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461700.9986A maximum likelihood QTL analysis reveals common genome regions controlling resistance to Salmonella colonization and carrier-state. BACKGROUND: The serovars Enteritidis and Typhimurium of the Gram-negative bacterium Salmonella enterica are significant causes of human food poisoning. Fowl carrying these bacteria often show no clinical disease, with detection only established post-mortem. Increased resistance to the carrier state in commercial poultry could be a way to improve food safety by reducing the spread of these bacteria in poultry flocks. Previous studies identified QTLs for both resistance to carrier state and resistance to Salmonella colonization in the same White Leghorn inbred lines. Until now, none of the QTLs identified was common to the two types of resistance. All these analyses were performed using the F2 inbred or backcross option of the QTLExpress software based on linear regression. In the present study, QTL analysis was achieved using Maximum Likelihood with QTLMap software, in order to test the effect of the QTL analysis method on QTL detection. We analyzed the same phenotypic and genotypic data as those used in previous studies, which were collected on 378 animals genotyped with 480 genome-wide SNP markers. To enrich these data, we added eleven SNP markers located within QTLs controlling resistance to colonization and we looked for potential candidate genes co-localizing with QTLs. RESULTS: In our case the QTL analysis method had an important impact on QTL detection. We were able to identify new genomic regions controlling resistance to carrier-state, in particular by testing the existence of two segregating QTLs. But some of the previously identified QTLs were not confirmed. Interestingly, two QTLs were detected on chromosomes 2 and 3, close to the locations of the major QTLs controlling resistance to colonization and to candidate genes involved in the immune response identified in other, independent studies. CONCLUSIONS: Due to the lack of stability of the QTLs detected, we suggest that interesting regions for further studies are those that were identified in several independent studies, which is the case of the QTL regions on chromosomes 2 and 3, involved in resistance to both Salmonella colonization and carrier state. These observations provide evidence of common genes controlling S. Typhimurium colonization and S. Enteritidis carrier-state in chickens.201222613937
516610.9985Illegitimate recombination: an efficient method for random mutagenesis in Mycobacterium avium subsp. hominissuis. BACKGROUND: The genus Mycobacterium (M.) comprises highly pathogenic bacteria such as M. tuberculosis as well as environmental opportunistic bacteria called non-tuberculous mycobacteria (NTM). While the incidence of tuberculosis is declining in the developed world, infection rates by NTM are increasing. NTM are ubiquitous and have been isolated from soil, natural water sources, tap water, biofilms, aerosols, dust and sawdust. Lung infections as well as lymphadenitis are most often caused by M. avium subsp. hominissuis (MAH), which is considered to be among the clinically most important NTM. Only few virulence genes from M. avium have been defined among other things due to difficulties in generating M. avium mutants. More efforts in developing new methods for mutagenesis of M. avium and identification of virulence-associated genes are therefore needed. RESULTS: We developed a random mutagenesis method based on illegitimate recombination and integration of a Hygromycin-resistance marker. Screening for mutations possibly affecting virulence was performed by monitoring of pH resistance, colony morphology, cytokine induction in infected macrophages and intracellular persistence. Out of 50 randomly chosen Hygromycin-resistant colonies, four revealed to be affected in virulence-related traits. The mutated genes were MAV_4334 (nitroreductase family protein), MAV_5106 (phosphoenolpyruvate carboxykinase), MAV_1778 (GTP-binding protein LepA) and MAV_3128 (lysyl-tRNA synthetase LysS). CONCLUSIONS: We established a random mutagenesis method for MAH that can be easily carried out and combined it with a set of phenotypic screening methods for the identification of virulence-associated mutants. By this method, four new MAH genes were identified that may be involved in virulence.201222966811
463320.9985Detection of Helicobacter pylori virulence-associated genes. Helicobacter pylori is an important human pathogen and persistent colonization of the human gastric mucosa can cause severe gastrointestinal diseases. The bacterium should not be considered as a uniform organism, but as a population of closely related and yet genetically diverse bacteria. Several genes of H. pylori (such as vacA and cagA) have been identified as being virulence-associated and may have important clinical and epidemiological implications. Assessment of virulence-associated genes of H. pylori should be included in clinical and epidemiological studies as well as therapeutic trials, in order to stratify between patient groups, harboring H. pylori strains with particular virulence genotypes. Molecular determination of antibiotic resistance will be especially useful for treatment studies. Together with our increasing knowledge about the human genome, typing of H. pylori will facilitate the management of gastroenterological pathologies.200111901834
