Search Results (273)

Proteins involved in antibiotic resistance (resistome) and with antimicrobial activity are present in biological specimens. This study aims to explore the presence and abundance of antimicrobial peptides (AMPs) and resistome proteins in bovine milk from diverse breeds and from intensive (Pezzata rossa, Bruna alpina, and Frisona) and non-intensive farming (Podolica breeds). Liquid atmospheric pressure matrix-assisted laser desorption/ionization (LAP-MALDI) mass spectrometry (MS) profiling, bottom-up proteomics, and metaproteomics were used to comprehensively analyze milk samples from various bovine breeds in order to identify and characterize AMPs and to investigate resistome proteins. LAP-MALDI MS coupled with linear discriminant analysis (LDA) machine learning was employed as a rapid classification method for Podolica milk recognition against the milk of other bovine species. The results of the LAP-MALDI MS analysis of milk coupled with the linear discriminant analysis (LDA) demonstrate the potential of distinguishing between Podolica and control milk samples based on MS profiles. The classification accuracy achieved in the training set is 86% while it reaches 98.4% in the test set. Bottom-up proteomics revealed approximately 220 quantified bovine proteins (identified using the Bos taurus database), with cathelicidins and annexins exhibiting higher abundance levels in control cows (intensive farming breeds). On the other hand, the metaproteomics analysis highlighted the diversity within the milk’s microbial ecosystem with interesting results that may reflect the diverse environmental variables. The bottom-up proteomics data analysis using the Comprehensive Antibiotic Resistance Database (CARD) revealed beta-lactamases and tetracycline resistance proteins in both control and Podolica milk samples, with no relevant breed-specific differences observed. Full article

Background and Objectives: In the undertaken study, proteomics alterations of blood-borne XDR S. Typhi isolated from Pakistan were investigated using mass spectrometry. Materials and Methods: MDR and XDR S. Typhi total protein lysates were fractionated, digested, and processed for nanoflow LC-LTQ-Orbitrap MS analysis. Results: Among the 1267 identified proteins, 37 were differentially regulated, of which 28 were up-regulated, and 9 were down-regulated in XDR S. Typhi as compared to MDR S. Typhi. Based on the functional annotation, proteins found up-regulated are involved mainly in metabolic pathways (ManA, FadB, DacC, GpmA, AphA, PfkB, TalA, FbaB, OtsA, 16504242), the biosynthesis of secondary metabolites (ManA, FadB, GlpB, GpmA, PfkB, TalA, FbaB, OtsA), microbial metabolism in diverse environments (FadB, GpmA, PfkB, NfnB, TalA, FbaB), and ABC transporters (PstS, YbeJ, MglB, RbsB, ArtJ). Proteins found down-regulated are involved mainly in carbon metabolism (FadB, GpmA, PfkB, FalA, FbaB) and the biosynthesis of amino acids (GpmA, PfkB, TalA, FbaB). Most of the identified differential proteins were predicted to be antigenic, and matched with resistome data. Conclusions: A total of 28 proteins were up-regulated, and 9 were down-regulated in XDR S. Typhi. Further characterization of the identified proteins will help in understanding the molecular signaling involved in the emergence of XDR S. Typhi. Full article

Here, we report on the emergence and spread of multidrug-resistant NDM-1-producing P. aeruginosa isolates from patients hospitalized in the Attica region, Greece, in 2022 to provide data on their resistome, their virulome, the genetic environment of bla_NDM-1, and their molecular epidemiology. A total of 17 carbapenem-resistant P. aeruginosa isolates identified as NDM-producers by immunochromatography at the hospital level were sent to the Central Public Health Laboratory, in the frame of the laboratory surveillance of carbapenem-resistant pathogens, for further characterization. The initial screening for genetic AMR determinants was carried out by PCR and the MDR Direct Flow Chip assay. Typing was performed by MLST and DLST, the latter in a subset of isolates. Further analysis was performed by whole-genome sequencing (WGS) of six isolates from both hospitals to analyze their entire genomes and elucidate their genetic relatedness. All isolates were allocated to international high-risk clones, sixteen to ST773 and one to ST308. Five ST773 and the sole ST308 isolate were found to harbor the bla_NDM-1 gene, along with various other ARGs integrated into their chromosomes, as well as with a wide variety of virulence genes. The bla_NDM-1 gene was located in the integrative and conjugative elements ICE6600-like and ICE_Tn43716385 in ST773 and ST308 isolates, respectively. Single-nucleotide polymorphism analysis of the five ST773 isolates indicated their clonal spread in both hospitals. These results suggested that two different molecular events contributed to the emergence of NDM-1-producing P. aeruginosa isolates in Athenian hospitals, highlighting the need for ongoing surveillance. Full article

