and NMR spectroscopy. Carlina oxide was found to be the main component of the oil (98.96% ± 0.15). The spectroscopic study showed the high stability of essential oil and Raman distribution analysis indicated that the oil reservoirs were localized mostly in the structures of outer layer of the root while the inner part showed nearly no signal assigned to the oil. Raman spectroscopy technique enabled rapid, non-destructive direct analysis of plant material with minimal sample preparation and allowed straightforward, unambiguous identification of the essential oil in the sample. Copyright © 2017. Published by Elsevier B.V.

Optimization of Exopolysaccharide Production by Lactobacillus delbrueckii subsp. bulgaricus RR Grown in a Semidefined Medium

PubMed Central

Kimmel, Stacy A.; Roberts, Robert F.; Ziegler, Gregory R.

1998-01-01

The optimal fermentation temperature, pH, and Bacto-casitone (Difco Laboratories, Detroit, Mich.) concentration for production of exopolysaccharide by Lactobacillus delbrueckii subsp. bulgaricus RR in a semidefined medium were determined by using response surface methods. The design consisted of 20 experiments, 15 unique combinations, and five replications. All fermentations were conducted in a fermentor with a 2.5-liter working volume and were terminated when 90% of the glucose in the medium had been consumed. The population of L. delbrueckii subsp. bulgaricus RR and exopolysaccharide content were measured at the end of each fermentation. The optimum temperature, pH, and Bacto-casitone concentration for exopolysaccharide production were 38°C, 5, and 30 g/liter, respectively, with a predicted yield of 295 mg of exopolysaccharide/liter. The actual yield under these conditions was 354 mg of exopolysaccharide/liter, which was within the 95% confidence interval (217 to 374 mg of exopolysaccharide/liter). An additional experiment conducted under optimum conditions showed that exopolysaccharide production was growth associated, with a specific production at the endpoint of 101.4 mg/g of dry cells. Finally, to obtain material for further characterization, a 100-liter fermentation was conducted under optimum conditions. Twenty-nine grams of exopolysaccharide was isolated from centrifuged, ultrafiltered fermentation broth by ethanol precipitation. PMID:9464404

Optimization of methods for the detection of Mycobacterium avium subsp. paratuberculosis in milk and colostrum of naturally infected dairy cows

USDA-ARS?s Scientific Manuscript database

Two decontamination chemicals, hexadecylpyridinium choride (HPC) and N-acetyl-L-cysteine-sodium hydroxide (NALC-NaOH), were compared for their efficacy of reducing the growth of non-specific microorganisms in milk while minimally affecting the recovery of Mycobacterium avium subsp. paratuberculosis ...

DsbA Plays a Critical and Multifaceted Role in the Production of Secreted Virulence Factors by the Phytopathogen Erwinia carotovora subsp. atroseptica*S⃞

PubMed Central

Coulthurst, Sarah J.; Lilley, Kathryn S.; Hedley, Peter E.; Liu, Hui; Toth, Ian K.; Salmond, George P. C.

2008-01-01

Erwinia carotovora subsp. atroseptica is an enterobacterial phytopathogen causing economically significant soft rot disease. Pathogenesis is mediated by multiple secreted virulence factors, many of which are secreted by the type II (Out) secretion system. DsbA catalyzes the introduction of disulfide bonds into periplasmic and secreted proteins. In this study, the extracellular proteome (secretome) of wild type E. carotovora subsp. atroseptica SCRI1043, and dsbA and out mutants, was analyzed by spectral counting mass spectrometry. This revealed that dsbA inactivation had a huge impact on the secretome and identified diverse DsbA- and Out-dependent secreted proteins, representing known, predicted, and novel candidate virulence factors. Further characterization of the dsbA mutant showed that secreted enzyme activities, motility, production of the quorumsensing signal, and virulence were absent or substantially reduced. The impact of DsbA on secreted virulence factor production was mediated at multiple levels, including impacting on the Out secretion system and the virulence gene regulatory network. Transcriptome analyses revealed that the abundance of a broad, but defined, set of transcripts, including many virulence factors, was altered in the dsbA mutant, identifying a new virulence regulon responsive to extracytoplasmic conditions. In conclusion, DsbA plays a crucial, multifaceted role in the pathogenesis of E. carotovora subsp. atroseptica. PMID:18562317

Genetic Diversity of Streptococcus equi subsp. zooepidemicus and Doxycycline Resistance in Kennelled Dogs

PubMed Central

Chalker, Victoria J.; Waller, Andrew; Webb, Katy; Spearing, Emma; Crosse, Patricia; Brownlie, Joe

2012-01-01

The genetic diversity and antibiotic resistance profiles of 38 Streptococcus equi subsp. zooepidemicus isolates were determined from a kennelled canine population during two outbreaks of hemorrhagic pneumonia (1999 to 2002 and 2007 to 2010). Analysis of the szp gene hypervariable region and the 16S-23S rRNA intergenic spacer region and multilocus sequence typing (MLST) indicated a predominant tetO-positive, doxycycline-resistant ST-10 strain during 1999 to 2002 and a predominant tetM-positive doxycycline-resistant ST-62 strain during 2007 to 2010. PMID:22495558

Detection of Mycobacterium avium subsp. paratuberculosis in Drinking Water and Biofilms by Quantitative PCR ▿ †

PubMed Central

Beumer, Amy; King, Dawn; Donohue, Maura; Mistry, Jatin; Covert, Terry; Pfaller, Stacy

2010-01-01

It has been suggested that Mycobacterium avium subspecies paratuberculosis has a role in Crohn's disease. The organism may be acquired but is difficult to culture from the environment. We describe a quantitative PCR (qPCR) method to detect M. avium subsp. paratuberculosis in drinking water and the results of its application to drinking water and faucet biofilm samples collected in the United States. PMID:20817803

Analysis of Mycobacterium avium subsp. paratuberculosis mutant libraries reveals loci-dependent transcription biases and strategies to novel mutant discovery

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) is the etiologic agent of Johne’s disease in ruminants and it has been implicated as a cause of Crohn’s disease in humans. The generation of comprehensive random mutant banks by transposon mutagenesis is a fundamental wide genomic technology utilized...