255430.9985Development of an antibiotic resistance monitoring system in Hungary. Because of the rapid development and spread of antimicrobial resistance it is important that a system be established to monitor antimicrobial resistance in pathogenic zoonotic and commensal bacteria of animal origin. Susceptibility testing of bacteria from carcasses and different samples of animal origin has been carried out in veterinary institutes for a long time but by an inconsistent methodology. The disc diffusion method proposed by the National Committee for Clinical Laboratory Standards (NCCLS) was introduced in all institutes in 1997. In order to obtain a coherent view of the antimicrobial resistance of bacteria a computer system was consulted, consisting of a central computer to store all data and some local computers attached to it through the network. At these local measuring stations computers are connected to a video camera, which displays the picture of Petri dishes on the monitor, and inhibition zone diameters of bacteria can be drawn with the mouse by the inspector. The software measures the diameters, evaluates whether or not the bacteria are sensitive, and stores the data. The evaluation is based upon the data of the NCCLS. The central computer can be connected to as many local computers with measuring stations as we wish, so it is suitable for an integrated system for monitoring trends in antimicrobial resistance of bacteria from animals, food and humans, facilitating comparison of the occurrence of resistance for each circumstance in the chain. It depends on the examiners which antibiotics they want to examine. Thirty-two different antibiotic panels were compiled, taking into consideration the active ingredients of medicinal products permitted for veterinary use in Hungary, natural resistance and cross-resistance, the mechanism of resistance and the animal species, i.e. which drugs were recommended for treatment in the given animal species, and the recommendations of the OIE Expert Group on Antimicrobial Resistance. The members of the panels can be changed any time, even during the measuring process. In addition to the inhibition zone diameters of bacteria the database also includes information about bacterial and animal species, the age of animals and the sample or organ where the bacteria are from. Since January 2001 the antibiotic susceptibility of E. coli, Salmonella, Campylobacter and Enterococcus strains isolated from the colons of slaughter cows, pigs and broiler chickens has also been examined. Each of the 19 counties of Hungary submits to the laboratory three tied colon samples from a herd of the above-mentioned animals every month.200212113174
478040.9984Characterization of two virulent Proteus mirabilis phages and application of phage cocktail in raw chicken safety. Proteus mirabilis is an opportunistic pathogen belonging to the genus Proteus in the Morganellaceae. It is closely associated with infections of the gastrointestinal and urinary systems in humans and animals, posing a threat to the safety of food industries. With the increasing abuse of antibiotics, the problem of bacterial resistance has become extremely urgent. Moreover, animal-source foods often have drug residues, making it an urgent need to explore alternatives to antibiotics. Bacteriophages, bacteria-lysing viruses, cause no biological pollution and do not induce bacterial antibiotic resistance, making them potential green biological bactericides. In this study, 24 pathogenic P. mirabilis strains were isolated as host bacteria. Two virulent bacteriophages, Proteus phage ABTNL-P14 (referred to as P14) and Proteus phage ABTNL-Pp2 (referred to as Pp2), were isolated and identified, both showing effective infectivity against P. mirabilis. Two phages were confirmed as virulent strains lacking virulence factors, lysogeny-related genes and antibiotic resistance genes, meeting safety criteria for food applications. This study also evaluated the effectiveness of a P14 and Pp2 phage cocktail in eliminating P. mirabilis from chicken breast meat at 4 °C under MOIs of 1, 100, and 10,000. Notably, at a high MOI (10,000), the phage cocktail eliminated two host strains on chicken breast meat surfaces within 1 h and 6 h. Therefore, the phage cocktail has the potential to eliminate P. mirabilis in foods at low temperatures, serving as a green antibacterial agent for food applications.202641136136
