75 DLL “Plan-Apochromat” or a 100×/1 4 DIC objective Metamorph 7

75 DLL “Plan-Apochromat” or a 100×/1.4 DIC objective. Metamorph 7.5 (Molecular Devices) or NIS-Element (Nikon) software was used for digital analysis and treatment of the images to extract the

number, specific fluorescence intensity and length of stained bacteria. This system allows enumeration and manual differentiation between individually labeled cells, cells in aggregations and/or auto fluorescent particles which can interfere with automated analysis. For the CV6 procedure, cells were scored as viable if their fluorescence intensity was at least 1.5 times greater than the fluorescence background noise. Under our conditions, detection limits for CV6 measurement were 3 × 104 cells ml-1. Co-culture of L. pneumophila and A. Castellanii Axenic cultures of A. INK 128 datasheet castellanii (ATCC 30234) were Roxadustat nmr prepared as previously described [48]. Briefly,

the A. castellanii strain was grown in a 150-cm2 cell culture flask in PYG medium (peptone-yeast extract-glucose) at 30°C for 3 days. Monolayers were developed in 24-well tissue culture plates using Page’s amoeba Saline (PAS) for 24 h at 30°C. Aliquots of 1 × 106 amoebae per well in 24-well tissue culture plates were infected with 10 × 1 ml of L. pneumophila at 1 × 108 cells ml-1 in PAS as described above (MOI 100). The plates were centrifuged at 500 × g for 5 min and incubated for 3 days at 37°C. Then, the monolayer and supernatant were removed and spread on BCYE agar plates. Colonies were counted after 3 days and 10 days of incubation at 37°C. Acknowledgement We thank Gail G. Hardy (Indiana university, Bloomington), Audrey Dumont (CNRS, Marseille), Emilie Fugier (CNRS, Marseille), and Sophie Jarraud (CNRL, France) for discussions and critical reading of the manuscript. This work was supported by a fund from the ANSES (Program ARCL-2005) and a doctoral fellowship (Adrien Ducret) from CIBA

SA. We also thank Yannick Fovet for his helpful comments on the manuscript. We thank Bernard Lascola and Isabelle Pagnier for their generous gifts of LP1 and amoebae. References 1. Fliermans: Ecology of legionella: from data to knowledge ZD1839 with a little wisdom. Microb Ecol 1996, 32:203–228.PubMedCrossRef 2. Muldrow LL, Tyndall RL, Fliermans CB: Application of flow cytometry to studies of pathogenic free-living amoebae. Appl Environ Microbiol 1982, 44:1258–1269.PubMedCentralPubMed 3. Fields BS, Benson RF, Besser RE: Legionella and legionnaires’ disease: 25 years of investigation. Clin Microbiol Rev 2002, 15:506–526.PubMedCentralPubMedCrossRef 4. Fields BS: The molecular ecology of legionellae. Trends Microbiol 1996, 4:286–290.PubMedCrossRef 5. Molmeret M, Horn M, Wagner M, Santic M, Abu Kwaik Y: Amoebae as training grounds for intracellular bacterial pathogens. Appl Environ Microbiol 2005, 71:20–28.PubMedCentralPubMedCrossRef 6. Newton HJ, Ang DKY, van Driel IR, Hartland EL: Molecular pathogenesis of infections caused by legionella pneumophila. Clin Microbiol Rev 2010, 23:274–298.

Poorly aligned positions and divergent regions were eliminated fr

Poorly aligned positions and divergent regions were eliminated from the alignment using Gblocks 0.91b with default settings (Castresana 2000). The congruency of the concatenated Trebouxia-alignment NVP-BEZ235 cost was tested by comparing the topology

in the single ITS and the concatenated ITS-psbF-L trees. Both phylogenies showed similar topologies and the same groups. Maximum parsimony analyses (MP) were performed using the program PAUP* (Swofford 2003). Heuristic searches with 1,000 random taxon addition replicates were conducted with TBR branch swapping and MulTrees option in operation, equally weighted characters and gaps treated as missing data. Bootstrapping was performed based on 2,000 replicates with random sequence additions. Homoplasy

levels were assessed by calculating consistency index (CI), retention index (RI), and rescaled consistency (RC) index from each parsimony search. Nucleotide substitution models were chosen using JModeltest 2.1.1. (Darriba et al. 2012). The Akaike information criterion selected the GTR model (Rodriguez et al. 1990) + I + Γ (estimation of invariant sites and a discrete gamma distribution) for the Trebouxia alignments and TRN model (Tamura and Nei 1993) + Γ for the Asterochloris Autophagy Compound Library mouse alignment as the optimal models. A maximum likelihood analysis (ML) was performed using the program Garli 0.96 (http://​www.​nescent.​org/​wg_​garli/​Main_​Page) with the estimated model (GTR > 6rate, TrN > 010020) and default settings. A nonparametric bootstrap was used to assess robustness of clades, running 2,000 pseudoreplicates. For Bayesian tree inference a Markov