Complete Genome Sequence of Clavibacter michiganensis subsp. insidiosus R1-1 Using PacBio Single-Molecule Real-Time Technology

PubMed Central

Lu, You; Samac, Deborah A.; Glazebrook, Jane

2015-01-01

We report here the complete genome sequence of Clavibacter michiganensis subsp. insidiosus R1-1, isolated in Minnesota, USA. The R1-1 genome, generated by a de novo assembly of PacBio sequencing data, is the first complete genome sequence available for this subspecies. PMID:25953184

Evaluation of two mutants of Mycobacterium avium subsp. paratuberculosis as candidates for a live attenuated vaccine for Johne's disease

USDA-ARS?s Scientific Manuscript database

Efforts to control Johne’s disease (JD), caused by Mycobacterium avium subsp. paratuberculosis (Map), has been difficult because of a lack of an effective vaccine. To address this problem we examined the potential of targeted gene disruption as a method to develop candidate vaccines with impaired c...

Three new flavonoids, hippophins K-M, from the seed residue of Hippophae rhamnoides subsp. sinensis.

PubMed

Chen, Chao; Gao, Wen; Ou-Yang, Dan-Wei; Zhang, Jing; Kong, De-Yun

2014-01-01

Three new flavonol glycosides (1-3), hippophins K-M, were isolated from the seed residue of Hippophae rhamnoides subsp. sinensis. The chemical structures of these three new compounds were identified by 1D, 2D NMR, mass spectroscopy and high resolution electrospray ionization mass spectrometry spectroscopic data and by comparison with the literature data. This report is a continuous research work on the systematic chemical investigation of plants of the genus Hippophae in our laboratory.

A comparative study of three detection techniques for Leifsonia xyli subsp. xyli, the causal pathogen of sugarcane ratoon stunting disease

USDA-ARS?s Scientific Manuscript database

Ratoon stunting disease (RSD), which is caused by the bacterium Leifsonia xyli subsp. xyli (Lxx), is now recognized worldwide as the most economically devastating disease impacting sugarcane. RSD causes significant yield losses and variety degradation. Diagnosis of RSD is challenging because it does...

Divergent cellular responses during asymptomatic subclinical and clinical states of disease in cows naturally infected with Mycobacterium avium subsp. paratuberculosis

USDA-ARS?s Scientific Manuscript database

Infection of the host with Mycobacterium avium subsp. paratuberculosis (MAP) results in a chronic and progressive enteritis that traverses both subclinical and clinical stages. The mechanism(s) for the shift from asymptomatic subclinical disease state to advanced clinical disease are not fully under...

Complete genomic sequence of Campylobacter jejuni subsp. jejuni HS:19 strain RM1285 that was isolated from packaged chicken

USDA-ARS?s Scientific Manuscript database

Poultry products serve as the main source of Campylobacter jejuni subsp. jejuni (Cjj) infections in humans. Cjj infections are a leading cause of foodborne gastroenteritis and are a prevalent antecedent to Guillain-Barré syndrome (GBS). This study describes the genome of Cjj HS:19 strain RM1285 isol...

Genome Sequence of Streptococcus phocae subsp. salmonis Strain C-4T, Isolated from Atlantic Salmon (Salmo salar).

PubMed

Avendaño-Herrera, Ruben; Suarez, Rudy; Lazo, Eduardo; Bravo, Diego; Llegues, Katerina O; Romalde, Jesús L; Godoy, Marcos G

2014-12-11

Streptococcus phocae subsp. salmonis is a fish pathogen that has an important impact on the Chilean salmon industry. Here, we report the genome sequence of the type strain C-4(T) isolated from Atlantic salmon (Salmo salar), showing a number of interesting features and genes related to its possible virulence factors. Copyright © 2014 Avendaño-Herrera et al.

Differential Expression of In Vivo and In Vitro Protein Profile of Outer Membrane of Acidovorax avenae Subsp. avenae

PubMed Central

Qiu, Hui; Li, Bin; Jabeen, Amara; Li, Liping; Liu, He; Kube, Michael; Xie, Guanlin; Wang, Yanli; Sun, Guochang

2012-01-01

Outer membrane (OM) proteins play a significant role in bacterial pathogenesis. In this work, we examined and compared the expression of the OM proteins of the rice pathogen Acidovorax avenae subsp. avenae strain RS-1, a Gram-negative bacterium, both in an in vitro culture medium and in vivo rice plants. Global proteomic profiling of A. avenae subsp. avenae strain RS-1 comparing in vivo and in vitro conditions revealed the differential expression of proteins affecting the survival and pathogenicity of the rice pathogen in host plants. The shotgun proteomics analysis of OM proteins resulted in the identification of 97 proteins in vitro and 62 proteins in vivo by mass spectrometry. Among these OM proteins, there is a high number of porins, TonB-dependent receptors, lipoproteins of the NodT family, ABC transporters, flagellins, and proteins of unknown function expressed under both conditions. However, the major proteins such as phospholipase and OmpA domain containing proteins were expressed in vitro, while the proteins such as the surface anchored protein F, ATP-dependent Clp protease, OmpA and MotB domain containing proteins were expressed in vivo. This may indicate that these in vivo OM proteins have roles in the pathogenicity of A. avenae subsp. avenae strain RS-1. In addition, the LC-MS/MS identification of OmpA and MotB validated the in silico prediction of the existance of Type VI secretion system core components. To the best of our knowledge, this is the first study to reveal the in vitro and in vivo protein profiles, in combination with LC-MS/MS mass spectra, in silico OM proteome and in silico genome wide analysis, of pathogenicity or plant host required proteins of a plant pathogenic bacterium. PMID:23166741

De Novo whole genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 from blueberry in Georgia, USA

USDA-ARS?s Scientific Manuscript database

This study reports a de novo assembled draft genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 causing blueberry bacterial leaf scorch in Georgia, USA. The BB01 genome is 2,517,579 bp with a G+C content of 51.8% and 2,943 open reading frames (ORFs) and 48 RNA genes....

Sensitive and Specific Enzyme-Linked Immunosorbent Assay for Detecting Serum Antibodies against Mycobacterium avium subsp. paratuberculosis in Fallow Deer

PubMed Central

Prieto, José M.; Balseiro, Ana; Casais, Rosa; Abendaño, Naiara; Fitzgerald, Liam E.; Garrido, Joseba M.; Juste, Ramon A.

