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453400.9970Microbiome diversity in Diaphorina citri populations from Kenya and Tanzania shows links to China. The Asian citrus psyllid (Diaphorina citri) is a key pest of Citrus spp. worldwide, as it acts as a vector for "Candidatus Liberibacter asiaticus (Las)", the bacterial pathogen associated with the destructive Huanglongbing (HLB) disease. Recent detection of D. citri in Africa and reports of Las-associated HLB in Ethiopia suggest that the citrus industry on the continent is under imminent threat. Endosymbionts and gut bacteria play key roles in the biology of arthropods, especially with regards to vector-pathogen interactions and resistance to antibiotics. Thus, we aim to profile the bacterial genera and to identify antibiotic resistance genes within the microbiome of different populations worldwide of D. citri. The metagenome of D. citri was sequenced using the Oxford Nanopore full-length 16S metagenomics protocol, and the "What's in my pot" (WIMP) analysis pipeline. Microbial diversity within and between D. citri populations was assessed, and antibiotic resistance genes were identified using the WIMP-ARMA workflow. The most abundant genera were key endosymbionts of D. citri ("Candidatus Carsonella", "Candidatus Profftella", and Wolbachia). The Shannon diversity index showed that D. citri from Tanzania had the highest diversity of bacterial genera (1.92), and D. citri from China had the lowest (1.34). The Bray-Curtis dissimilarity showed that China and Kenya represented the most diverged populations, while the populations from Kenya and Tanzania were the least diverged. The WIMP-ARMA analyses generated 48 CARD genes from 13 bacterial species in each of the populations. Spectinomycin resistance genes were the most frequently found, with an average of 65.98% in all the populations. These findings add to the knowledge on the diversity of the African D. citri populations and the probable introduction source of the psyllid in these African countries.202032589643
491810.9968Time trends for bacterial species and resistance patterns in semen in patients undergoing evaluation for male infertility. Semen from asymptomatic men who are being evaluated as male partners in interfile couples have been reported to contain a variety of bacteria. Longitudinal studies of the variation of these bacteria over time and their resistance patterns have not been commonly reported. At our institution, residues from semen samples are routinely evaluated for bacteria, including antibiotic sensitivity profiles. We set out to profile the changes in semen bacteria and antibiotic resistance at our institution over time. A total of 72 semen isolates were examined for type of bacteria and sensitivity to a panel of antibiotics. The results were divided into two separate 5-year intervals (the first beginning in 2006, the second in 2011) and compared. The majority of bacteria were skin flora, with Streptococcus and Staphylococcus being the most prevalent. The resistance data for these two pathogens showed minimal statistically significant difference between the two time periods, although the Staphylococcus species did show a trend toward increasing resistance, suggesting that antibiotics currently used in sperm cell preparations may need to be varied.201829706808
341520.9968Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches. The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces.201526114765
354030.9968Microbial contents of vacuum cleaner bag dust and emitted bioaerosols and their implications for human exposure indoors. Vacuum cleaners can release large concentrations of particles, both in their exhaust air and from resuspension of settled dust. However, the size, variability, and microbial diversity of these emissions are unknown, despite evidence to suggest they may contribute to allergic responses and infection transmission indoors. This study aimed to evaluate bioaerosol emission from various vacuum cleaners. We sampled the air in an experimental flow tunnel where vacuum cleaners were run, and their airborne emissions were sampled with closed-face cassettes. Dust samples were also collected from the dust bag. Total bacteria, total archaea, Penicillium/Aspergillus, and total Clostridium cluster 1 were quantified with specific quantitative PCR protocols, and emission rates were calculated. Clostridium botulinum and antibiotic resistance genes were detected in each sample using endpoint PCR. Bacterial diversity was also analyzed using denaturing gradient gel electrophoresis (DGGE), image analysis, and band sequencing. We demonstrated that emission of bacteria and molds (Penicillium/Aspergillus) can reach values as high as 1E5 cell equivalents/min and that those emissions are not related to each other. The bag dust bacterial and mold content was also consistent across the vacuums we assessed, reaching up to 1E7 bacterial or mold cell equivalents/g. Antibiotic resistance genes were detected in several samples. No archaea or C. botulinum was detected in any air samples. Diversity analyses showed that most bacteria are from human sources, in keeping with other recent results. These results highlight the potential capability of vacuum cleaners to disseminate appreciable quantities of molds and human-associated bacteria indoors and their role as a source of exposure to bioaerosols.201323934489