254050.9984Equine sinusitis aetiology is linked to sinus microbiome by amplicon sequencing. BACKGROUND: Information regarding the microbiome in sinusitis using genetic sequencing is lacking and more-in-depth understanding of the microbiome could improve antimicrobial selection and treatment outcomes for cases of primary sinusitis. OBJECTIVES: To describe sinus microbiota in samples from horses with sinusitis and compare microbiota and the presence of antimicrobial resistance genes between primary, dental-related and other secondary causes of sinusitis. STUDY DESIGN: Retrospective case series. METHODS: Records of equine sinusitis from 2017 to 2021 were reviewed and historical microbial amplicon sequence data were obtained from clinical diagnostic testing of sinus secretions. Following bioinformatic processing of bacterial and fungal sequence data, the sinus microbiota and importance of sinusitis aetiology among other factors were investigated from the perspectives of alpha diversity (e.g., number of operational taxonomic units [OTUs], Hill1 Diversity), beta diversity, and differentially abundant taxa. Quantitative PCR allowed for comparisons of estimated bacterial abundance and detection rate of common antibiotic resistance-associated genes. In a smaller subset, longitudinal analysis was performed to evaluate similarity in samples over time. RESULTS: Of 81 samples analysed from 70 horses, the bacterial microbiome was characterised in 66, and fungal in five. Only sinusitis aetiology was shown to significantly influence microbiome diversity and composition (p < 0.05). Dental-related sinusitis (n = 44) was associated with a significantly higher proportion of obligate anaerobic bacteria, whereas primary sinusitis (n = 12) and other (n = 10) groups were associated with fewer bacteria and higher proportions of facultative anaerobic and aerobic genera. Antimicrobial resistance genes and fungal components were exclusively identified in dental-related sinusitis. MAIN LIMITATIONS: Retrospective nature, incomplete prior antimicrobial administration data. CONCLUSIONS: Molecular characterisation in sinusitis identifies microbial species which may be difficult to isolate via culture, and microbiome profiling can differentiate sinusitis aetiology, which may inform further treatment, including antimicrobial therapy.202336199163
568660.9984Founding of the culture collection of antibiotic-resistant strains of zoonotic bacteria in the Russian Federation. BACKGROUND AND AIM: The main purpose of a national bioresource center is to standardize, centralize, preserve, and ensure accessibility of microbial bioresources that accumulate there because of state research programs. The establishment of national bioresource centers for antibiotic-resistant microorganisms allows to solve practical problems in the field of veterinary service, as well as to develop effective chemotherapeutic and disinfectant drugs to overcome the mechanisms of resistance. This study aimed to outline the process of forming a national culture collection of antibiotic-resistant strains of zoonotic bacteria in the Russian Federation using two microbial strains. MATERIALS AND METHODS: The object of research was isolates of Salmonella spp., Escherichia coli, Enterococcus spp., Campylobacter spp., Listeria monocytogenes, and Staphylococcus spp., all of which were obtained from biomaterials of farm animals, feed samples, bedding, water from livestock buildings, washouts from environmental objects, and food products. The resistance of bacterial isolates was determined using microbiological and molecular-genetic research methods. RESULTS: During monitoring studies, 1489 bacterial isolates were isolated. In total, 408 bacterial isolates were tested for sensitivity to antimicrobial agents, including E. coli (47.6%), Salmonella spp. (30.4%), Enterococcus spp. (11.3%), and Campylobacter spp. (10.8%). For genetic characterization, 95 isolates of Salmonella enterica, E. coli, Campylobacter spp., L. monocytogenes, Staphylococcus spp., Enterococcus spp. were chosen from the research collection, which was formed as part of the monitoring program for antibiotic resistance. CONCLUSION: Deposited isolates that underwent whole-genome analysis can be used as positive control samples both in the development and use of methods or test systems for the detection of various resistance genes in zoonotic bacteria. In addition, such isolates can also be used for microbiological studies related to determining the sensitivity of microorganisms to antibacterial drugs, for phenotypic studies in the diagnosis of various bacterial infections in animals and birds, and retrospective analysis of strains from numerous collections.202337621551
475970.9984Recent advances in rapid antimicrobial susceptibility testing systems. INTRODUCTION: Until recently antimicrobial susceptibility testing (AST) methods based on the demonstration of phenotypic susceptibility in 16-24 h remained largely unchanged. AREAS COVERED: Advances in rapid phenotypic and molecular-based AST systems. EXPERT OPINION: AST has changed over the past decade, with many rapid phenotypic and molecular methods developed to demonstrate phenotypic or genotypic resistance, or biochemical markers of resistance such as β-lactamases associated with carbapenem resistance. Most methods still require isolation of bacteria from specimens before both legacy and newer methods can be used. Bacterial identification by MALDI-TOF mass spectroscopy is now widely used and is often key to the interpretation of rapid AST results. Several PCR arrays are available to detect the most frequent pathogens associated with bloodstream infections and their major antimicrobial resistance genes. Many advances in whole-genome sequencing of bacteria and fungi isolated by culture as well as directly from clinical specimens have been made but are not yet widely available. High cost and limited throughput are the major obstacles to uptake of rapid methods, but targeted use, continued development and decreasing costs are expected to result in more extensive use of these increasingly useful methods.202133926351