2014-01-01

The enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies against Mycobacterium avium subsp. paratuberculosis in wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deer M. avium subsp. paratuberculosis isolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detect M. avium subsp. paratuberculosis antibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations. PMID:24872517

Kinetics of Inactivation of Bacillus subtilis subsp. niger Spores and Staphylococcus albus on Paper by Chlorine Dioxide Gas in an Enclosed Space

PubMed Central

Wang, Tao; Wu, Jinhui; Hao, Limei; Yi, Ying; Zhang, Zongxing

2016-01-01

ABSTRACT Bacillus subtilis subsp. niger spore and Staphylococcus albus are typical biological indicators for the inactivation of airborne pathogens. The present study characterized and compared the behaviors of B. subtilis subsp. niger spores and S. albus in regard to inactivation by chlorine dioxide (ClO2) gas under different gas concentrations and relative humidity (RH) conditions. The inactivation kinetics under different ClO2 gas concentrations (1 to 5 mg/liter) were determined by first-order and Weibull models. A new model (the Weibull-H model) was established to reveal the inactivation tendency and kinetics for ClO2 gas under different RH conditions (30 to 90%). The results showed that both the gas concentration and RH were significantly (P < 0.05) and positively correlated with the inactivation of the two chosen indicators. There was a rapid improvement in the inactivation efficiency under high RH (>70%). Compared with the first-order model, the Weibull and Weibull-H models demonstrated a better fit for the experimental data, indicating nonlinear inactivation behaviors of the vegetative bacteria and spores following exposure to ClO2 gas. The times to achieve a six-log reduction of B. subtilis subsp. niger spore and S. albus were calculated based on the established models. Clarifying the kinetics of inactivation of B. subtilis subsp. niger spores and S. albus by ClO2 gas will allow the development of ClO2 gas treatments that provide an effective disinfection method. IMPORTANCE Chlorine dioxide (ClO2) gas is a novel and effective fumigation agent with strong oxidization ability and a broad biocidal spectrum. The antimicrobial efficacy of ClO2 gas has been evaluated in many previous studies. However, there are presently no published models that can be used to describe the kinetics of inactivation of airborne pathogens by ClO2 gas under different gas concentrations and RH conditions. The first-order and Weibull (Weibull-H) models established in this study can

Kinetics of Inactivation of Bacillus subtilis subsp. niger Spores and Staphylococcus albus on Paper by Chlorine Dioxide Gas in an Enclosed Space.

PubMed

Wang, Tao; Wu, Jinhui; Qi, Jiancheng; Hao, Limei; Yi, Ying; Zhang, Zongxing

2016-05-15

Bacillus subtilis subsp. niger spore and Staphylococcus albus are typical biological indicators for the inactivation of airborne pathogens. The present study characterized and compared the behaviors of B. subtilis subsp. niger spores and S. albus in regard to inactivation by chlorine dioxide (ClO2) gas under different gas concentrations and relative humidity (RH) conditions. The inactivation kinetics under different ClO2 gas concentrations (1 to 5 mg/liter) were determined by first-order and Weibull models. A new model (the Weibull-H model) was established to reveal the inactivation tendency and kinetics for ClO2 gas under different RH conditions (30 to 90%). The results showed that both the gas concentration and RH were significantly (P < 0.05) and positively correlated with the inactivation of the two chosen indicators. There was a rapid improvement in the inactivation efficiency under high RH (>70%). Compared with the first-order model, the Weibull and Weibull-H models demonstrated a better fit for the experimental data, indicating nonlinear inactivation behaviors of the vegetative bacteria and spores following exposure to ClO2 gas. The times to achieve a six-log reduction of B. subtilis subsp. niger spore and S. albus were calculated based on the established models. Clarifying the kinetics of inactivation of B. subtilis subsp. niger spores and S. albus by ClO2 gas will allow the development of ClO2 gas treatments that provide an effective disinfection method. Chlorine dioxide (ClO2) gas is a novel and effective fumigation agent with strong oxidization ability and a broad biocidal spectrum. The antimicrobial efficacy of ClO2 gas has been evaluated in many previous studies. However, there are presently no published models that can be used to describe the kinetics of inactivation of airborne pathogens by ClO2 gas under different gas concentrations and RH conditions. The first-order and Weibull (Weibull-H) models established in this study can characterize and

Purulent pericarditis and pneumonia caused by Streptococcus equi subsp. zooepidemicus.

PubMed

Held, Jürgen; Schmitz, Roland; van der Linden, Mark; Nührenberg, Thomas; Häcker, Georg; Neumann, Franz-Josef

2014-02-01

Purulent pericarditis is a life-threatening disease that usually manifests following bacteraemia or through spreading from an intrathoracic focus. Only a few cases of this disease have been reported with Lancefield group C streptococci as aetiological agents, and the primary focus in these infections remains unknown. We report a case of purulent pericarditis with septic and cardiogenic shock, caused by Streptococcus equi subsp. zooepidemicus (group C) in a 51-year-old patient. The pathogen was possibly contracted through contact with horses. Most probably, it caused initially pneumonia before spreading to the pericardium, either directly or via the bloodstream. A combined therapeutic approach, consisting of antibiotic therapy and repeated pericardial drainage, was necessary to ensure a clinical cure. After discharge, long-term follow-up for development of constrictive pericarditis is considered mandatory.

New genes of Xanthomonas citri subsp. citri involved in pathogenesis and adaptation revealed by a transposon-based mutant library