Chain Monte Carlo (MCMC) procedure as implemented in the program MrBayes 3.2. was used (Ronquist and Huelsenbeck 2003). The analyses were performed assuming the general time reversible model of nucleotide substitution including estimation of invariant sites and a discrete gamma distribution with six rate categories (GTR + I + Γ, Rodriguez et al. 1990). A run with 5 million generations starting with a random tree and employing four simultaneous chains was executed. Every 100th tree was saved into a file. Subsequently, the first 25 % of trees were deleted as the “burn in” of the chain. A consensus topology with posterior probabilities for each clade was calculated Prostatic acid phosphatase from the remaining 37,501 trees. Results The final data matrix of the molecular phylogeny of Trebouxia ITS (see Online Resource 2) comprised 101 OTUs with a length of 431 characters, 226 positions of the alignment were parsimony-informative with the following homoplasy levels CI = 0.647, RI = 0.953, RC = 0.617. The concatenated Trebouxia ITS/psbL-J (Fig. 2) phylogeny comprised 75 OTUs with 694 characters, 461 positions were parsimony informative and the homoplasy levels amounted CI = 0.765, RI = 0.958, RC = 0.733. Finally, the Asterochloris ITS phylogeny (Fig.

The t½ was calculated as 0 693/λz [19] The total clearance after

The t½ was calculated as 0.693/λz [19]. The total clearance after oral administration (CL/F) was calculated as dose/AUC∞. Descriptive statistics, including mean values and standard deviations (SDs), were used to summarize the pharmacokinetic data for the two drugs. Statistical analyses were performed using SAS version

9.0.2 software (SAS Institute Inc., Cary, NC, USA). An analysis of variance (ANOVA) was performed on the natural logarithm (ln)-transformed pharmacokinetic parameters (the AUCt, AUC∞, and Cmax), using the general linear models procedures in SAS. The ANOVA model had fixed factors for sequence, treatment, period, and subject within Selleck 5-Fluoracil sequence. The Wilcoxon signed-rank test was used for nonparametric analysis to determine differences in the tmax. If the 90% confidence intervals (CIs) of the AUC and Cmax were located within 80–125% of the statistical interval proposed by the FDA [20], the two drugs would be considered bioequivalent. On the basis of the variability reported in a previous trial in India and the Chinese SFDA guidance [19], the number of subjects required to demonstrate bioequivalence at a significance level of 5% with 90% power was calculated

Bortezomib to be 24. 3 Results 3.1 Demographic Data A total of 24 healthy male Chinese volunteers were enrolled, and all completed the study. The demographic characteristics of the study population are summarized in heptaminol Table 1. Table 1 Baseline demographic and clinical characteristics of the study population (n = 24 healthy Chinese male volunteers) Characteristic Value Age

(years)  Mean [SD] 22.9 [2.7]  Range 19.2–27.1 Weight (kg)  Mean [SD] 63.2 [7.0]  Range 52.0–78.0 Height (cm)  Mean [SD] 171.3 [6.1]  Range 162.0–187.0 Body mass index (kg/m2)  Mean [SD] 21.5 [1.3]  Range 19.3–23.7 SD standard deviation 3.2 Tolerability The tolerability of the two formulations of risperidone, each given in a single administration, was acceptable. No serious AEs occurred during treatment with the test formulation or the reference formulation. A total of 73 AEs were observed in 24 subjects during the study, and the event rate was similar with both formulations (37 AEs occurred after intake of the test formulation, while 36 AEs occurred after intake of the reference formulation). The most common AE was sedation (48 events), followed by nasal reactions (14 events), postural hypotension (3 events), hypertriglyceridemia (2 events), dizziness (4 events), nausea (1 events), and anorexia (1 events). Their severity was as follows: 16 were mild, 57 were moderate, and none were severe. The majority of the AEs were considered to be related (48 events) or probably related (23 events) to the study medication. No clinically significant abnormalities on physical examination, vital sign measurements, or electrocardiographic recordings were reported. 3.

