The role of the glycome in symbiotic host-microbe interactions.

Glycosylation plays a crucial role in many aspects of cell biology, including cellular and organismal integrity, structure-and-function of many glycosylated molecules in the cell, signal transduction, development, cancer, and in a number of diseases. Besides, at the inter-organismal level of interaction, a variety of glycosylated molecules are involved in the host-microbiota recognition and initiation of downstream signalling cascades depending on the outcomes of the glycome-mediated ascertainment. The role of glycosylation in host-microbe interactions is better elaborated within the context of virulence and pathogenicity in bacterial infection processes but the symbiotic host-microbe relationships also involve substantive glycome-mediated interactions. The works in the latter field have been reviewed to a much lesser extent, and the main aim of this mini-review is to compensate for this deficiency and summarise the role of glycomics in host-microbe symbiotic interactions.

Molecular Epidemiology and Virulence of Non-Typhoidal Salmonella in Armenia.

In this work, we analysed human isolates of nontyphoidal Salmonella enterica subsp. enterica (NTS), which were collected from salmonellosis cases in Armenia from 1996 to 2019. This disease became a leading food-borne bacterial infection in the region, with the younger age groups especially affected. The isolates were characterised by serotyping, Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR) typing, and whole genome sequencing (WGS). The main serotypes were S. Typhimurium, S. Enteritidis, and S. Arizonae. ERIC-PCR indicated a high degree of clonality among S. Typhimurium strains, which were also multidrug-resistant and produced extended spectrum beta-lactamases. During the study period, the frequency of S. Typhimurium and S. Arizonae isolations decreased, but with the increase in S. Enteritidis and other NTS. A total of 42 NTS isolates were subjected to WGS and explored for virulence-related traits and the corresponding genetic elements. Some virulence and genetic factors were shared by all NTS serotypes, while the main differences were attributed to the serotype-specific diversity of virulence genes, SPIs, virulence plasmids, and phages. The results indicated the variability and dynamics in the epidemiology of salmonellosis and a high virulence potential of human NTS isolates circulating in the region.

Acquisition and Spread of Antimicrobial Resistance: A tet(X) Case Study.

Understanding the mechanisms leading to the rise and dissemination of antimicrobial resistance (AMR) is crucially important for the preservation of power of antimicrobials and controlling infectious diseases. Measures to monitor and detect AMR, however, have been significantly delayed and introduced much later after the beginning of industrial production and consumption of antimicrobials. However, monitoring and detection of AMR is largely focused on bacterial pathogens, thus missing multiple key events which take place before the emergence and spread of AMR among the pathogens. In this regard, careful analysis of AMR development towards recently introduced antimicrobials may serve as a valuable example for the better understanding of mechanisms driving AMR evolution. Here, the example of evolution of tet(X), which confers resistance to the next-generation tetracyclines, is summarised and discussed. Initial mechanisms of resistance to these antimicrobials among pathogens were mostly via chromosomal mutations leading to the overexpression of efflux pumps. High-level resistance was achieved only after the acquisition of flavin-dependent monooxygenase-encoding genes from the environmental microbiota. These genes confer resistance to all tetracyclines, including the next-generation tetracyclines, and thus were termed tet(X). ISCR2 and IS26, as well as a variety of conjugative and mobilizable plasmids of different incompatibility groups, played an essential role in the acquisition of tet(X) genes from natural reservoirs and in further dissemination among bacterial commensals and pathogens. This process, which took place within the last decade, demonstrates how rapidly AMR evolution may progress, taking away some drugs of last resort from our arsenal.

Segmented filamentous bacteria are a major group in terminal ileum of piglets.

Metabolically active microbiota of the porcine terminal ileum mucosa was analyzed by RT-PCR of 16S rRNAs. The majority of 1951 sequences retrieved (24.8%) displayed the closest similarity to segmented filamentous bacteria (SFB). Phylogenetic analysis inferred the host-specific clusters of SFB sequences suggesting the host-specific selection of this group of bacteria.