PubMed Central

2009-01-01

Background Citrus canker is a disease caused by the phytopathogens Xanthomonas citri subsp. citri, Xanthomonas fuscans subsp. aurantifolli and Xanthomonas alfalfae subsp. citrumelonis. The first of the three species, which causes citrus bacterial canker type A, is the most widely spread and severe, attacking all citrus species. In Brazil, this species is the most important, being found in practically all areas where citrus canker has been detected. Like most phytobacterioses, there is no efficient way to control citrus canker. Considering the importance of the disease worldwide, investigation is needed to accurately detect which genes are related to the pathogen-host adaptation process and which are associated with pathogenesis. Results Through transposon insertion mutagenesis, 10,000 mutants of Xanthomonas citri subsp. citri strain 306 (Xcc) were obtained, and 3,300 were inoculated in Rangpur lime (Citrus limonia) leaves. Their ability to cause citrus canker was analyzed every 3 days until 21 days after inoculation; a set of 44 mutants showed altered virulence, with 8 presenting a complete loss of causing citrus canker symptoms. Sequencing of the insertion site in all 44 mutants revealed that 35 different ORFs were hit, since some ORFs were hit in more than one mutant, with mutants for the same ORF presenting the same phenotype. An analysis of these ORFs showed that some encoded genes were previously known as related to pathogenicity in phytobacteria and, more interestingly, revealed new genes never implicated with Xanthomonas pathogenicity before, including hypothetical ORFs. Among the 8 mutants with no canker symptoms are the hrpB4 and hrpX genes, two genes that belong to type III secretion system (TTSS), two hypothetical ORFS and, surprisingly, the htrA gene, a gene reported as involved with the virulence process in animal-pathogenic bacteria but not described as involved in phytobacteria virulence. Nucleic acid hybridization using labeled cDNA probes showed

Complete genomic sequences of two salmonella enterica subsp. enterica serogroup C2 (O:6,8) strains from central California

USDA-ARS?s Scientific Manuscript database

Salmonella enteric subsp. enterica strains RM11060, serotype 6,8:d:-, and RM11065, serotype 6,8:-:e,n,z15, were isolated from environmental sampling in Central California in 2009. We report the complete genome sequences and annotation of these two strains. These genomic sequences are distinct and wi...

Draft Genome Sequence of Salmonella enterica subsp. enterica Serovar Infantis Strain SPE101, Isolated from a Chronic Human Infection

PubMed Central

Iriarte, Andrés; Giner-Lamia, Joaquín; Betancor, Laura; Astocondor, Lizeth; Cestero, Juan J.; Ochoa, Theresa; García, Coralith; Puente, José L.; Chabalgoity, José A.

2017-01-01

ABSTRACT We report a 4.99-Mb draft genome sequence of Salmonella enterica subsp. enterica serovar Infantis strain SPE101, isolated from feces of a 5-month-old breast-fed female showing diarrhea associated with severe dehydration and malnutrition. The infection prolonged for 6 months despite antibiotic treatment. PMID:28729277

Complete Genome Sequence of Clavibacter michiganensis subsp. insidiosus R1-1 Using PacBio Single-Molecule Real-Time Technology.

PubMed

Lu, You; Samac, Deborah A; Glazebrook, Jane; Ishimaru, Carol A

2015-05-07

We report here the complete genome sequence of Clavibacter michiganensis subsp. insidiosus R1-1, isolated in Minnesota, USA. The R1-1 genome, generated by a de novo assembly of PacBio sequencing data, is the first complete genome sequence available for this subspecies. Copyright © 2015 Lu et al.

Structural characterization of the lipid A region of Aeromonas salmonicida subsp. salmonicida lipopolysaccharide.

PubMed

Wang, Zhan; Li, Jianjun; Altman, Eleonora

2006-12-11

The lipid A components of Aeromonas salmonicida subsp. salmonicida from strains A449, 80204-1 and an in vivo rough isolate were isolated by mild acid hydrolysis of the lipopolysaccharide. Structural studies carried out by a combination of fatty acid, electrospray ionization-mass spectrometry and nuclear magnetic resonance analyses confirmed that the structure of lipid A was conserved among different isolates of A. salmonicida subsp. salmonicida. All analyzed strains contained three major lipid A molecules differing in acylation patterns corresponding to tetra-, penta- and hexaacylated lipid A species and comprising 4'-monophosphorylated beta-2-amino-2-deoxy-d-glucopyranose-(1-->6)-2-amino-2-deoxy-d-glucopyranose disaccharide, where the reducing end 2-amino-2-deoxy-d-glucose was present primarily in the alpha-pyranose form. Electrospray ionization-tandem mass spectrometry fragment pattern analysis, including investigation of the inner-ring fragmentation, allowed the localization of fatty acyl residues on the disaccharide backbone of lipid A. The tetraacylated lipid A structure containing 3-(dodecanoyloxy)tetradecanoic acid at N-2',3-hydroxytetradecanoic acid at N-2 and 3-hydroxytetradecanoic acid at O-3, respectively, was found. The pentaacyl lipid A molecule had a similar fatty acid distribution pattern and, additionally, carried 3-hydroxytetradecanoic acid at O-3'. In the hexaacylated lipid A structure, 3-hydroxytetradecanoic acid at O-3' was esterified with a secondary 9-hexadecenoic acid. Interestingly, lipid A of the in vivo rough isolate contained predominantly tetra- and pentaacylated lipid A species suggesting that the presence of the hexaacyl lipid A was associated with the smooth-form lipopolysaccharide.

A combination of direct viable count and fluorescence in situ hybridization for specific enumeration of viable Lactobacillus delbrueckii subsp.bulgaricus and Streptococcus thermophilus.

PubMed

García-Hernández, J; Moreno, Y; Amorocho, C M; Hernández, M

2012-03-01

We have developed a direct viable count (DVC)-FISH procedure for quickly and easily discriminating between viable and nonviable cells of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains, the traditional yogurt bacteria. direct viable count method has been modified and adapted for Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus analysis by testing different times of incubation and concentrations of DNA-gyrase inhibitors. DVC procedure has been combined with fluorescent in situ hybridization (FISH) for the specific detection of viable cells of both bacteria with specific rRNA oligonucleotide probes (DVC-FISH). Of the four antibiotics tested (novobiocin, nalidixic acid, pipemidic acid and ciprofloxacin), novobiocin was the most effective for DVC method and the optimum incubation time was 7 h for both bacteria. The number of viable cells was obtained by the enumeration of specific hybridized cells that were elongated at least twice their original length for Lactobacillus and twice their original size for Streptococcus. This technique was successfully applied to detect viable cells in inoculated faeces. Results showed that this DVC-FISH procedure is a quick and culture-independent useful method to specifically detect viable Lact. delbrueckii subsp. bulgaricus and Strep. thermophilus in different samples, being applied for the first time to lactic acid bacteria. © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

Influence of different proteolytic strains of Streptococcus thermophilus in co-culture with Lactobacillus delbrueckii subsp. bulgaricus on the metabolite profile of set-yoghurt.