Abbreviations: nac, nicotinic acid; ppbng, porphobilinogen; thm,

Abbreviations: nac, nicotinic acid; ppbng, porphobilinogen; thm, thiamin; pan4p, pantotheine-4-phosphate; dhor-S, S-dihydroorotate. Figure 3 Effect of different metabolites on the performance of the metabolic models. Biomass production

rates (mmol g DW-1 h-1) in the two networks (strain Bge, green bars; strain Pam, red bars) were measured Selleck Copanlisib under minimal conditions (see Fig. 2 and main text) or considering the uptake of different metabolites. FBA was also used to predict the behavior of the strain Bge in terms of growth rate when an additional metabolite was considered in the medium. We tested several metabolites with transport systems encoded by genes present in both B. cuenoti genomes (L-Asp, D-fructose, fumarate, L-Glu, glycerol, L-malate, succinate and urea) and also the input of the TCA cycle intermediate 2-oxoglutarate, as a simulation of an anaplerotic reaction. All

the considered additions had a positive effect on the biomass production rate by the Bge network, compared to the minimal medium (Fig. 3). In particular, some intermediates of the TCA cycle improved the performance of both networks with a remarkable ten-fold enhancement of biomass production by the Pam network in the presence of L-Glu and 2-oxoglutarate. This result can be correlated with the fact that the strain Pam possesses an incomplete TCA cycle. We decided to focus our attention on how the metabolic flux should be completely redirected click here through the different reactions leaving or entering this pathway (see Fig. 1). Thus, the fluxes through the transaminase reactions generating 2-oxoglutarate were particularly

important because they feed the enzymatic steps of the TCA cycle downstream of the isocitrate dehydrogenase reaction (Fig. 4). In summary, the positive effect of L-Glu (and 2-oxoglutarate) on the Pam network achieved a similar performance to the Bge network due to the anaplerotic effect of these metabolites (Fig. 4). Figure 4 Flux distribution through the TCA cycle and adjacent reactions. FBA simulations 17-DMAG (Alvespimycin) HCl of both models (strain Bge, left; strain Pam, right) were performed under minimal medium (green values) or with L-Glu uptake (red values). EC numbers are indicated (for enzyme names, see Fig. 1). The excretion of ammonia from the system, a phenomenon compatible with the physiological and experimental observations (for review see [8] and [1] and references therein), was always observed in simulations with both models under minimal conditions. The efflux of ammonia reached maximum levels when L-Glu uptake was simulated by the system. However, the efflux of ammonia stopped and could even be reversed when 2-oxoglutarate or succinate were provided to both metabolic networks. This was due to an increased assimilation of ammonia through displacement of the glutamate dehydrogenase reaction (EC 1.4.1.4) in the assimilative direction.

Mol Biol Cell 2008,19(12):5214–5225 PubMedCrossRef 3 Walther TC,

Mol Biol Cell 2008,19(12):5214–5225.PubMedCrossRef 3. Walther TC, Brickner JH, Aguilar PS, Bernales S, Pantoja C, Walter P: Eisosomes mark static sites of endocytosis. Nature 2006,439(7079):998–1003.PubMedCrossRef 4. Young ME, Karpova TS, Brugger B, Moschenross DM, Wang GK, Schneiter R, Wieland FT, Cooper JA: The Sur7p family defines novel cortical domains

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BC: Additional background research and paper sourcing for literat

BC: Additional background research and paper sourcing for literature review. RS: Image acquisition. Anonymised radiographic data. AH: Additional key source acquisition. Proof read and helped

edit paper. MB: Consultant surgeon responsible for overall patient care and patient data. Read and approved manuscript. All authors read and approved the final manuscript.”
“Introduction Intra-abdominal infections (IAIs) include a wide spectrum of pathological conditions, ranging from uncomplicated appendicitis to faecal peritonitis [1]. In the event of complicated IAI the infection proceeds beyond a singularly affected organ and causes either localized peritonitis (intra-abdominal abscesses) or diffuse peritonitis. Effectively treating patients with complicated intra-abdominal infections find more involves both source control and antimicrobial therapy [2, 3]. In order to describe the epidemiological, clinical, microbiological, and surgical treatment profiles of complicated intra-abdominal infections (IAIs) in Europe, the World Society of Emergency Surgery (WSES) designed the CIAO Study (Complicated intra-abdominal infections observational study). The CIAO Study was conducted during 2012 across twenty European countries [4]. Given the interesting results of the CIAO Study, WSES designed a prospective observational study investigating the management of complicated intra-abdominal

infections in a worldwide context. The CIAOW Selumetinib study (Complicated intra-abdominal infections worldwide observational study) is a multicenter observational study underwent in 68 medical institutions worldwide during a six-month study period (October 2012-March 2013). In January 2013 the preliminary results (2-month study period) of the CIAOW study were published [5]. WSES presents the definitive data of the CIAOW Study. Methods Aim The purpose of the P-type ATPase study was to describe the clinical, microbiological, and treatment profiles of both community- and healthcare-acquired complicated