581940.9967Application of mNGS in the Etiological Analysis of Lower Respiratory Tract Infections and the Prediction of Drug Resistance. Lower respiratory tract infections (LRTIs) have high morbidity and mortality rates. However, traditional etiological detection methods have not been able to meet the needs for the clinical diagnosis and prognosis of LRTIs. The rapid development of metagenomic next-generation sequencing (mNGS) provides new insights for the diagnosis and treatment of LRTIs; however, little is known about how to interpret the application of mNGS results in LRTIs. In this study, lower respiratory tract specimens from 46 patients with suspected LRTIs were tested simultaneously using conventional microbiological detection methods and mNGS. Receiver operating characteristic (ROC) curves were used to evaluate the performance of the logarithm of reads per kilobase per million mapped reads [lg(RPKM)], genomic coverage, and relative abundance of the organism in predicting the true-positive pathogenic bacteria. True-positive viruses were identified according to the lg(RPKM) threshold of bacteria. We also evaluated the ability to predict drug resistance genes using mNGS. Compared to that using conventional detection methods, the false-positive detection rate of pathogenic bacteria was significantly higher using mNGS. It was concluded from the ROC curves that the lg(RPKM) and genomic coverage contributed to the identification of pathogenic bacteria, with the performance of lg(RPKM) being the best (area under the curve [AUC] = 0.99). The corresponding lg(RPKM) threshold for identifying the pathogenic bacteria was -1.35. Thirty-five strains of true-positive virus were identified based on the lg(RPKM) threshold of bacteria, with the detection of human gammaherpesvirus 4 being the highest and prone to coinfection with Pseudomonas aeruginosa, Acinetobacter baumannii, and Stenotrophomonas maltophilia. Antimicrobial susceptibility tests (AST) revealed the resistance of bacteria containing drug resistance genes (detected by mNGS). However, the drug resistance genes of some multidrug-resistant bacteria were not detected. As an emerging technology, mNGS has shown many advantages for the unbiased etiological detection and the prediction of antibiotic resistance. However, a correct understanding of mNGS results is a prerequisite for its clinical application, especially for LRTIs. IMPORTANCE LRTIs are caused by hundreds of pathogens, and they have become a great threat to human health due to the limitations of traditional etiological detection methods. As an unbiased approach to detect pathogens, mNGS overcomes such etiological diagnostic challenges. However, there is no unified standard on how to use mNGS indicators (the sequencing reads, genomic coverage, and relative abundance of each organism) to distinguish between pathogens and colonizing microorganisms or contaminant microorganisms. Here, we selected the mNGS indicator with the best identification performance and established a cutoff value for the identification of pathogens in LRTIs using ROC curves. In addition, we also evaluated the accuracy of antibiotic resistance prediction using mNGS.202235171007
461450.9967Listeria monocytogenes ability to survive desiccation: Influence of serotype, origin, virulence, and genotype. Listeria monocytogenes, a bacterium that is responsible for listeriosis, is a very diverse species. Desiccation resistance has been rarely studied in L. monocytogenes, although it is a stress that is largely encountered by this microorganism in food-processing environments and that could be managed to prevent its presence. The objective of this study was to evaluate the resistance of 30 L. monocytogenes strains to moderate desiccation (75% relative humidity) and evaluate the correlation of such resistance with the strains' virulence, serotype and genotype. The results showed a great heterogeneity of strains regarding their ability to survive (loss of cultivability between 0.4 and 2.0 log). Strains were classified into three groups according to desiccation resistance (sensitive, intermediate, or resistant), and the