PubMed

Settachaimongkon, Sarn; Nout, M J Robert; Antunes Fernandes, Elsa C; Hettinga, Kasper A; Vervoort, Jacques M; van Hooijdonk, Toon C M; Zwietering, Marcel H; Smid, Eddy J; van Valenberg, Hein J F

2014-05-02

Proto-cooperation between Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus is one of the key factors that determine the fermentation process and final quality of yoghurt. In this study, the interaction between different proteolytic strains of S. thermophilus and L. delbrueckii subsp. bulgaricus was investigated in terms of microbial growth, acidification and changes in the biochemical composition of milk during set-yoghurt fermentation. A complementary metabolomics approach was applied for global characterization of volatile and non-volatile polar metabolite profiles of yoghurt associated with proteolytic activity of the individual strains in the starter cultures. The results demonstrated that only non-proteolytic S. thermophilus (Prt-) strain performed proto-cooperation with L. delbrueckii subsp. bulgaricus. The proto-cooperation resulted in significant higher populations of the two species, faster milk acidification, significant abundance of aroma volatiles and non-volatile metabolites desirable for a good organoleptic quality of yoghurt. Headspace SPME-GC/MS and (1)H NMR resulted in the identification of 35 volatiles and 43 non-volatile polar metabolites, respectively. Furthermore, multivariate statistical analysis allows discriminating set-yoghurts fermented by different types of starter cultures according to their metabolite profiles. Our finding underlines that selection of suitable strain combinations in yoghurt starters is important for achieving the best technological performance regarding the quality of product. Copyright © 2014 Elsevier B.V. All rights reserved.

Secondary Metabolites, Glandular Trichomes and Biological Activity of Sideritis montana L. subsp. montana from Central Italy.

PubMed

Venditti, Alessandro; Bianco, Armandodoriano; Frezza, Claudio; Serafini, Mauro; Giacomello, Ginevra; Giuliani, Claudia; Bramucci, Massimo; Quassinti, Luana; Lupidi, Giulio; Lucarini, Domenico; Papa, Fabrizio; Maggi, Filippo

2016-10-01

Sideritis montana subsp. montana is a small annual herb occurring in countries bordering the Mediterranean and Balkan regions. The secondary metabolism of this plant has not been fully explored so far. The aim of the present study was to understand the complex mixture of secondary metabolites and the type of secretory structures. The polar constituents were isolated by column chromatography from the ethanolic extract, and their structure was elucidated by NMR and MS. The essential oil was isolated by hydrodistillation and analysed by GC/MS. The plant indumentum was studied by light and scanning electron microscopy. To complete the work, the essential oil antioxidant activity and cytotoxicity on tumor cells were evaluated by DPPH, ABTS, FRAP, and MTT methods. Four different classes of secondary metabolites were isolated, namely flavonoids, caffeoylquinic derivatives, glycosidic hydroquinones and iridoids. The essential oil was mainly characterized by sesquiterpenene hydrocarbons. Peltate and long-capitate hairs were the main sites where terpenes and polar constituents are produced. The secondary metabolites found in S. montana subsp. montana are of chemotaxonomic interest, some of them being typical of the genus Sideritis. The trichomes types observed partially differ from those described in other members of the genus Sideritis. The essential oil showed noteworthy inhibition on tumor cells. © 2016 Wiley-VHCA AG, Zürich.

Multilocus Sex Determination Revealed in Two Populations of Gynodioecious Wild Strawberry, Fragaria vesca subsp. bracteata.

PubMed

Ashman, Tia-Lynn; Tennessen, Jacob A; Dalton, Rebecca M; Govindarajulu, Rajanikanth; Koski, Matthew H; Liston, Aaron

2015-10-19

Gynodioecy, the coexistence of females and hermaphrodites, occurs in 20% of angiosperm families and often enables transitions between hermaphroditism and dioecy. Clarifying mechanisms of sex determination in gynodioecious species can thus illuminate sexual system evolution. Genetic determination of gynodioecy, however, can be complex and is not fully characterized in any wild species. We used targeted sequence capture to genetically map a novel nuclear contributor to male sterility in a self-pollinated hermaphrodite of Fragaria vesca subsp. bracteata from the southern portion of its range. To understand its interaction with another identified locus and possibly additional loci, we performed crosses within and between two populations separated by 2000 km, phenotyped the progeny and sequenced candidate markers at both sex-determining loci. The newly mapped locus contains a high density of pentatricopeptide repeat genes, a class commonly involved in restoration of fertility caused by cytoplasmic male sterility. Examination of all crosses revealed three unlinked epistatically interacting loci that determine sexual phenotype and vary in frequency between populations. Fragaria vesca subsp. bracteata represents the first wild gynodioecious species with genomic evidence of both cytoplasmic and nuclear genes in sex determination. We propose a model for the interactions between these loci and new hypotheses for the evolution of sex determining chromosomes in the subdioecious and dioecious Fragaria. Copyright © 2015 Ashman et al.

Biofilm Formation and Morphotypes of Salmonella enterica subsp.arizonae Differs from Those of Other Salmonella enterica Subspecies in Isolates from Poultry Houses.

PubMed

Lamas, A; Fernandez-No, I C; Miranda, J M; Vázquez, B; Cepeda, A; Franco, C M

2016-07-01

Salmonella serovars are responsible for foodborne diseases around the world. The ability to form biofilms allows microorganisms to survive in the environment. In this study, 73 Salmonella strains, belonging to four different subspecies, were isolated from poultry houses and foodstuffs and tested. Biofilm formation was measured at four different temperatures and two nutrient concentrations. Morphotypes and cellulose production were evaluated at three different temperatures. The presence of several genes related to biofilm production was also examined. All strains and subspecies of Salmonella had the ability to form biofilms, and 46.57% of strains produced biofilms under all conditions tested. Biofilm formation was strain dependent and varied according to the conditions. This is the first study to analyze biofilm formation in a wide number of Salmonella enterica subsp. arizonae strains, and no direct relationship between the high prevalence of Salmonella enterica subsp. arizonae strains and their ability to form biofilm was established. Morphotypes and cellulose production varied as the temperature changed, with 20°C being the optimum temperature for expression of the red, dry, and rough morphotype and cellulose. Salmonella enterica subsp. arizonae, whose morphotype is poorly studied, only showed a smooth and white morphotype and lacked the csgD and gcpA genes that are implicated in biofilm production. Thus, Salmonella biofilm formation under different environmental conditions is a public health problem because it can survive and advance through the food chain to reach the consumer.