IAIs in a worldwide context. Patients older than 18 years with both community-acquired and healthcare-associated IAIs were included in the database. Study population The CIAOW study is a multicenter observational study underwent in 68 medical institutions worldwide. The study included patients undergoing surgery or interventional drainage to address complicated IAIs. Medical institutions from each continent participated in the study. The geographical distribution of the participating centers are represented in Figure 1. Figure 1 Participating centers for each continent. Study design The study did not attempt to change or modify the laboratory or clinical practices of the participating physicians, and neither informed consent nor formal approval by an Ethics Committee were required. The study met the standards outlined in the Declaration of Helsinki and Good Epidemiological Practices.

Data indicate that treatment with

vitamin D could be bene

Data indicate that treatment with

vitamin D could be beneficial in reducing the risk of developing multiple sclerosis and diminishing its exacerbations [102]. Although contradictory ABT 888 data exist concerning supplementation benefits in rheumatoid arthritis (RA) and systemic lupus erythematosus, an association between low levels of 25(OH) vitamin D levels and activity of both diseases has been reported [103, 104]. Furthermore, an inverse association between higher intake of vitamin D and risk of rheumatoid arthritis was demonstrated in the Iowa Women’s Health Study [105]. However, we still lack non-biased large cohort studies that can sustain the proposed benefits of vitamin D supplementation for optimal immune function. Large-scale intervention trials in humans that support the findings in preclinical or observational studies are lacking [96]. Vitamin D and cancer treatment and prevention Many experimental data show that calcitriol stimulates apoptosis and differentiation and inhibits angiogenesis and proliferation in tumour cells [106]. Numerous association studies suggest that serum 25(OH) vitamin D levels are inversely associated with the risk of many types of cancer. Further, in some studies of patients with cancer, an association between low 25(OH) vitamin D levels and poor prognosis has been observed [107,

108]. A meta-analysis of available studies indicated that there is a trend for lower incidence of colorectal carcinoma and adenoma with 25(OH) vitamin D levels >20 ng/ml in a dose–response association [109]. For breast cancer, a pooled analysis of two studies Galeterone with 880 cases CDK and cancer and 880 controls demonstrated that individuals with sufficient serum 25(OH) vitamin D levels had 50% lower risk of breast cancer

than those with levels <13 ng/ml [110]. In addition, a large case–control study on 1,394 post-menopausal breast cancer patients and 1,365 controls also showed that the 25(OH) vitamin D level was significantly associated with lower breast cancer risk, particularly at levels above 20 ng/ml [111]. Most evidence concerning the link between vitamin D and cancer is derived from laboratory studies and observational investigations of 25(OH) vitamin D levels in association with cancer incidence and outcome. There are, however, several possible confounding factors and association cannot prove causation. Moreover, results from prospective studies only are more heterogeneous and do not support a significant association between vitamin D status and breast cancer [112]. There have been no clinical trials with cancer incidence or mortality as a primary outcome to support causality between vitamin D status and cancer. One population-based randomised clinical trial found that calcium plus vitamin D supplementation decreased cancer incidence as a secondary outcome. In that study including 1,179 healthy postmenopausal women aged >55 years, the mean level of 25(OH) vitamin D at baseline was 29 ng/ml.