Antibiotic resistance is a global health problem. The human gut microbiome is implicated in the dynamics of antibiotic resistance acquisition and transmission, with the gut microbiota thought to play a crucial role. This study aimed to determine the potential influence of the human gut bacteria microbiota on the gut resistome and the relationship between the gut microbiota and Escherichia coli resistome. The PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guideline was used to systematically review studies that characterized the gut microbiota and resistome using metagenomic analysis and/or those that reported gut E. coli resistome in healthy individuals. Changes in the diversity and abundance of the bacterial gut microbiota and the resistome across different time points and participant groups were summarized. Additionally, using E. coli resistome as a proxy for the gut resistome, the microbiota composition of the gut harboring antibiotic-resistant E. coli was examined. The findings suggest that lower bacterial microbiota diversity is likely associated with an increased abundance of the overall gut resistome. Age-related differences were observed, with younger infants exhibiting lower microbiota diversity and higher antibiotic resistance gene (ARG) abundance compared to older infants and adults. Studies that reported positive correlations between the relative abundance of Proteobacteria and ARGs were mainly driven by members within the Enterobacteriaceae family, mainly E. coli. This study also reveals that human gut microbiome studies investigating the gut resistome using metagenomic sequencing approaches in healthy individuals are uncommon. Full article

Antimicrobial resistance (AMR) is a growing global health challenge, compromising bacterial infection treatments and necessitating robust surveillance and mitigation strategies. The overuse of antimicrobials in humans and farm animals has made them hotspots for AMR. However, the spread of AMR genes in wildlife and the environment represents an additional challenge, turning these areas into new AMR hotspots. Among the AMR bacteria considered to be of high concern for public health, Campylobacter has been the leading cause of foodborne infections in the European Union since 2005. This study examines the prevalence of AMR genes and virulence factors in Campylobacter isolates from wild birds and surface waters in Luxembourg. The findings reveal a significant prevalence of resistant Campylobacter strains, with 12% of C. jejuni from wild birds and 37% of C. coli from surface waters carrying resistance genes, mainly against key antibiotics like quinolones and tetracycline. This study underscores the crucial role of the environment in the spread of AMR bacteria and genes, highlighting the urgent need for enhanced surveillance and control measures to curb AMR in wildlife and environmental reservoirs and reduce transmission risks to humans. This research supports One Health approaches to tackling antimicrobial resistance and protecting human, animal, and environmental health. Full article

Antibiotics and antibiotic growth promoters have been extensively employed in poultry farming to enhance growth performance, maintain bird health, improve nutrient uptake efficiency, and mitigate enteric diseases at both sub-therapeutic and therapeutic doses. However, the extensive use of antimicrobials in poultry farming has led to the emergence of antimicrobial resistance (AMR) in microbial reservoirs, representing a significant global public health concern. In response, non-antibiotic dietary interventions, such as yeast mannan-rich fraction (MRF), have emerged as a promising alternative to modulate the gut microbiota and combat the AMR crisis. This study investigated whether a yeast mannan-rich fraction containing feed supplement impacted the performance of laying hens, their microbiomes, and the associated carriage of antimicrobial resistance genes under commercial conditions. High-throughput DNA sequencing was utilised to profile the bacterial community and assess changes in the antibiotic resistance genomes detected in the metagenome, the “resistome”, in response to MRF supplementation. It was found that supplementation favourably influenced laying hen performance and microbial composition. Notably, there was a compositional shift in the MRF supplemented group associated with a lower relative abundance of pathobionts, e.g., Escherichia, Brachyspira and Trueperella, and their AMR-encoded genes, relative to beneficial microbes. Overall, the findings further demonstrate the ability of prebiotics to improve laying hen performance through changes associated with their microbiome and resistome. Full article