Phages for treatment of Escherichia coli infections.

Diseases due to infections by pathogenic Escherichia coli strains are on the rise and with the growing antimicrobial resistance among bacterial pathogens, including this group. Thus, alternative therapeutic options are actively investigated. Among these alternatives is phage therapy. In the case of E. coli, the combination of the well understood biology of this species and its bacteriophages represents a good guiding example for the establishment of phage therapy principles against this and other pathogenic bacteria. In this chapter, the procedures toward the development of phage therapy against pathogenic E. coli with the use of T-even group of phages are discussed. These steps involve the isolation, purification, characterisation and large-scale production of these phages, with formulation of phage cocktails for in vitro and in vivo studies. The main emphasis is made on phage therapy of enteropathogenic E. coli O157:H, which is one of the prominent human pathogens but persists as a commensal bacterium in many food animals. The implementation of phage therapy against E. coli O157:H within the One Health framework in carrier animals and for treatment of meat, vegetables, fruits and other agricultural produce thus would allow controlling and interrupting the transmission routes of this pathogen to the human food chain and preventing human disease. Examples of successful control and elimination of E. coli O157:H are given, while the problems encountered in phage treatment of this pathogen are also discussed.

Metabolomics in antimicrobial drug discovery.

INTRODUCTION: In light of the ever-escalating problem of antimicrobial resistance, there is an urgent need for the development of new antimicrobials. In this review, the role of metabolomics in antimicrobial drug discovery and development is summarized and discussed. For this, ScienceDirect, PubMed, Web of Science and Google Scholar databases were searched with the article's keywords and their combinations to retrieve the most relevant and up-to-date information. AREAS COVERED: The areas covered include the metabolomic concepts and techniques and bioinformatic tools used in metabolomics as well as recent developments in these areas. Also, examples of the use of metabolomics tools in several areas of antimicrobial drug discovery are given. EXPERT OPINION: Metabolomics, with the corresponding bioinformatic support and combination with other omics technologies, represents an integral and essential part of antimicrobial drug discovery and development. Metabolomics contributes to the mechanism-based approach in antimicrobial drug discovery, reveals the mechanisms of action of antimicrobials and non-antimicrobial compounds, identifies new targets, and opens new ways to manage and control bacterial infections.

Antimicrobial drug resistance mechanisms among Mollicutes.

Representatives of the Mollicutes class are the smallest, wall-less bacteria capable of independent reproduction. They are widespread in nature, most are commensals, and some are pathogens of humans, animals and plants. They are also the main contaminants of cell cultures and vaccine preparations. Despite limited biosynthetic capabilities, they are highly adaptable and capable of surviving under various stress and extreme conditions, including antimicrobial selective pressure. This review describes current understanding of antibiotic resistance (ABR) mechanisms in Mollicutes. Protective mechanisms in these bacteria include point mutations, which may include non-target genes, and unique gene exchange mechanisms, contributing to transfer of ABR genes. Better understanding of the mechanisms of emergence and dissemination of ABR in Mollicutes is crucial to control these hypermutable bacteria and prevent the occurrence of highly ABR strains.

Comparative analysis of cytokine profiles in autoinflammatory and autoimmune conditions.

BACKGROUND: Many disease states are characterized by a biased cytokine profile and determining the level of certain cytokines becomes an important diagnostic and research tool in clinical medicine. We hypothesized that, despite the highly dynamic nature, the profile of cytokines may be characteristic of disease. METHODS: The level of systemic cytokines was studied in an autoinflammatory condition, familial Mediterranean fever; in an autoimmune disease, systemic lupus erythematosus (SLE), and in healthy controls. RESULTS: Multivariate statistics with the use of seven variables clustered the patients and control subjects into the four well separated and distinct groups, corresponding to the SLE, FMF attack, FMF remission, and healthy states. CONCLUSIONS: The model suggested the existence of specific patterns in cytokine levels reflecting the healthy and specific disease states. These findings warrant further investigations to establish whether this approach may serve as a potential meta-biomarker for other inflammatory disorders.