Assessment of stress tolerance acquisition in the heat-tolerant derivative strains of Bifidobacterium animalis subsp. lactis BB-12 and Lactobacillus rhamnosus GG.

PubMed

Aakko, J; Sánchez, B; Gueimonde, M; Salminen, S

2014-07-01

The purpose of this study was to investigate the heat-shock response at molecular level in Lactobacillus rhamnosus GG, Bifidobacterium animalis subsp. lactis BB-12 and their heat-tolerant derivatives and to characterize the changes that make the derivatives more robust in terms of heat stress. The study strains were exposed for 2 h to a heat-shock treatment, Bif. animalis subsp. lactis BB-12 and its derivative at 50°C and the Lact. rhamnosus GG and its derivative at 60°C. Protein synthesis before and after heat shock was examined using proteomics and RT-qPCR. The analysis revealed that the regulation of seven proteins in both strain pairs was modified as a response to heat or between the original and the derivative strain. The comparison of wild-type strains and the heat-tolerant derivatives suggests that the acquisition of heat tolerance in the Bif. animalis subsp. lactis BB-12 derivative is due to a slightly increased constitutive level of chaperones, while in Lact. rhamnosus GG derivative, the main reason seems to be a higher ability to induce the production of chaperones. This study revealed possible markers of heat tolerance in B. lactis and Lact. rhamnosus strains. This study increases our knowledge on how Lactobacillus and Bifidobacterium strains may acquire heat tolerance. These findings may be useful for improving the heat tolerance of existing probiotic strains as well as screening new heat-tolerant strains. © 2014 The Society for Applied Microbiology.

Sideritis romana L. subsp. purpurea (Tal. ex Benth.) Heywood, a new chemotype from Montenegro.

PubMed

Garzoli, Stefania; Božović, Mijat; Baldisserotto, Anna; Andreotti, Elisa; Pepi, Federico; Tadić, Vanja; Manfredini, Stefano; Ragno, Rino

2018-05-01

A study on essential oil fractions of the Western Balkan endemic Sideritis romana L. subsp. purpurea (Tal. ex Benth.) Heywood collected in Montenegro is reported. The 24-h systematic steam distillation extraction procedure was performed. The gas chromatographic/mass spectrometric (GC/MS) analysis of the fractions showed γ-elemene and spathulenol as two main constituents, revealing a new chemotype of this plant species. Although varying in the content of these two main compounds, which makes the fractions quite different between each other, evaluation of the anti-Candida activity showed the lack of any significant efficacy.

Neisseria elongata subsp elongata infective endocarditis following endurance exercise

PubMed Central

Jenkins, Joanne May; Fife, Amanda; Baghai, Max; Dworakowski, Rafal

2015-01-01

A 31-year-old Argentinian woman presented with a 3-week history of fever, night sweats, myalgia and lethargy following a work trip to Uganda where she ran a marathon. Malarial screens were negative but C reactive protein, erythrocyte sedimentation rate and neutrophil count were raised and she was anaemic. A new pansystolic murmur was heard over the mitral valve and the transthoracic echocardiogram showed a large vegetation (>1 cm) with at least moderate mitral regurgitation. Blood cultures grew Neisseria elongata, subsp elongata treated initially with ceftriaxone then oral ciprofloxacin to complete 4 weeks of treatment. CT scan revealed a wedge-shaped area of low attenuation in the spleen in keeping with a splenic infarct. Seven days postadmission, the patient underwent a successful mitral valve repair. Recovery was complicated by a likely embolic infarct in the right frontal lobe, but the patient was discharged 12 days postoperative with no neurological sequelae. PMID:26655669

A novel dimeric flavonol glycoside from Cynanchum acutum subsp. sibiricum.

PubMed

Yuan, Si-Wen; Dai, Wei; Pan, Xin-Hui; Lu, Yan; Chen, Dao-Feng; Wang, Qi

2018-06-11

A novel dimeric flavonol glycoside, Cynanflavoside A (1), together with six analogues, kaempferol-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside (2), quercetin-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside (3), kaempferol-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranoside (4), quercetin-3-O-α-L-rhamnopyranosyl-(1→2)-β-D-xylopyranoside (5), kaempferol-3-O-β-D-glucopyranosyl-7-O-α-L-rhamnopyranoside (6), and quercetin-3-O-galactoside (7) were isolated from the n-butyl alcohol extract of Cynanchum acutum subsp. sibiricum. Their structures were determined spectroscopically and compared with previously reported spectral data. All compounds were evaluated for their anti-complementary activity in vitro, and only compound 5 exhibited anti-complement effects with CH 50 value of 0.33 mM.

Sensitive and specific enzyme-linked immunosorbent assay for detecting serum antibodies against Mycobacterium avium subsp. paratuberculosis in fallow deer.

PubMed

Prieto, José M; Balseiro, Ana; Casais, Rosa; Abendaño, Naiara; Fitzgerald, Liam E; Garrido, Joseba M; Juste, Ramon A; Alonso-Hearn, Marta

2014-08-01

The enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies against Mycobacterium avium subsp. paratuberculosis in wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deer M. avium subsp. paratuberculosis isolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detect M. avium subsp. paratuberculosis antibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Draft Genome Sequences of 20 Salmonella enterica subsp. enterica Serovar Typhimurium Strains Isolated from Swine in Santa Catarina, Brazil.

PubMed

Seribelli, Amanda Aparecida; Frazão, Miliane Rodrigues; Gonzales, Júlia Cunha; Cao, Guojie; Leon, Maria Sanchez; Kich, Jalusa Deon; Allard, Marc William; Falcão, Juliana Pfrimer

2018-04-19

Salmonellosis is a disease with a high incidence worldwide, and Salmonella enterica subsp. enterica serovar Typhimurium is one of the most clinically important serovars. We report here the draft genome sequences of 20 S. Typhimurium strains isolated from swine in Santa Catarina, Brazil. These draft genomes will improve our understanding of S. Typhimurium in Brazil.

Draft Genome Sequence of Salmonella enterica subsp. enterica Serovar Infantis Strain SPE101, Isolated from a Chronic Human Infection.