strain repartition was analyzed relative to serotype, virulence level and environmental origin of the strains. No correlation was found between isolate origin and desiccation resistance. All serotype 1/2b strains were classified into the group of resistant strains. Virulent and hypovirulent strains were distributed among the three groups of desiccation resistance. Finally, a genomic comparison was performed based on 31 genes that were previously identified as being involved in desiccation resistance. The presence of those genes was localized among the genomes of some strains and compared regarding strain-resistance levels. High nucleotide conservation was identified between resistant and desiccation-sensitive strains. In conclusion, the findings regarding the strains of serotype 1/2b indicate potential serotype-specific resistance to desiccation, and thus, to relative humidity fluctuations potentially encountered in food-related environments. The genomic comparison of 31 genes associated to desiccation tolerance did not reveal differences among four strains which have different level of resistance to desiccation.201728288399
254460.9967Antibiotic resistance potential of the healthy preterm infant gut microbiome. BACKGROUND: Few studies have investigated the gut microbiome of infants, fewer still preterm infants. In this study we sought to quantify and interrogate the resistome within a cohort of premature infants using shotgun metagenomic sequencing. We describe the gut microbiomes from preterm but healthy infants, characterising the taxonomic diversity identified and frequency of antibiotic resistance genes detected. RESULTS: Dominant clinically important species identified within the microbiomes included C. perfringens, K. pneumoniae and members of the Staphylococci and Enterobacter genera. Screening at the gene level we identified an average of 13 antimicrobial resistance genes per preterm infant, ranging across eight different antibiotic classes, including aminoglycosides and fluoroquinolones. Some antibiotic resistance genes were associated with clinically relevant bacteria, including the identification of mecA and high levels of Staphylococci within some infants. We were able to demonstrate that in a third of the infants the S. aureus identified was unrelated using MLST or metagenome assembly, but low abundance prevented such analysis within the remaining samples. CONCLUSIONS: We found that the healthy preterm infant gut microbiomes in this study harboured a significant diversity of antibiotic resistance genes. This broad picture of resistances and the wider taxonomic diversity identified raises further caution to the use of antibiotics without consideration of the resident microbial communities.201728149696
464270.9967Characterization of antibiotic resistance and host-microbiome interactions in the human upper respiratory tract during influenza infection. BACKGROUND: The abundance and diversity of antibiotic resistance genes (ARGs) in the human respiratory microbiome remain poorly characterized. In the context of influenza virus infection, interactions between the virus, the host, and resident bacteria with pathogenic potential are known to complicate and worsen disease, resulting in coinfection and increased morbidity and mortality of infected individuals. When pathogenic bacteria acquire antibiotic resistance, they are more difficult to treat and of global health concern. Characterization of ARG expression in the upper respiratory tract could help better understand the role antibiotic resistance plays in the pathogenesis of influenza-associated bacterial secondary infection. RESULTS: Thirty-seven individuals participating in the Household Influenza Transmission Study (HITS) in Managua, Nicaragua, were selected for this study. We performed metatranscriptomics and 16S rRNA gene sequencing analyses on nasal and throat swab samples, and host transcriptome profiling on blood samples. Individuals clustered into two groups based on their microbial gene expression profiles, with several microbial pathways enriched with genes differentially expressed between groups. We also analyzed antibiotic resistance gene expression and determined that approximately 25% of the sequence reads that corresponded to antibiotic resistance genes mapped to Streptococcus pneumoniae and Staphylococcus aureus. Following construction of an integrated network of ARG expression with host gene co-expression, we identified several host key regulators involved in the host response to influenza virus and bacterial infections, and host gene pathways associated with specific antibiotic resistance genes. CONCLUSIONS: This study indicates the host response to influenza infection could indirectly affect antibiotic resistance gene expression in the respiratory tract by impacting the microbial community structure and overall microbial gene expression. Interactions between the host systemic responses to influenza infection and antibiotic resistance gene expression highlight the importance of viral-bacterial co-infection in acute respiratory infections like influenza. Video abstract.202032178738