508780.9983Sensitive colorimetric detection of antibiotic resistant Staphylococcus aureus on dairy farms using LAMP with pH-responsive polydiacetylene. Rapidly and accurately detecting antibiotic-resistant pathogens in agriculture and husbandry is important since these represent a major threat to public health. While much attention has been dedicated to detecting now-common resistant bacteria, such as methicillin-resistant Staphylococcus aureus, fewer methods have been developed to assess resistance against macrolides in Staphylococcus aureus (SA). Here, we report a visual on-site detection system for macrolide resistant SA in dairy products. First, metagenomic sequencing in raw milk, cow manure, water and aerosol deposit collected from dairy farms around Tianjin was used to identify the most abundant macrolide resistance gene, which was found to be the macB gene. In parallel, SA housekeeping genes were screened to allow selective identification of SA, which resulted in the selection of the SAOUHSC_01275 gene. Next, LAMP assays targeting the above-mentioned genes were developed and interpreted by agarose gel electrophoresis. For on-site application, different pH-sensitive colorimetric LAMP indicators were compared, which resulted in selection of polydiacetylene (PDA) as the most sensitive candidate. Additionally, a semi-quantitative detection could be realized by analyzing the RGB information via smartphone with a LOD of 1.344 × 10(-7) ng/μL of genomic DNA from a milk sample. Finally, the proposed method was successfully carried out at a real farm within 1 h from sample to result by using freeze-dried reagents and portable devices. This is the first instance in which PDA is used to detect LAMP products, and this generic read-out system can be expanded to other antibiotic resistant genes and bacteria.202336327562
417690.9983Plants with Antimicrobial Activity Growing in Italy: A Pathogen-Driven Systematic Review for Green Veterinary Pharmacology Applications. Drug resistance threatening humans may be linked with antimicrobial and anthelmintic resistance in other species, especially among farm animals and, more in general, in the entire environment. From this perspective, Green Veterinary Pharmacology was proven successful for the control of parasites in small ruminants and for the control of other pests such as varroa in bee farming. As in anthelmintic resistance, antimicrobial resistance (AMR) represents one of the major challenges against the successful treatment of infectious diseases, and antimicrobials use in agriculture contributes to the spread of more AMR bacterial phenotypes, genes, and proteins. With this systematic review, we list Italian plants with documented antimicrobial activity against possible pathogenic microbes. Methods: The literature search included all the manuscripts published since 1990 in PubMed, Web of Science, and Scopus using the keywords (i) "antimicrobial, plants, Italy"; (ii) "antibacterial, plant, Italy"; (iii) "essential oil, antibacterial, Italy"; (iv) "essential oil, antimicrobial, Italy"; (v) "methanol extract, antibacterial, Italy"; (vi) "methanol extract, antimicrobial, Italy". Results: In total, 105 manuscripts that documented the inhibitory effect of plants growing in Italy against bacteria were included. One hundred thirty-five plants were recorded as effective against Gram+ bacteria, and 88 against Gram-. This will provide a ready-to-use comprehensive tool to be further tested against the indicated list of pathogens and will suggest new alternative strategies against bacterial pathogens to be employed in Green Veterinary Pharmacology applications.202235884173
4629100.9983Screening and in silico characterization of prophages in Helicobacter pylori clinical strains. The increase of antibiotic resistance calls for alternatives to control Helicobacter pylori, a Gram-negative bacterium associated with various gastric diseases. Bacteriophages (phages) can be highly effective in the treatment of pathogenic bacteria. Here, we developed a method to identify prophages in H. pylori genomes aiming at their future use in therapy. A polymerase chain reaction (PCR)-based technique tested five primer pairs on 74 clinical H. pylori strains. After the PCR screening, 14 strains most likely to carry prophages were fully sequenced. After that, a more holistic approach was taken by studying the complete genome of the strains. This study allowed us to identify 12 intact prophage sequences, which were then characterized concerning their morphology, virulence, and antibiotic-resistance genes. To understand the variability of prophages, a phylogenetic analysis using the sequences of all H. pylori phages reported to date was performed. Overall, we increased the efficiency of identifying complete prophages to 54.1 %. Genes with homology to potential virulence factors were identified in some new prophages. Phylogenetic analysis revealed a close relationship among H. pylori-phages, although there are phages with different geographical origins. This study provides a deeper understanding of H. pylori-phages, providing valuable insights into their potential use in therapy.202539368610