PubMed

Iriarte, Andrés; Giner-Lamia, Joaquín; Silva, Claudia; Betancor, Laura; Astocondor, Lizeth; Cestero, Juan J; Ochoa, Theresa; García, Coralith; Puente, José L; Chabalgoity, José A; García-Del Portillo, Francisco

2017-07-20

We report a 4.99-Mb draft genome sequence of Salmonella enterica subsp. enterica serovar Infantis strain SPE101, isolated from feces of a 5-month-old breast-fed female showing diarrhea associated with severe dehydration and malnutrition. The infection prolonged for 6 months despite antibiotic treatment. Copyright © 2017 Iriarte et al.

Application of halophilic nuclease H of Micrococcus varians subsp. halophilus to commercial production of flavoring agent 5'-GMP.

PubMed Central

Kamekura, M; Hamakawa, T; Onishi, H

1982-01-01

RNA was degraded at 60 degrees C for 24 h by halophilic nuclease H in supernatants from broth cultures of Micrococcus varians subsp. halophilus containing 12% NaCl. Since contaminating 5'-nucleotidase exhibited almost no activity under these conditions, the 5'-GMP formed could be recovered from the reaction mixture, and the yield was 805 mg from 5 g of RNA. PMID:6184020

First report of bacterial blight of crucifers caused by Pseudomonas cannabina pv. alisalensis in Minnesota on arugula (Eruca vesicaria subsp. sativa)

USDA-ARS?s Scientific Manuscript database

In 2011, bacterial blight of arugula (Eruca vesicaria subsp. sativa; cv. Roquette) was observed in organically grown plants under overhead irrigation near Delano, MN. Approximately 80 to 100% of each planting was affected. Blue-green fluorescent pseudomonads were isolated consistently on King’s Medi...

Analysis of the Immune Response to a Major Membrane Protein of Mycobacterium avium subsp. paratuberculosis in Experimentally and Naturally Infected Cattle

USDA-ARS?s Scientific Manuscript database

The 35 kDa major membrane protein (MMP) of Mycobacterium avium subsp. paratuberculosis (Map) is implicated in the pathogenesis of paratuberculosis (Ptb) in cattle. Understanding the immune response to MMP could reveal how Map evades immune elimination and provide information needed for developing a ...

Mycobacterium avium subsp. paratuberculosis PPE Protein MAP1152 and Conserved Protein MAP1156 are Antigenic in Experimentally and Naturally Infected Cattle

USDA-ARS?s Scientific Manuscript database

Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne’s Disease (JD) in ruminants resulting in significant production losses. An insertion mutation upstream from the MAP1152-MAP1156 region causes a change in colony morphotype and results in an attenuated phenotype in bovine monocyte derive...

Sex ratio and reproductive effort in the dioecious Juniperus communis subsp. alpina (Suter) Celak. (Cupressaceae) along an altitudinal gradient.

PubMed

Ortiz, Pedro Luis; Arista, Montserrat; Talavera, Salvador

2002-02-01

The hypothesis that reproductive cost differs between sexes was tested in Juniperus communis subsp. alpina along an altitudinal gradient. Sex ratio (male : female) increased significantly with elevation, and above 2,600 m it was significantly male-biased. The reproductive effort was markedly greater for females than for males at all elevations. However, over 3 years of study, the growth of the females, measured as elongation of the main axes, was similar to that of the males. In both sexes, growth decreased with increasing elevation. Neither size of the ripe seed cones, nor the number of developed seeds per cone varied with elevation. The percentage of filled seeds was significantly greater at higher elevations indicating more favourable conditions for wind pollination in these stands. However, cone production decreased with elevation and so, reproductive success of J. communis subsp. alpina in Sierra Nevada decreases towards both upper and lower altitudinal distribution limits. The results do not support the hypothesis of differential reproductive cost between sexes; thus, alternative arguments to explain the altitudinal variation of sex ratio are discussed.

Development and Validation of a Liquid Medium (M7H9C) for Routine Culture of Mycobacterium avium subsp. paratuberculosis To Replace Modified Bactec 12B Medium

PubMed Central

Whittington, Ann-Michele; Waldron, Anna; Begg, Douglas J.; de Silva, Kumi; Purdie, Auriol C.; Plain, Karren M.

2013-01-01

Liquid culture of Mycobacterium avium subsp. paratuberculosis from clinical samples, such as feces, is the most sensitive antemortem test for the diagnosis of Johne's disease in ruminants. In Australia, New Zealand, the United States, and some other countries, the Bactec 460 system with modified Bactec 12B medium (Becton, Dickinson) has been the most commonly used liquid culture system, but it was discontinued in 2012. In this study, a new liquid culture medium, M7H9C, was developed. It consists of a Middlebrook 7H9 medium base with added Casitone, albumin, dextrose, catalase, egg yolk, mycobactin J, and a cocktail of antibiotics. We found that polyoxyethylene stearate (POES) was not essential for the cultivation of M. avium subsp. paratuberculosis in either the Bactec 12B or the M7H9C medium. The limit of detection determined using pure cultures of the C and S strains of M. avium subsp. paratuberculosis was 7 bacilli per 50 μl inoculum in the two media. The new medium was validated using 784 fecal and tissue samples from sheep and cattle, >25% of which contained viable M. avium subsp. paratuberculosis. Discrepant results for the clinical samples between the two media were mostly associated with samples that contained <10 viable bacilli per gram, but these results were relatively uncommon, and the performances of the two media were not significantly different. M7H9C medium was less than half the cost of the Bactec 12B medium and did not require regular examination during incubation, but a confirmatory IS900 PCR test had to be performed on every culture after the predetermined incubation period. PMID:24048541

Protein expression vector and secretion signal peptide optimization to drive the production, secretion, and functional expression of the bacteriocin enterocin A in lactic acid bacteria.