491980.9967Distribution of β-Lactamase Genes in Clinical Isolates from California Central Valley Hospital Deviates from the United States Nationwide Trends. The evolution and dissemination of antibiotic resistance genes throughout the world are clearly affected by the selection and migration of resistant bacteria. However, the relative contributions of selection and migration at a local scale have not been fully explored. We sought to identify which of these factors has the strongest effect through comparisons of antibiotic resistance gene abundance between a distinct location and its surroundings over an extended period of six years. In this work, we used two repositories of extended spectrum β-lactamase (ESBL)-producing isolates collected since 2013 from patients at Dignity Health Mercy Medical Center (DHMMC) in Merced, California, USA, and a nationwide database compiled from clinical isolate genomes reported by the National Center for Biotechnology Information (NCBI) since 2013. We analyzed the stability of average resistance gene frequencies over the years since collection of these clinical isolates began for each repository. We then compared the frequencies of resistance genes in the DHMMC collection with the averages of the nationwide frequencies. We found DHMMC gene frequencies are stable over time and differ significantly from nationwide frequencies throughout the period of time we examined. Our results suggest that local selective pressures are a more important influence on the population structure of resistance genes in bacterial populations than migration. This, in turn, indicates the potential for antibiotic resistance to be controlled at a regional level, making it easier to limit the spread through local stewardship.202133925352
241590.9967Profiles of Staphyloccocus aureus isolated from goat persistent mastitis before and after treatment with enrofloxacin. BACKGROUND: Staphylococcus aureus is one of the main causative agents of mastitis in small ruminants. Antimicrobial use is the major treatment, but there are many flaws linked to resistance, tolerance or persistence. This study aimed to verify changes in resistance, virulence and clonal profiles of S. aureus isolated from persistent mastitis goat milk before and after enrofloxacin treatment. RESULTS: MIC increased to at least one antimicrobial in S. aureus isolates after enrofloxacin treatment compared to before. The most detected resistance genes before and after treatment were tetK, tetM, and blaZ, with more resistance genes detected after enrofloxacin treatment (p < 0.05). Occasional variations in efflux system gene detection were observed before and after treatment. Nine virulence genes (hla, fnbA, fnbB, eta, etb, sea, sec, seh, and sej) were detected at both times, and between these, the hla and eta genes were detected more in isolates after treatment. All isolates of S. aureus belonged to the same sequence type (ST) 133, except for two S. aureus isolates prior to enrofloxacin treatment which were classified as ST5 and the other as a new one, ST4966. Isolates of S. aureus 4, 8, and 100 from before and after treatment had identical pulse types, while others obtained from other animals before and after treatment were classified into distinct pulse types. CONCLUSION: There were occasional changes in the studied profiles of S. aureus isolated before and after treatment of animals with enrofloxacin, which may have contributed to the permanence of bacteria in the mammary gland, even when using traditional treatment, resulting in persistent mastitis.202032448145
5820100.9966Sequencing Methods to Study the Microbiome with Antibiotic Resistance Genes in Patients with Pulmonary Infections. Various antibiotic-resistant bacteria (ARB) are known to induce repeated pulmonary infections and increase morbidity and mortality. A thorough knowledge of antibiotic resistance is imperative for clinical practice to treat resistant pulmonary infections. In this study, we used a reads-based method and an assembly-based method according to the metagenomic next-generation sequencing (mNGS) data to reveal the spectra of ARB and corresponding antibiotic resistance genes (ARGs) in samples from patients with pulmonary infections. A total of 151 clinical samples from 144 patients with pulmonary infections were collected for retrospective analysis. The ARB and ARGs detection performance was compared by the reads-based method and assembly-based method with the culture method and antibiotic susceptibility testing (AST), respectively. In addition, ARGs and the attribution relationship of common ARB were analyzed by the two methods. The comparison results showed that the assembly-based method could assist in determining pathogens detected by the reads-based method as true ARB and improve the predictive capabilities (46% > 13%). ARG-ARB network analysis revealed that assembly-based method could promote determining clear ARG-bacteria attribution and 101 ARGs were detected both in two methods. 