2555110.9983Results of the antimicrobial agent susceptibility study raised in a representative, cross-sectional monitoring study on a national basis. The use of antimicrobial substances in human and veterinary medicine inevitably results in a selection pressure for drug resistance in exposed bacteria. Preventive measures, apt to avoid the consequent development of new resistances and selection for existing ones, respectively, have to be elaborated. Moreover, it has to be ensured that neither resistant bacteria nor resistance genes are spread to and consequently via the food chain. Respiratory diseases as well as mastitis in dairy cattle belong to the most frequently occurring diseases in food-producing animals. For the first time in Germany, a comprehensive, cross-sectional study into the antimicrobial susceptibility of bacteria associated with these disease patterns in food-producing animals was conducted by the Federal Office of Consumer Protection and Food Safety (BVL) in 2001. The selection of examined bacterial species comprised Pasteurella multocida and Mannheimia haemolytica associated with respiratory disease in pigs, and Escherichia coli, Streptococcus spp. and Staphylococcus spp. causing mastitis in dairy cattle. Bacterial strains were collected following a representative sampling scheme, taking into account the total number of animals in the individual German federal Länder. In an analogous study conducted in 2002/2003, this selection was extended by the indication respiratory disease in juvenile cattle, caused by P. multocida and M. haemolytica, respectively. In comparison with data from 2001, MIC values determined in 2002/2003 suggested significantly lower or higher degrees of drug susceptibility only for a few antimicrobial agents. Comparison was carried out on the basis of bacterial species and individual federal Länder, respectively. Overall, the data raised in both studies revealed substantially lower resistance rates than published for Germany so far. This is particularly true for results from those Länder, whose animal health services had implemented preventive strategies to control infectious diseases. No correlation could be established between differing animal population densities and differences in the prevalence of resistance in corresponding Lander. However, the geographical distribution of occurrence of resistance against beta-lactam antimicrobial agents suggests different therapeutic strategies employed in different sized animal flocks. In federal Länder marked by large-scale livestock farming, significantly higher resistance values could be measured for cephalosporins than for penicillins, whereas in Länder with rather traditional farming structures, resistance to penicillins was predominant. Assuming otherwise similar factors of influence on the emergence of resistance, this pattern suggests that cephalosporins are preferably used in large enterprises and penicillins in smaller farms, respectively. Currently, mechanisms effecting changes in antimicrobial resistance are being further investigated in a successive study.200616531116
4618120.9983Genetic control of resistance to salmonellosis and to Salmonella carrier-state in fowl: a review. Salmonellosis is a frequent disease in poultry stocks, caused by several serotypes of the bacterial species Salmonella enterica and sometimes transmitted to humans through the consumption of contaminated meat or eggs. Symptom-free carriers of the bacteria contribute greatly to the propagation of the disease in poultry stocks. So far, several candidate genes and quantitative trait loci (QTL) for resistance to carrier state or to acute disease have been identified using artificial infection of S. enterica serovar Enteritidis or S. enterica serovar Typhimurium strains in diverse genetic backgrounds, with several different infection procedures and phenotypic assessment protocols. This diversity in experimental conditions has led to a complex sum of results, but allows a more complete description of the disease. Comparisons among studies show that genes controlling resistance to Salmonella differ according to the chicken line studied, the trait assessed and the chicken's age. The loci identified are located on 25 of the 38 chicken autosomal chromosomes. Some of these loci are clustered in several genomic regions, indicating the possibility of a common genetic control for different models. In particular, the genomic regions carrying the candidate genes TLR4 and SLC11A1, the Major Histocompatibility Complex (MHC) and the QTL SAL1 are interesting for more in-depth studies. This article reviews the main Salmonella infection models and chicken lines studied under a historical perspective and then the candidate genes and QTL identified so far.201020429884