The global antibiotic resistance crisis, driven by overuse and misuse of antibiotics, is multifaceted. This study aimed to assess the microbiological and genetic characteristics of raw retail pork meat through various methods, including the isolation, antibiotic susceptibility testing (AST), whole-genome sequencing (WGS) of selected indicator bacteria, antibiotic residue testing, and metagenomic sequencing. Samples were purchased from 10 pre-selected retail stores in Gauteng, South Africa. The samples were aseptically separated, with portions sent to an external laboratory for isolating indicator bacteria and testing for antibiotic residues. Identification of the isolated bacteria was reconfirmed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). AST was performed using the Microscan Walkaway system (Beckman Coulter, Brea, CA, USA). WGS and metagenomic sequencing were performed using the Illumina NextSeq 550 instrument (San Diego, CA, USA). The isolated E. coli and E. faecalis exhibited minimal phenotypic resistance, with WGS revealing the presence of tetracycline resistance genes. Both the isolated bacteria and meat samples harboured tetracycline resistance genes and the antibiotic residue concentrations were within acceptable limits for human consumption. In the metagenomic context, most identified bacteria were of food/meat spoilage and environmental origin. The resistome analysis primarily indicated beta-lactam, tetracycline and multidrug resistance genes. Further research is needed to understand the broader implications of these findings on environmental health and antibiotic resistance. Full article

The objective of this investigation is to ascertain the distinctive profile of the rhizospheric soil resistome within the Mecca region, while also evaluating the potential risks associated with the horizontal transfer of resistome determinants to the open environment and human clinical isolates. We have made metagenomic whole-genome shotgun sequencing for rhizospheric microbiomes of two endemic plants, namely Moringa oleifera and Abutilon fruticosum. The rhizospheric resistomes of the two plants and the abundance of antibiotic resistance genes (ARGs) were identified by cross-referencing encoded proteins with the comprehensive antibiotic resistance database (CARD). The identified ARGs were then analyzed for their antimicrobial resistance (AMR) mechanisms. Predominantly within this soil are the two bacterial species Pseudomonas aeruginosa and Mycobacterium tuberculosis. These opportunistic human pathogens are implicated in respiratory infections and are correlated with heightened mortality rates. The most prevalent array of ARGs existing in this soil comprises mexA, mexC, mexE, and cpxR, associated with mechanisms of antibiotic active efflux, along with ACC(2), ACC(3), AAC(6), and APH(6), in addition to arr1, arr3, arr4, iri, rphA, and rphB, implicated in antibiotic inactivation. Furthermore, vanS, vanR, and vanJ are identified for antibiotic target alteration, while rpoB2 and RbpA are noted for antibiotic target replacement and protection, respectively. These mechanisms confer resistance against a diverse spectrum of drug classes encompassing fluoroquinolones, aminoglycosides, glycopeptides, and rifampicins. This study underscores the potential hazards posed to human health by the presence of these pathogenic bacteria within the rhizospheric soil of the Mecca region, particularly in scenarios where novel ARGs prevalent in human populations are harbored and subsequently transmitted through the food chain to human clinical isolates. Consequently, stringent adherence to good agricultural and food transportation practices is imperative, particularly with regard to edible plant parts and those utilized in folkloric medicine. Full article

Antibiotic Resistance Genes (ARGs) are contaminants of emerging concern with marked potential to impact public and environmental health. This review focusses on factors that influence the presence, abundance, and dissemination of ARGs within Wastewater Treatment Plants (WWTPs) and associated effluents. Antibiotic-Resistant Bacteria (ARB) and ARGs have been detected in the influent and the effluent of WWTPs worldwide. Different levels of wastewater treatment (primary, secondary, and tertiary) show different degrees of removal efficiency of ARGs, with further differences being observed when ARGs are captured as intracellular or extracellular forms. Furthermore, routinely used molecular methodologies such as quantitative polymerase chain reaction or whole genome sequencing may also vary in resistome identification and in quantifying ARG removal efficiencies from WWTP effluents. Additionally, we provide an overview of the One Health risk assessment framework, as well as future strategies on how WWTPs can be assessed for environmental and public health impact. Full article

This study aimed to assess the suitability of fermented food interventions to replenish damaged gut microbiota. Metagenomic assessment of published sequencing data found that fermented food interventions led to a significant increase in the gut antibiotic resistome in healthy human subjects. Antibiotic resistome and viable antibiotic-resistant (AR) bacteria were further highly prevalent in retail kimchi and artisan cheeses by metagenomic and culture analyses. Representative AR pathogens of importance in nosocomial infections, such as Klebsiella pneumoniae, Serratia marcescens, and vancomycin-resistant Enterococcus (VRE), as well as commensals and lactic acid bacteria, were characterized; some exhibited an extremely high minimum inhibitory concentration (MIC) against antibiotics of clinical significance. Exposing fermented food microbiota to representative antibiotics further led to a boost of the corresponding antibiotic and multidrug-resistance gene pools, as well as disturbed microbiota, including the rise of previously undetectable pathogens. These results revealed an underestimated public health risk associated with fermented food intervention at the current stage, particularly for susceptible populations with compromised gut integrity and immune functions seeking gut microbiota rescue. The findings call for productive intervention of foodborne AR via technology innovation and strategic movements to mitigate unnecessary, massive damages to the host gut microbiota due to orally administered or biliary excreted antibiotics. Full article