PubMed

Borrero, Juan; Jiménez, Juan J; Gútiez, Loreto; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2011-10-20

Replacement of the leader sequence (LS) of the bacteriocin enterocin A (LS(entA)) by the signal peptides (SP) of the protein Usp45 (SP(usp45)), and the bacteriocins enterocin P (SP(entP)), and hiracin JM79 (SP(hirJM79)) permits the production, secretion, and functional expression of EntA by different lactic acid bacteria (LAB). Chimeric genes encoding the SP(usp45), the SP(entP), and the SP(hirJM79) fused to mature EntA plus the EntA immunity genes (entA+entiA) were cloned into the expression vectors pNZ8048 and pMSP3545, under control of the inducible P(nisA) promoter, and in pMG36c, under control of the constitutive P(32) promoter. The amount, antimicrobial activity, and specific antimicrobial activity of the EntA produced by the recombinant Lactococcus lactis, Enterococcus faecium, E. faecalis, Lactobacillus sakei and Pediococcus acidilactici hosts varied depending on the signal peptide, the expression vector, and the host strain. However, the antimicrobial activity and the specific antimicrobial activity of the EntA produced by most of the LAB transformants was lower than expected from their production. The supernatants of the recombinant L. lactis NZ9000 (pNZUAI) and L. lactis NZ9000 (pNZHAI), overproducers of EntA, showed a 1.2- to 5.1-fold higher antimicrobial activity than that of the natural producer E. faecium T136 against different Listeria spp. Copyright © 2011 Elsevier B.V. All rights reserved.

Biodiversity and dynamics of meat fermentations: the contribution of molecular methods for a better comprehension of a complex ecosystem.

PubMed

Cocolin, Luca; Dolci, Paola; Rantsiou, Kalliopi

2011-11-01

The ecology of fermented sausages is complex and includes different species and strains of bacteria, yeasts and molds. The developments in the field of molecular biology, allowed for new methods to become available, which could be applied to better understand dynamics and diversity of the microorganisms involved in the production of sausages. Methods, such as denaturing gradient gel electrophoresis (DGGE), employed as a culture-independent approach, allow to define the microbial dynamics during the fermentation and ripening. Such approach has highlighted that two main species of lactic acid bacteria, namely Lactobacillus sakei and Lb. curvatus, are involved in the transformation process and that they are accompanied by Staphylococcus xylosus, as representative of the coagulase-negative cocci. These findings were repeatedly confirmed in different regions of the world, mainly in the Mediterranean countries where dry fermented sausages have a long tradition and history. The application of molecular methods for the identification and characterization of isolated strains from fermentations highlighted a high degree of diversity within the species mentioned above, underlining the need to better follow strain dynamics during the transformation process. While there is an important number of papers dealing with bacterial ecology by using molecular methods, studies on mycobiota of fermented sausages are just a few. This review reports on how the application of molecular methods made possible a better comprehension of the sausage fermentations, opening up new fields of research that in the near future will allow to unravel the connection between sensory properties and co-presence of multiple strains of the same species. Copyright © 2011 Elsevier Ltd. All rights reserved.

Detection and Genomic Characterization of Motility in Lactobacillus curvatus: Confirmation of Motility in a Species outside the Lactobacillus salivarius Clade

PubMed Central

Cousin, Fabien J.; Lynch, Shónagh M.; Harris, Hugh M. B.; McCann, Angela; Lynch, Denise B.; Neville, B. Anne; Irisawa, Tomohiro; Okada, Sanae; Endo, Akihito

2014-01-01

Lactobacillus is the largest genus within the lactic acid bacteria (LAB), with almost 180 species currently identified. Motility has been reported for at least 13 Lactobacillus species, all belonging to the Lactobacillus salivarius clade. Motility in lactobacilli is poorly characterized. It probably confers competitive advantages, such as superior nutrient acquisition and niche colonization, but it could also play an important role in innate immune system activation through flagellin–Toll-like receptor 5 (TLR5) interaction. We now report strong evidence of motility in a species outside the L. salivarius clade, Lactobacillus curvatus (strain NRIC 0822). The motility of L. curvatus NRIC 0822 was revealed by phase-contrast microscopy and soft-agar motility assays. Strain NRIC 0822 was motile at temperatures between 15°C and 37°C, with a range of different carbohydrates, and under varying atmospheric conditions. We sequenced the L. curvatus NRIC 0822 genome, which revealed that the motility genes are organized in a single operon and that the products are very similar (>98.5% amino acid similarity over >11,000 amino acids) to those encoded by the motility operon of Lactobacillus acidipiscis KCTC 13900 (shown for the first time to be motile also). Moreover, the presence of a large number of mobile genetic elements within and flanking the motility operon of L. curvatus suggests recent horizontal transfer between members of two distinct Lactobacillus clades: L. acidipiscis in the L. salivarius clade and L. curvatus in the L. sakei clade. This study provides novel phenotypic, genetic, and phylogenetic insights into flagellum-mediated motility in lactobacilli. PMID:25501479

Detection and genomic characterization of motility in Lactobacillus curvatus: confirmation of motility in a species outside the Lactobacillus salivarius clade.

PubMed

Cousin, Fabien J; Lynch, Shónagh M; Harris, Hugh M B; McCann, Angela; Lynch, Denise B; Neville, B Anne; Irisawa, Tomohiro; Okada, Sanae; Endo, Akihito; O'Toole, Paul W

2015-02-01

Lactobacillus is the largest genus within the lactic acid bacteria (LAB), with almost 180 species currently identified. Motility has been reported for at least 13 Lactobacillus species, all belonging to the Lactobacillus salivarius clade. Motility in lactobacilli is poorly characterized. It probably confers competitive advantages, such as superior nutrient acquisition and niche colonization, but it could also play an important role in innate immune system activation through flagellin–Toll-like receptor 5 (TLR5) interaction. We now report strong evidence of motility in a species outside the L. salivarius clade, Lactobacillus curvatus (strain NRIC0822). The motility of L. curvatus NRIC 0822 was revealed by phase-contrast microscopy and soft-agar motility assays. Strain NRIC 0822 was motile at temperatures between 15 °C and 37 °C, with a range of different carbohydrates, and under varying atmospheric conditions. We sequenced the L. curvatus NRIC 0822 genome, which revealed that the motility genes are organized in a single operon and that the products are very similar (>98.5% amino acid similarity over >11,000 amino acids) to those encoded by the motility operon of Lactobacillus acidipiscis KCTC 13900 (shown for the first time to be motile also). Moreover, the presence of a large number of mobile genetic elements within and flanking the motility operon of L. curvatus suggests recent horizontal transfer between members of two distinct Lactobacillus clades: L. acidipiscis in the L. salivarius clade and L. curvatus inthe L. sakei clade. This study provides novel phenotypic, genetic, and phylogenetic insights into flagellum-mediated motility in lactobacilli.