25 ARB were obtained by both methods, of which the most predominant ARB and its ARGs in the samples of pulmonary infections were Acinetobacter baumannii (ade), Pseudomonas aeruginosa (mex), Klebsiella pneumoniae (emr), and Stenotrophomonas maltophilia (sme). Collectively, our findings demonstrated that the assembly-based method could be a supplement to the reads-based method and uncovered pulmonary infection-associated ARB and ARGs as potential antibiotic treatment targets.202439113195
5823110.9966Comparing Patient Risk Factor-, Sequence Type-, and Resistance Locus Identification-Based Approaches for Predicting Antibiotic Resistance in Escherichia coli Bloodstream Infections. Rapid diagnostic tests for antibiotic resistance that identify the presence or absence of antibiotic resistance genes/loci are increasingly being developed. However, these approaches usually neglect other sources of predictive information which could be identified over shorter time periods, including patient epidemiologic risk factors for antibiotic resistance and markers of lineage. Using a data set of 414 Escherichia coli isolates recovered from separate episodes of bacteremia at a single academic institution in Toronto, Ontario, Canada, between 2010 and 2015, we compared the potential predictive ability of three approaches (epidemiologic risk factor-, pathogen sequence type [ST]-, and resistance gene identification-based approaches) for classifying phenotypic resistance to three antibiotics representing classes of broad-spectrum antimicrobial therapy (ceftriaxone [a 3rd-generation cephalosporin], ciprofloxacin [a fluoroquinolone], and gentamicin [an aminoglycoside]). We used logistic regression models to generate model receiver operating characteristic (ROC) curves. Predictive discrimination was measured using apparent and corrected (bootstrapped) areas under the curves (AUCs). Epidemiologic risk factor-based models based on two simple risk factors (prior antibiotic exposure and recent prior susceptibility of Gram-negative bacteria) provided a modest predictive discrimination, with AUCs ranging from 0.65 to 0.74. Sequence type-based models demonstrated strong discrimination (AUCs, 0.83 to 0.94) across all three antibiotic classes. The addition of epidemiologic risk factors to sequence type significantly improved the ability to predict resistance for all antibiotics (P < 0.05). Resistance gene identification-based approaches provided the highest degree of discrimination (AUCs, 0.88 to 0.99), with no statistically significant benefit being achieved by adding the patient epidemiologic predictors. In summary, sequence type or other lineage-based approaches could produce an excellent discrimination of antibiotic resistance and may be improved by incorporating readily available patient epidemiologic predictors but are less discriminatory than identification of the presence of known resistance loci.201930894438
3122120.9966Hybrid sequence-based analysis reveals the distribution of bacterial species and genes in the oral microbiome at a high resolution. Bacteria in the oral microbiome are poorly identified owing to the lack of established culture methods for them. Thus, this study aimed to use culture-free analysis techniques, including bacterial single-cell genome sequencing, to identify bacterial species and investigate gene distribution in saliva. Saliva samples from the same individual were classified as inactivated or viable and then analyzed using 16S rRNA sequencing, metagenomic shotgun sequencing, and bacterial single-cell sequencing. The results of 16S rRNA sequencing revealed similar microbiota structures in both samples, with Streptococcus being the predominant genus. Metagenomic shotgun sequencing showed that approximately 80 % of the DNA in the samples was of non-bacterial origin, whereas single-cell sequencing showed an average contamination rate of 10.4 % per genome. Single-cell sequencing also yielded genome sequences for 43 out of 48 wells for the inactivated samples and 45 out of 48 wells for the viable samples. With respect to resistance genes, four out of 88 isolates carried cfxA, which encodes a β-lactamase, and four isolates carried erythromycin resistance genes. Tetracycline resistance genes were found in nine bacteria. Metagenomic shotgun sequencing provided complete sequences of cfxA, ermF, and ermX, whereas other resistance genes, such as tetQ and tetM, were detected as fragments. In addition, virulence factors from Streptococcus pneumoniae were the most common, with 13 genes detected. Our average nucleotide identity analysis also suggested five single-cell-isolated bacteria as potential novel species. These data would contribute to expanding the oral microbiome data resource.202438708423