CrossRefPubMed 6 Vautrin E, Genieys S, Charles S, Vavre F: Do ve

CrossRefPubMed 6. Vautrin E, Genieys S, Charles S, Vavre F: Do vertically transmitted symbionts co-existing in a single host compete Protein Tyrosine Kinase inhibitor or cooperate? A modelling approach. J Evol Biol 2008, 21:145–161.CrossRefPubMed 7. Lombardo M: Access to mutualistic endosymbiotic microbes: an underappreciated benefit of group living. Behav Ecol Sociobiol 2008, 62:479–497.CrossRef 8. Krause J, Ruxton GD: Living in groups New York, Oxford

University Press 2002. 9. Cremer S, Armitage SAO, Schmid-Hempel P: Social Immunity. Curr Biol 2007, 17:R693-R702.CrossRefPubMed 10. Degnan PH, Lazarus AB, Brock CD, Wernegreen JJ: Host-symbiont stability and fast evolutionary rates in an ant-bacterium association: Cospeciation of Camponotus species and their endosymbionts, Candidatus Blochmannia. Syst Biol 2004, 53:95–110.CrossRefPubMed 11. Gaudermann P, Vogl I, Zientz E, Silva FJ, Moya A, Gross R, Dandekar T: Analysis of and function predictions for previously conserved hypothetical or putative proteins in Blochmannia floridanus. Bmc Microbiol 2006, 6:1.CrossRefPubMed 12. Degnan PH, Lazarus AB, Wernegreen JJ: Genome sequence of Blochmannia pennsylvanicus

indicates parallel evolutionary trends among bacterial mutualists of insects. Genome Res 2005, 15:1023–1033.CrossRefPubMed 13. Gil R, Silva FJ, Zientz E, Delmotte F, Gonzalez-Candelas F, Latorre A, Rausell C, Kamerbeek J, Gadau J, Holldobler B, et al.: The genome sequence of Blochmannia floridanus : Comparative analysis of reduced genomes. Proc Natl Acad Sci USA 2003, 100:9388–9393.CrossRefPubMed 14. Zientz E, Beyaert N, Gross R, Feldhaar H: Relevance of the endosymbiosis of Blochmannia floridanus and carpenter Stem Cells inhibitor ants at different stages of the life cycle of the host. Appl Environ Microbiol 2006, 72:6027–6033.CrossRefPubMed 15. Wernegreen JJ, Degnan PH, Lazarus AB, Palacios

C, Bordenstein SR: Genome evolution in an insect cell: Distinct features of an ant-bacterial partnership. Biol Bull 2003, 204:221–231.CrossRefPubMed 16. Sauer C, Stackebrandt E, Gadau J, Holldobler B, Gross R: Systematic relationships and cospeciation of bacterial Sclareol endosymbionts and their carpenter ant host species: proposal of the new taxon Candidatus Blochmannia gen. nov. Int J Syst Evol Microbiol 2000, 50:1877–1886.PubMed 17. Moran NA: Symbiosis. Curr Biol 2006, 16:R866-R871.CrossRefPubMed 18. Oliver KM, Russell JA, Moran NA, Hunter MS: Facultative bacterial symbionts in aphids confer resistance to parasitic wasps. Proc Natl Acad Sci USA 2003, 100:1803–1807.CrossRefPubMed 19. Hedges LM, Brownlie JC, O’Neill SL, Johnson KN:Wolbachia and virus protection in insects. Science 2008, 322:702.CrossRefPubMed 20. Kaltenpoth M, Gottler W, Herzner G, Strohm E: Symbiotic bacteria protect wasp larvae from fungal infestation. Curr Biol 2005, 15:475–479.CrossRefPubMed 21. Wang Q, Garrity GM, Tiedje JM, Cole JR: Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy. Appl Environ Microbiol 2007, 73:5261–5267.CrossRefPubMed 22.

PNP also accumulates in the

PNP also accumulates in the FDA approved Drug Library soil due to the hydrolysis of organophosphorus insecticides such as parathion or methyl parathion (MP) [1]. Although PNP is less toxic than MP, it is also considered a significant potential toxic contaminant [2, 3] and belongs to one of 275 hazardous substances commonly found at Superfund sites [4, 5]. Many PNP-degrading bacteria have been isolated and their PNP degradation pathways studied [2, 6, 7]. In general, there are two alternative oxidative pathways that have been identified based

on their distinct intermediates. The hydroquinone (HQ) pathway, in which PNP is degraded via HQ, is the predominant pathway in gram-negative bacteria such as Moraxella sp. [2] and Pseudomonas sp. strain WBC-3 (Figure 1, upper)