Molecular ecology of macrolide-lincosamide-streptogramin B methylases in waste lagoons and subsurface waters associated with swine production.

RNA methylase genes are common antibiotic resistance determinants for multiple drugs of the macrolide, lincosamide, and streptogramin B (MLS(B)) families. We used molecular methods to investigate the diversity, distribution, and abundance of MLS(B) methylases in waste lagoons and groundwater wells at two swine farms with a history of tylosin (a macrolide antibiotic structurally related to erythromycin) and tetracycline usage. Phylogenetic analysis guided primer design for quantification of MLS(B) resistance genes found in tylosin-producing Streptomyces (tlr(B), tlr(D)) and commensal/pathogenic bacteria (erm(A), erm(B), erm(C), erm(F), erm(G), erm(Q)). The near absence of tlr genes at these sites suggested a lack of native antibiotic-producing organisms. The gene combination erm(ABCF) was found in all lagoon samples analyzed. These four genes were also detected with high frequency in wells previously found to be contaminated by lagoon leakage. A weak correlation was found between the distribution of erm genes and previously reported patterns of tetracycline resistance determinants, suggesting that dissemination of these genes into the environment is not necessarily linked. Considerations of gene origins in history (i.e., phylogeny) and gene distributions in the landscape provide a useful "molecular ecology" framework for studying environmental spread of antibiotic resistance.

Environmentally-acquired bacteria influence microbial diversity and natural innate immune responses at gut surfaces.

BACKGROUND: Early microbial colonization of the gut reduces the incidence of infectious, inflammatory and autoimmune diseases. Recent population studies reveal that childhood hygiene is a significant risk factor for development of inflammatory bowel disease, thereby reinforcing the hygiene hypothesis and the potential importance of microbial colonization during early life. The extent to which early-life environment impacts on microbial diversity of the adult gut and subsequent immune processes has not been comprehensively investigated thus far. We addressed this important question using the pig as a model to evaluate the impact of early-life environment on microbe/host gut interactions during development. RESULTS: Genetically-related piglets were housed in either indoor or outdoor environments or in experimental isolators. Analysis of over 3,000 16S rRNA sequences revealed major differences in mucosa-adherent microbial diversity in the ileum of adult pigs attributable to differences in early-life environment. Pigs housed in a natural outdoor environment showed a dominance of Firmicutes, in particular Lactobacillus, whereas animals housed in a hygienic indoor environment had reduced Lactobacillus and higher numbers of potentially pathogenic phylotypes. Our analysis revealed a strong negative correlation between the abundance of Firmicutes and pathogenic bacterial populations in the gut. These differences were exaggerated in animals housed in experimental isolators. Affymetrix microarray technology and Real-time Polymerase Chain Reaction revealed significant gut-specific gene responses also related to early-life environment. Significantly, indoor-housed pigs displayed increased expression of Type 1 interferon genes, Major Histocompatibility Complex class I and several chemokines. Gene Ontology and pathway analysis further confirmed these results. CONCLUSION: Early-life environment significantly affects both microbial composition of the adult gut and mucosal innate immune function. We observed that a microbiota dominated by lactobacilli may function to maintain mucosal immune homeostasis and limit pathogen colonization.

Fermented liquid feed enhances bacterial diversity in piglet intestine.

Because of limitations imposed on the antibiotic use in animal industry, there is a need for alternatives to maintain the efficiency of production. One of them may be the use of fermented liquid feed (FLF) but how it affects gut ecology is poorly understood. We investigated the effect of three diets, standard dry feed (control), dry feed supplemented with antibiotics, and fermented liquid feed (FLF, fermented with Lactobacillus plantarum), on gut bacterial diversity in piglets. The structure of the ileal and caecal communities was estimated by sequencing the SSU rRNA gene libraries. Antibiotic-supplemented feed slightly increased bacterial diversity in the ileum but reduced it in the caecum while in FLF-fed animals bacterial diversity was elevated. The majority of bacterial sequences in the ileum of all three groups belonged to lactobacilli (92-98%). In the caecum the lactobacilli were still dominant in control and antibiotic-fed animals (59% and 64% of total bacterial sequences, respectively) but in FLF-fed animals they fell to 31% with the concomitant increase in the Firmicutes diversity represented by the Dorea, Coprococcus, Roseburia and Faecalibacterium genera. Thus FLF affects the gut ecology in a different way than antibiotics and contributes to the enhanced bacterial diversity in the gastrointestinal tract.