4180130.9983Toward integrative genomics study of genetic resistance to Salmonella and Campylobacter intestinal colonization in fowl. Salmonella enterica serotypes Enteritidis and Typhimurium and Campylobacter jejuni are responsible for most cases of food poisoning in Europe. These bacteria do not cause severe disease symptoms in chicken, but they are easily propagated by symptomless chicken carriers which cannot be easily isolated. This animal tolerance is detrimental to food safety. In this particular case, increasing animal's resistance is not sufficient, since some animals considered as resistant are able to carry bacteria during several weeks without displaying disease symptoms. We review studies aimed at evaluating the resistance of chicken to Salmonella and Campylobacter intestinal colonization, either a few days or several weeks after infection. While studies of the genetic control of Campylobacter colonization are only beginning, mostly due to technical difficulties in infection protocols, genetic studies of Salmonella colonization have been conducted for now more than 20 years. They have initially reported an estimation of the genetic parameters associated with resistance to Salmonella colonization and are now aimed at identifying the genomic regions controlling variation of this trait in experimental lines and commercial populations. With the advent of high-throughput genomics, we are closer than ever to identify the true genes controlling resistance to Enterobacteria colonization in chicken. The comparison of genes involved in early resistance to intestinal colonization with genes controlling resistance to bacteria persistence several weeks after infection (i.e., carrier-state) should soon highlight the differences between the molecular mechanisms underlying those two distinct phenotypes. It will also be highly interesting to compare the genes or genomic regions controlling Campylobacter and Salmonella, in order to evaluate the feasibility of a selection conducted on both bacteria simultaneously.201223412643
6280140.9983Genomic variation in Pseudomonas aeruginosa clinical respiratory isolates with de novo resistance to a bacteriophage cocktail. Pseudomonas aeruginosa is an opportunistic pathogen that can cause sinus infections and pneumonia in cystic fibrosis (CF) patients. Bacteriophage therapy is being investigated as a treatment for antibiotic-resistant P. aeruginosa infections. Although virulent bacteriophages have shown promise in treating P. aeruginosa infections, the development of bacteriophage-insensitive mutants (BIMs) in the presence of bacteriophages has been described. The aim of this study was to examine the genetic changes associated with the BIM phenotype. Biofilms of three genetically distinct P. aeruginosa strains, including PAO1 (ATCC 15692), and two clinical respiratory isolates (one CF and one non-CF) were grown for 7 days and treated with either a cocktail of four bacteriophages or a vehicle control for 7 consecutive days. BIMs isolated from the biofilms were detected by streak assays, and resistance to the phage cocktail was confirmed using spot test assays. Comparison of whole genome sequencing between the recovered BIMs and their respective vehicle control-treated phage-sensitive isolates revealed structural variants in two strains, and several small variants in all three strains. These variations involved a TonB-dependent outer membrane receptor in one strain, and mutations in lipopolysaccharide synthesis genes in two strains. Prophage deletion and induction were also noted in two strains, as well as mutations in several genes associated with virulence factors. Mutations in genes involved in susceptibility to conventional antibiotics were also identified in BIMs, with both decreased and increased antibiotic sensitivity to various antibiotics being observed. These findings may have implications for future applications of lytic phage therapy.IMPORTANCELytic bacteriophages are viruses that infect and kill bacteria and can be used to treat difficult-to-treat bacterial infections, including biofilm-associated infections and multidrug-resistant bacteria. Pseudomonas aeruginosa is a bacterium that can cause life-threatening infections. Lytic bacteriophage therapy has been trialed in the treatment of P. aeruginosa infections; however, sometimes bacteria develop resistance to the bacteriophages. This study sheds light on the genetic mechanisms of such resistance, and how this might be harnessed to restore the sensitivity of multidrug-resistant P. aeruginosa to conventional antibiotics.202540162801