3246130.9966Antibiotic Resistance Gene Detection in the Microbiome Context. Within the past decade, microbiologists have moved from detecting single antibiotic resistance genes (ARGs) to detecting all known resistance genes within a sample due to advances in next generation sequencing. This has provided a wealth of data on the variation and relative abundances of ARGs present in a total bacterial population. However, to use these data in terms of therapy or risk to patients, they must be analyzed in the context of the background microbiome. Using a quantitative PCR ARG chip and 16S rRNA amplicon sequencing, we have sought to identify the ARGs and bacteria present in a fecal sample of a healthy adult using genomic tools. Of the 42 ARGs detected, 12 fitted into the ResCon1 category of ARGs: cfxA, cphA, bacA, sul3, aadE, bla(TEM), aphA1, aphA3, aph(2')-Id, aacA/aphd, catA1, and vanC. Therefore, we describe these 12 genes as the core resistome of this person's fecal microbiome and the remaining 30 ARGs as descriptors of the microbial population within the fecal microbiome. The dominant phyla and genera agree with those previously detected in the greatest abundances in fecal samples of healthy humans. The majority of the ARGs detected were associated with the presence of specific bacterial taxa, which were confirmed using microbiome analysis. We acknowledge the limitations of the data in the context of the limited sample set. However, the principle of combining qPCR and microbiome analysis was shown to be helpful to identify the association of the ARGs with specific taxa.201829185915
2543140.9966Capturing the antibiotic resistome of preterm infants reveals new benefits of probiotic supplementation. BACKGROUND: Probiotic use in preterm infants can mitigate the impact of antibiotic exposure and reduce rates of certain illnesses; however, the benefit on the gut resistome, the collection of antibiotic resistance genes, requires further investigation. We hypothesized that probiotic supplementation of early preterm infants (born < 32-week gestation) while in hospital reduces the prevalence of antibiotic resistance genes associated with pathogenic bacteria in the gut. We used a targeted capture approach to compare the resistome from stool samples collected at the term corrected age of 40 weeks for two groups of preterm infants (those that routinely received a multi-strain probiotic during hospitalization and those that did not) with samples from full-term infants at 10 days of age to identify if preterm birth or probiotic supplementation impacted the resistome. We also compared the two groups of preterm infants up to 5 months of age to identify persistent antibiotic resistance genes. RESULTS: At the term corrected age, or 10 days of age for the full-term infants, we found over 80 antibiotic resistance genes in the preterm infants that did not receive probiotics that were not identified in either the full-term or probiotic-supplemented preterm infants. More genes associated with antibiotic inactivation mechanisms were identified in preterm infants unexposed to probiotics at this collection time-point compared to the other infants. We further linked these genes to mobile genetic elements and Enterobacteriaceae, which were also abundant in their gut microbiomes. Various genes associated with aminoglycoside and beta-lactam resistance, commonly found in pathogenic bacteria, were retained for up to 5 months in the preterm infants that did not receive probiotics. CONCLUSIONS: This pilot survey of preterm infants shows that probiotics administered after preterm birth during hospitalization reduced the diversity and prevented persistence of antibiotic resistance genes in the gut microbiome. The benefits of probiotic use on the microbiome and the resistome should be further explored in larger groups of infants. Due to its high sensitivity and lower sequencing cost, our targeted capture approach can facilitate these surveys to further address the implications of resistance genes persisting into infancy without the need for large-scale metagenomic sequencing. Video Abstract.202236008821