[3]. The hydroxyquinol (BT) pathway is always used in gram-positive bacteria such as Bacillus sphaericus JS905 [7] and Rhodococcus opacus SAO101 [5]. PNP is degraded via 4-NC and BT in this pathway (Figure 1, lower). However, recently a gram negative organism, Burkholderia sp. strain SJ98, was reported to degrade PNP through the BT pathway, with no HQ pathway being detected [8]. In a gram positive organism, Rhodococcus sp. strain PN1, a two component PNP monooxygenase NpsA1A2 was found to catalyze PNP to both HQ and BT in the JQ1 presence of ascorbic acid as a reducing reagent. However, no microbial degradation data or results from direct enzyme analyses were provided [9]. We are not aware of any reports of one bacterium being able to degrade PNP utilizing two different pathways. Figure 1 Two alternative oxidative pathways

for the metabolism of PNP. Although some studies examining PNP degradation have been reported, genetic information related to the PNP degradation pathways remains limited. In the BT pathway, two enzymes were first characterized from Rhodococcus opacus SAO101: Palmatine one was the two-component PNP monooxygenase NpcAB; the other was the one-component BT 1,2-dioxygenase NpcC. However, the other enzymes involved in this pathway have not been identified [5]. In Arthrobacter sp. strain JS443, another two-component monooxygenase gene NpdA1A2 has been identified [4]. Recently, Chauhan A et al. identified two lower stream genes (pnpCD) encoding BT 1,2-dioxygenase and maleylacetate (MA) reductase in this pathway [8]. It is worth mentioning that there are two clusters involved in PNP degradation in the gram-positive bacterium Rhodococcus sp. strain PN1. Within these two clusters, two kinds of two-component PNP monooxygenase genes (nphA1A2 and npsA1A2), a regulator protein gene (npcR) and a BT 1,2-dioxygenase gene (npsB) have been identified [9, 10]. For the HQ pathway, the first gene cluster was obtained from Pseudomonas sp. strain WBC-3, and three enzymes involved in PNP degradation, PNP 4-monooxygenase (PnpA), p-benzoquinone (BQ) reductase (PnpB) and BT 1,2-dioxygenase (PnpG), have been characterized [3, 11].

Isolate IMAU20185 belonging to ST9 was a six-locus variant of ST1

Isolate IMAU20185 belonging to ST9 was a six-locus variant of ST1 to which it was connected by a dotted line. Isolate IMAU80137 belonging to ST19 was a six-locus variant of ST14 to which it was also connected by a dotted line. UPGMA tree based on MLST data Genetic relatedness amongst the L. lactis isolates investigated in this study showed they were well clustered within two major groups, A and B. Group A was comprised of 34 isolates and group B of only 16 isolates. Group A was the better supported

group and included two subgroups. Group B was a weakly supported group that included four subgroups (Figure  3). With the exception of ST19, isolates in group A were closely related only differing in CP 868596 two out of the eight loci from the primary founder, ST14. The isolate that belonged to ST19 was a six-locus variant of the primary founder. Isolates in Group B were distantly related and differed in between two and six of the eight loci from the primary founder ST1. Figure 3 UPGMA dendrogram showing the genetic relationships between the 20 STs that belong to L . lactis through Alectinib ic50 MLST typing in this study. The Phylogenetic tree was produced using START 2.0 software and the UPGMA method.

The numbering in the figure refers to the ST. Two major phylogroups were designated as A and B. Discussion MLST is considered to be the best method for studying molecular epidemiology and population structure of bacteria [29–31]. Although this approach has been developed for several LAB, such as Lb. plantarum, Lb. delbrueckii, Lb. casei, and O. oeni[25, 26, 32], until this study there had been no MLST protocol used for L. lactis. In this study, we used MLST with eight housekeeping genes on 50 L. lactis isolates from a relatively large geographic area including Mongolia, a number of Chinese Provinces and an Autonomous region. These representative isolates are unique in their diversity of sources and provide the relevant information required for a better understanding of genetic diversity, persistence and movement. The first step in development of a MLST typing

method required analysis of the sequence diversity of eight housekeeping genes from the 50 L. lactis isolates under evaluation, to ensure that the MLST protocol had Cobimetinib mouse the discriminatory power to type isolates within a single species. The two loci that had low polymorphism, contained three and four polymorphic sites in the recA and carB loci respectively (Table  1). The low level of biodiversity in recA and carB suggested they had similar sequences at the species level and would, therefore, have a lower discriminatory ability than the other housekeeping loci used in this study. The remaining six loci, groEL, pheS, uvrC, rpoB, pyrG, murC had more polymorphic sites (between five and nine), suggesting that they would have a good discriminatory ability when used in MLST. A total of 47 polymorphic sites were detected in the eight loci giving a polymorphism rate of 0.88% of the 5,325 nucleotides present.