4388150.9983Detection of Genes Related to Antibiotic Resistance in Leptospira. Leptospirosis is a disease caused by the bacteria of the Leptospira genus, which can usually be acquired by humans through contact with urine from infected animals; it is also possible for this urine to contaminate soils and bodies of water. The disease can have deadly consequences in some extreme cases. Fortunately, until now, patients with leptospirosis have responded adequately to treatment with doxycycline and azithromycin, and no cases of antibiotic resistance have been reported. However, with the extensive use of such medications, more bacteria, such as Staphylococci and Enterococci, are becoming resistant. The purpose of this study is to determine the presence of genes related to antibiotic resistance in the Leptospira genus using bioinformatic tools, which have not been undertaken in the past. Whole genomes from the 69 described Leptospira species were downloaded from NCBI's GeneBank and analyzed using CARD (The Comprehensive Antibiotic Resistant Database) and RAST (Rapid Annotations using Subsystem Technology). After a detailed genomic search, 12 genes associated with four mechanisms were found: resistance to beta-lactamases, vancomycin, aminoglycoside adenylyltransferases, as well as multiple drug efflux pumps. Some of these genes are highly polymorphic among different species, and some of them are present in multiple copies in the same species. In conclusion, this study provides evidence of the presence of genes related to antibiotic resistance in the genomes of some species of the genus Leptospira, and it is the starting point for future experimental evaluation to determine whether these genes are transcriptionally active in some species and serovars.202439330892
3945160.9983Vancomycin-resistant enterococci: why are they here, and where do they come from? Vancomcyin-resistant enterococci (VRE) have emerged as nosocomial pathogens in the past 10 years, causing epidemiological controversy. In the USA, colonisation with VRE is endemic in many hospitals and increasingly causes infection, but colonisation is absent in healthy people. In Europe, outbreaks still happen sporadically, usually with few serious infections, but colonisation seems to be endemic in healthy people and farm animals. Vancomycin use has been much higher in the USA, where emergence of ampicillin-resistant enterococci preceded emergence of VRE, making them very susceptible to the selective effects of antibiotics. In Europe, avoparcin, a vancomycin-like glycopeptide, has been widely used in the agricultural industry, explaining the community reservoir in European animals. Avoparcin has not been used in the USA, which is consistent with the absence of colonisation in healthy people. From the European animal reservoir, VRE and resistance genes have spread to healthy human beings and hospitalised patients. However, certain genogroups of enterococci in both continents seem to be more capable of causing hospital outbreaks, perhaps because of the presence of a specific virulence factor, the variant esp gene. By contrast with the evidence of a direct link between European animal and human reservoirs, the origin of American resistance genes remains to be established. Considering the spread of antibiotic-resistant bacteria and resistance genes, the emergence of VRE has emphasised the non-existence of boundaries between hospitals, between people and animals, between countries, and probably between continents.200111871804
4776170.9983Integrate genome-based assessment of safety for probiotic strains: Bacillus coagulans GBI-30, 6086 as a case study. Probiotics are microorganisms that confer beneficial effects on the host; nevertheless, before being allowed for human consumption, their safety must be verified with accurate protocols. In the genomic era, such procedures should take into account the genomic-based approaches. This study aims at assessing the safety traits of Bacillus coagulans GBI-30, 6086 integrating the most updated genomics-based procedures and conventional phenotypic assays. Special attention was paid to putative virulence factors (VF), antibiotic resistance (AR) genes and genes encoding enzymes responsible for harmful metabolites (i.e. biogenic amines, BAs). This probiotic strain was phenotypically resistant to streptomycin and kanamycin, although the genome analysis suggested that the AR-related genes were not easily transferrable to other bacteria, and no other genes with potential safety risks, such as those related to VF or BA production, were retrieved. Furthermore, no unstable elements that could potentially lead to genomic rearrangements were detected. Moreover, a workflow is proposed to allow the proper taxonomic identification of a microbial strain and the accurate evaluation of risk-related gene traits, combining whole genome sequencing analysis with updated bioinformatics tools and standard phenotypic assays. The workflow presented can be generalized as a guideline for the safety investigation of novel probiotic strains to help stakeholders (from scientists to manufacturers and consumers) to meet regulatory requirements and avoid misleading information.201626952108