Extended-Spectrum ß-Lactamases in Human Isolates of Multidrug-Resistant Non-typhoidal Salmonella enterica.

A total of 291 non-duplicate isolates of non-typhoidal Salmonella (NTS) were collected from the fecal samples of patients with salmonellosis in Armenia and Georgia during 1996-2016. The isolates were tested for resistance to antimicrobials, including extended-spectrum ß-lactamases (ESBL). The high prevalence of multidrug-resistance (MDR) and ESBL-producer phenotypes was detected among Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) isolates collected from patients in Armenia between 1996 and 2016. A total of 36 MDR NTS isolates were subjected to whole genome sequencing (WGS) to determine the genetic background of antimicrobial resistance (AMR) and mobile genetic elements. All ESBL-producing S. Typhimurium isolates belonged to the same sequence type (ST328). The ESBL-producer phenotype was associated with plasmid-encoded CTX-M-5 production. A range of other plasmids was associated with resistance to other antimicrobials, including the MDR phenotype.

The role of antibiotics and antibiotic resistance in nature.

Investigations of antibiotic resistance from an environmental prospective shed new light on a problem that was traditionally confined to a subset of clinically relevant antibiotic-resistant bacterial pathogens. It is clear that the environmental microbiota, even in apparently antibiotic-free environments, possess an enormous number and diversity of antibiotic resistance genes, some of which are very similar to the genes circulating in pathogenic microbiota. It is difficult to explain the role of antibiotics and antibiotic resistance in natural environments from an anthropocentric point of view, which is focused on clinical aspects such as the efficiency of antibiotics in clearing infections and pathogens that are resistant to antibiotic treatment. A broader overview of the role of antibiotics and antibiotic resistance in nature from the evolutionary and ecological prospective suggests that antibiotics have evolved as another way of intra- and inter-domain communication in various ecosystems. This signalling by non-clinical concentrations of antibiotics in the environment results in adaptive phenotypic and genotypic responses of microbiota and other members of the community. Understanding the complex picture of evolution and ecology of antibiotics and antibiotic resistance may help to understand the processes leading to the emergence and dissemination of antibiotic resistance and also help to control it, at least in relation to the newer antibiotics now entering clinical practice.

Tetracycline resistome of the organic pig gut.

The occurrence of genes conferring resistance to tetracyclines in the organic pig gut was assessed through the metagenomic approach. Of 9,000 bacterial artificial chromosome clones analyzed, 10 were identified as carrying the known tet(C), tet(W), and tet(40) genes, as well as novel genes encoding resistance to the tetracyclines minocycline and doxycycline. The latter are different from the known tet genes and are homologous to genes encoding UDP-glucose 4-epimerases, with the domain structure characteristic for these enzymes. The majority of the resistance genes were associated with putative mobile genetic elements. The sequence of a novel 9.7-kb plasmid carrying tet(W) and tet(40) was also identified. Conserved flanking regions identified around the tet(W) and tet(40) genes in our metagenomic library may play a role in genetic exchange of these genes. This is the first report describing the occurrence of tet(40) outside the human intestine. The maintenance of antibiotic resistance genes in apparently antibiotic-free animals is probably due to their presence on mobile genetic elements, the fitness cost of which for the cell is ameliorated during the previous antibiotic selection.

Fate and transport of antibiotic residues and antibiotic resistance genes following land application of manure waste.