3159150.9966Longitudinal development of the dust microbiome in a newly opened Norwegian kindergarten. BACKGROUND: In Norway, 91% of children aged 1-5 attend kindergarten where they are exposed to indoor microbiomes which can have relevance for development and health. In order to gain a better understanding of the composition of the indoor microbiome and how it is affected by occupancy over time, floor dust samples from a newly opened kindergarten were investigated. Samples were collected during an 11-month period. Samples were analyzed for bacterial composition using 16S rRNA gene sequencing. Samples were also screened for four clinically relevant antibiotic resistance genes. In addition, Petrifilm analyses were used to evaluate surface hygiene. RESULTS: Significant changes in the microbial community composition were observed over time (PERMANOVA, P < 0.05). Particularly, changes in the abundance and the proportions of human associated bacteria were found. A decrease in the prevalence of Propionibacterium from over 16% abundance to less than 1% and an increase in Streptococcus from 10 to 16% were the most significant findings. Four classes of clinically relevant antibiotic resistance genes were tested for; three were detected in the dust, indicating the presence of resistant bacteria and a potential for resistance spread. Petrifilm analysis showed that some surfaces in the kindergarten were of consistent poor hygienic quality, and new hygienic routines are required. CONCLUSIONS: This study, which is the first of its kind performed at a newly opened kindergarten, reveals changes in the microbiome over time as well as the presence of antibiotic resistance genes and hygiene issues which are of relevance for occupant health.201830219104
5809160.9966Genomic epidemiology of Streptococcus agalactiae ST283 in Southeast Asia. Streptococcus agalactiae, also known as Lancefield Group B Streptococcus (GBS), is typically regarded as a neonatal pathogen; however, several studies have shown that the bacteria are capable of causing invasive diseases in non-pregnant adults as well. The majority of documented cases were from Southeast Asian countries, and the most common genotype found was ST283, which is also known to be able to infect fish. This study sequenced 12 GBS ST283 samples collected from adult patients in Thailand. Together with publicly available sequences, we performed temporo-spatial analysis and estimated population dynamics of the bacteria. Putative drug resistance genes were also identified and characterized, and the drug resistance phenotypes were validated experimentally. The results, together with historical records, draw a detailed picture of the past transmission history of GBS ST283 in Southeast Asia.202235264716
5811170.9966Antimicrobial susceptibility testing and tentative epidemiological cut-off values for Lactobacillaceae family species intended for ingestion. INTRODUCTION: In this work, 170 strains covering 13 species from the Lactobacillaceae family were analyzed to determine minimal inhibitory concentration (MIC) distributions to nine antimicrobial agents, and genes potentially conferring resistance. This allows a proposal of tentative Epidemiological Cut-Offs (ECOFFs) that follows the phylogeny for interpretation of resistance in the 13 species. METHODS: The 170 strains originated from different sources, geographical areas, and time periods. MICs for nine antibiotics were determined according to the ISO 10932 standard for lactobacillia and by a modified CLSI-method for Leuconostoc and Pediococcus which ensured sufficient growth. The strains were whole genome sequenced, subtyped by core genome analysis, and assessed for the presence of antibiotic resistance genes using the ResFinder and NCBI AMRFinder databases. RESULTS AND DISCUSSION: The data provide evidence that antimicrobial susceptibility follows phylogeny instead of fermentation pattern and accordingly, tentative ECOFFs were defined. For some species the tentative ECOFFs for specific antibiotics are above the cut-off values set by the European Food Safety Authority (EFSA) which are primarily defined according to fermentation pattern or at genus level. The increased tolerance for specific antibiotics observed for some species was evaluated to be innate, as only for one strain phenotypic resistance was found to be related to an acquired resistance gene. In general, more data are needed to define ECOFFs and since the number of isolates available for industrial relevant bacterial species are often limited compared to clinically relevant species, it is important; 1) that strains are unambiguously defined at species level and subtyped through core genome analysis, 2) MIC determination are performed by use of a standardized method to define species-specific MIC distributions and 3) that known antimicrobial resistance genes are determined in whole genome sequences to support the MIC determinations.202339816654