4181180.9983The place of molecular genetic methods in the diagnostics of human pathogenic anaerobic bacteria. A minireview. Anaerobic infections are common and can cause diseases associated with severe morbidity, but are easily overlooked in clinical settings. Both the relatively small number of infections due to exogenous anaerobes and the much larger number of infections involving anaerobic species that are originally members of the normal flora, may lead to a life-threatening situation unless appropriate treatment is instituted. Special laboratory procedures are needed for the isolation, identification and susceptibility testing of this diverse group of bacteria. Since many anaerobes grow more slowly than the facultative or aerobic bacteria, and particularly since clinical specimens yielding anaerobic bacteria commonly contain several organisms and often very complex mixtures of aerobic and anaerobic bacteria, considerable time may elapse before the laboratory is able to provide a final report. Species definition based on phenotypic features is often time-consuming and is not always easy to carry out. Molecular genetic methods may help in the everyday clinical microbiological practice in laboratories dealing with the diagnostics of anaerobic infections. Methods have been introduced for species diagnostics, such as 16S rRNA PCR-RFLP profile determination, which can help to distinguish species of Bacteroides, Prevotella, Actinomyces, etc. that are otherwise difficult to differentiate. The use of DNA-DNA hybridization and the sequencing of special regions of the 16S rRNA have revealed fundamental taxonomic changes among anaerobic bacteria. Some anaerobic bacteria are extremely slow growing or not cultivatable at all. To detect them in special infections involving flora changes due to oral malignancy or periodontitis, for instance, a PCR-based hybridization technique is used. Molecular methods have demonstrated the spread of specific resistance genes among the most important anaerobic bacteria, the members of the Bacteroides genus. Their detection and investigation of the IS elements involved in their expression may facilitate following of the spread of antibiotic resistance among anaerobic bacteria involved in infections and in the normal flora members. Molecular methods (a search for toxin genes and ribotyping) may promote a better understanding of the pathogenic features of some anaerobic infections, such as the nosocomial diarrhoea caused by C. difficile and its spread in the hospital environment and the community. The investigation of toxin production at a molecular level helps in the detection of new toxin types. This mini-review surveys some of the results obtained by our group and others using molecular genetic methods in anaerobic diagnostics.200616956128
4634190.9983Genome analysis reveals a biased distribution of virulence and antibiotic resistance genes in the genus Enterococcus and an abundance of safe species. Enterococci are lactic acid bacteria (LAB) that, as their name implies, often are found in the gastrointestinal tract of animals. Like many other gut-dwelling LAB, for example, various lactobacilli, they are frequently found in other niches as well, including plants and fermented foods from all over the world. In fermented foods, they contribute to flavor and other organoleptic properties, help extend shelf life, and some even possess probiotic properties. There are many positive attributes of enterococci; however, they have been overshadowed by the occurrence of antibiotic-resistant and virulent strains, often reported for the two species, Enterococcus faecalis and Enterococcus faecium. More than 40,000 whole-genome sequences covering 64 Enterococcus type species are currently available in the National Center for Biotechnology Information repository. Closer inspection of these sequences revealed that most represent the two gut-dwelling species E. faecalis and E. faecium. The remaining 62 species, many of which have been isolated from plants, are thus quite underrepresented. Of the latter species, we found that most carried no potential virulence and antibiotic resistance genes, an observation that is aligned with these species predominately occupying other niches. Thus, the culprits found in the Enterococcus genus mainly belong to E. faecalis, and a biased characterization has resulted in the opinion that enterococci do not belong in food. Since enterococci possess many industrially desirable traits and frequently are found in other niches besides the gut of animals, we suggest that their use as food fermentation microorganisms is reconsidered.IMPORTANCEWe have retrieved a large number of Enterococcus genome sequences from the National Center for Biotechnology Information repository and have scrutinized these for the presence of virulence and antibiotic resistance genes. Our results show that such genes are prevalently found in the two species Enterococcus faecalis and Enterococcus faecium. Most other species do not harbor any virulence and antibiotic resistance genes and display great potential for use as food fermentation microorganisms or as probiotics. The study contributes to the current debate on enterococci and goes against the mainstream perception of enterococci as potentially dangerous microorganisms that should not be associated with food and health.202540202320