Antibiotics are used in animal livestock production for therapeutic treatment of disease and at subtherapeutic levels for growth promotion and improvement of feed efficiency. It is estimated that approximately 75% of antibiotics are not absorbed by animals and are excreted in waste. Antibiotic resistance selection occurs among gastrointestinal bacteria, which are also excreted in manure and stored in waste holding systems. Land application of animal waste is a common disposal method used in the United States and is a means for environmental entry of both antibiotics and genetic resistance determinants. Concerns for bacterial resistance gene selection and dissemination of resistance genes have prompted interest about the concentrations and biological activity of drug residues and break-down metabolites, and their fate and transport. Fecal bacteria can survive for weeks to months in the environment, depending on species and temperature, however, genetic elements can persist regardless of cell viability. Phylogenetic analyses indicate antibiotic resistance genes have evolved, although some genes have been maintained in bacteria before the modern antibiotic era. Quantitative measurements of drug residues and levels of resistance genes are needed, in addition to understanding the environmental mechanisms of genetic selection, gene acquisition, and the spatiotemporal dynamics of these resistance genes and their bacterial hosts. This review article discusses an accumulation of findings that address aspects of the fate, transport, and persistence of antibiotics and antibiotic resistance genes in natural environments, with emphasis on mechanisms pertaining to soil environments following land application of animal waste effluent.

Environmental Triggers of Autoreactive Responses: Induction of Antiphospholipid Antibody Formation.

Antiphospholipid antibodies (aPLs) comprise a diverse family of autoantibodies targeted against proteins with the affinity toward negatively charged phospholipids or protein-phospholipid complexes. Their clinical significance, including prothrombotic potential of anti-cardiolipin antibodies (aCLs), anti-ß2-glycoprotein I antibodies (aß2-GPIs), and lupus anti-coagulant (LA), is well-established. However, the ontogeny of these pathogenic aPLs remains less clear. While transient appearance of aPLs could be induced by various environmental factors, in genetically predisposed individuals these factors may eventually lead to the development of the antiphospholipid syndrome (APS). Since the first description of APS, it has been found that a wide variety of microbial and viral agents influence aPLs production and contribute to clinical manifestations of APS. Many theories attempted to explain the pathogenic potential of different environmental factors as well as a phenomenon termed molecular mimicry between ß2-GPI molecule and infection-relevant structures. In this review, we summarize and critically assess the pathogenic and non-pathogenic formation of aPLs and its contribution to the development of APS.

Omics of antimicrobials and antimicrobial resistance.

INTRODUCTION: The development of new antimicrobials has become an urgent priority because of a global challenge emerging from the rise of antimicrobial resistant pathogens. Areas covered: In this review, the authors discuss the opportunities offered by modern omics approaches to address the challenge and the use of this approach in antimicrobial development. Specifically, the authors focus on the role of omics technologies and bioinformatics for the revelation of the effects of antimicrobials in a variety of microbial cellular processes, as well as the identification of potential cellular targets, the mechanisms of antimicrobial resistance, and the development of new antimicrobials. Expert opinion: Prevention of antimicrobial resistance does not only depend on rational drug design such as narrow-spectrum antimicrobials but on several factors. It is the opinion of the authors that the use of a multi-omics bioinformatics approach should become an integral part of antimicrobial drug discovery as well as in the prevention of antimicrobial resistance.

Potential Involvement of Salmonella Infection in Autoimmunity.

In this work, we investigated the potential effects of nontyphoidal Salmonella infection on autoantibody (AA) formation. The titer and profiles of autoantibodies in the sera of patients with acute salmonellosis due to Salmonella enterica serovar Typhimurium (S. Typhimurium) or Salmonella enterica serovar Enteritidis (S. Enteritidis) infection, as well as in convalescent patients, were determined with indirect immunofluorescence. A significant increase of autoantibodies in acute diseases caused by both serotypes of Salmonella and during post infection by S. Enteritidis was detected. Antibody profile analysis by multivariate statistics revealed that this increase was non-specific and was not dependent on the infectious agent or disease stage. The results obtained suggest that nontyphoidal Salmonella infection contributes to the generation of autoantibodies and may play a role in autoimmune disease.