2553180.9966Early-life gut microbiome modulation reduces the abundance of antibiotic-resistant bacteria. BACKGROUND: Antibiotic-resistant (AR) bacteria are a global threat. AR bacteria can be acquired in early life and have long-term sequelae. Limiting the spread of antibiotic resistance without triggering the development of additional resistance mechanisms is of immense clinical value. Here, we show how the infant gut microbiome can be modified, resulting in a significant reduction of AR genes (ARGs) and the potentially pathogenic bacteria that harbor them. METHODS: The gut microbiome was characterized using shotgun metagenomics of fecal samples from two groups of healthy, term breastfed infants. One group was fed B. infantis EVC001 in addition to receiving lactation support (n = 29, EVC001-fed), while the other received lactation support alone (n = 31, controls). Coliforms were isolated from fecal samples and genome sequenced, as well as tested for minimal inhibitory concentrations against clinically relevant antibiotics. RESULTS: Infants fed B. infantis EVC001 exhibited a change to the gut microbiome, resulting in a 90% lower level of ARGs compared to controls. ARGs that differed significantly between groups were predicted to confer resistance to beta lactams, fluoroquinolones, or multiple drug classes, the majority of which belonged to Escherichia, Clostridium, and Staphylococcus. Minimal inhibitory concentration assays confirmed the resistance phenotypes among isolates with these genes. Notably, we found extended-spectrum beta lactamases among healthy, vaginally delivered breastfed infants who had never been exposed to antibiotics. CONCLUSIONS: Colonization of the gut of breastfed infants by a single strain of B. longum subsp. infantis had a profound impact on the fecal metagenome, including a reduction in ARGs. This highlights the importance of developing novel approaches to limit the spread of these genes among clinically relevant bacteria. Future studies are needed to determine whether colonization with B. infantis EVC001 decreases the incidence of AR infections in breastfed infants. TRIAL REGISTRATION: This clinical trial was registered at ClinicalTrials.gov, NCT02457338.201931423298
5687190.9966The effect of short-course antibiotics on the resistance profile of colonizing gut bacteria in the ICU: a prospective cohort study. BACKGROUND: The need for early antibiotics in the intensive care unit (ICU) is often balanced against the goal of antibiotic stewardship. Long-course antibiotics increase the burden of antimicrobial resistance within colonizing gut bacteria, but the dynamics of this process are not fully understood. We sought to determine how short-course antibiotics affect the antimicrobial resistance phenotype and genotype of colonizing gut bacteria in the ICU by performing a prospective cohort study with assessments of resistance at ICU admission and exactly 72 h later. METHODS: Deep rectal swabs were performed on 48 adults at the time of ICU admission and exactly 72 h later, including patients who did and did not receive antibiotics. To determine resistance phenotype, rectal swabs were cultured for methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE). In addition, Gram-negative bacterial isolates were cultured against relevant antibiotics. To determine resistance genotype, quantitative PCR (qPCR) was performed from rectal swabs for 87 established resistance genes. Within-individual changes in antimicrobial resistance were calculated based on culture and qPCR results and correlated with exposure to relevant antibiotics (e.g., did β-lactam antibiotic exposure associate with a detectable change in β-lactam resistance over this 72-h period?). RESULTS: Of 48 ICU patients, 41 (85%) received antibiotics. Overall, there was no increase in the antimicrobial resistance profile of colonizing gut bacteria during the 72-h study period. There was also no increase in antimicrobial resistance after stratification by receipt of antibiotics (i.e., no detectable increase in β-lactam, vancomycin, or macrolide resistance regardless of whether patients received those same antibiotics). This was true for both culture and PCR. Antimicrobial resistance pattern at ICU admission strongly predicted resistance pattern after 72 h. CONCLUSIONS: Short-course ICU antibiotics made little detectable difference in the antimicrobial resistance pattern of colonizing gut bacteria over 72 h in the ICU. This provides an improved understanding of the dynamics of antimicrobial resistance in the ICU and some reassurance that short-course antibiotics may not adversely impact the stewardship goal of reducing antimicrobial resistance.202032646458