Probiotics are widely used in broiler chickens to support the gut microbiome, gut health, and to reduce the amount of antibiotics used. Despite their benefits, there is concern over their ability to carry and spread antimicrobial resistance genes (ARGs), posing a significant public health risk. This study utilized next-generation sequencing to investigate ARGs in probiotics approved for poultry, focusing on their potential to be transferred via mobile genetic elements such as plasmids and phages. We examined the gut microbiome and resistome changes in 60 broiler chickens over their rearing period, correlating these changes with different probiotic treatments. Specific resistance mechanisms against critically important antibiotics were identified, including genes related to fluoroquinolone resistance and peptide antibiotic resistance. We also found genes with significant relevance to public health (aadK, AAC(6′)-Ii) and multiple drug-resistance genes (vmlR, ykkC, ykkD, msrC, clbA, eatAv). Only one phage-encoded gene (dfrA43) was detected, with no evidence of plasmid or mobile genetic element transmission. Additionally, metagenomic analysis of fecal samples showed no significant changes corresponding to time or diet across groups. Our findings highlight the potential risks associated with the use of probiotics in poultry, particularly regarding the carriage of ARGs. It is crucial to conduct further research into the molecular genetics of probiotics to develop strategies that mitigate the risk of resistance gene transfer in agriculture, ensuring the safe and effective use of probiotics in animal husbandry. Full article

Waste Water Treatment Plants (WWTP) aim to reduce contamination in effluent water; however, studies indicate antimicrobial resistance genes (ARGs) persist post-treatment, potentially leading to their spread from human populated areas into the environment. This study evaluated the impact of a large WWTP serving 125,000 people on the Iskar River in Bulgaria, by characterizing the spatial and short-term temporal dynamics in bacterial community dynamics and resistance profiles of the surface water. Pairs of samples were collected biweekly on four dates from two different locations, one about 800 m after the WWTP effluents and the other 10 km downstream. Taxonomic classification revealed the dominance of Pseudomonodota and Bacteriodota, notably the genera Flavobacterium, Aquirufa, Acidovorax, Polynucleobacter, and Limnohabitans. The taxonomic structure corresponded with both lentic and lotic freshwater habitats, with Flavobacterium exhibiting a significant decrease over the study period. Principal Coordinate Analysis revealed statistically significant differences in bacterial community composition between samples collected on different dates. Differential abundance analysis identified notable enrichment of Polynucleobacter and Limnohabitans. There were shifts within the enriched or depleted bacterial taxa between early and late sampling dates. High relative abundance of the genes erm(B), erm(F), mph(E), msr(E) (macrolides); tet(C), tet(O), tet(W), tet(Q) and tet(X) (tetracyclines); sul1 and sul2 (sulphonamides); and cfxA3, cfxA6 (beta-lactams) were detected, with trends of increased presence in the latest sampling dates and in the location closer to the WWTP. Of note, genes conferring resistance to carbapenems blaOXA-58 and blaIMP-33-like were identified. Co-occurrence analysis of ARGs and mobile genetic elements on putative plasmids showed few instances, and the estimated human health risk score (0.19) according to MetaCompare2.0 was low. In total, 29 metagenome-assembled genomes were recovered, with only a few harbouring ARGs. This study enhances our understanding of freshwater microbial community dynamics and antibiotic resistance profiles, highlighting the need for continued ARGs monitoring. Full article

The metagenomic surveillance of antimicrobial resistance in wastewater has been suggested as a methodological tool to characterize the distribution, status, and trends of antibiotic-resistant bacteria. In this study, a cross-sectional collection of samples of hospital-associated raw and treated wastewater were obtained from February to March 2020. Shotgun metagenomic sequencing and bioinformatic analysis were performed to characterize bacterial abundance and antimicrobial resistance gene analysis. The main bacterial phyla found in all the samples were as follows: Proteobacteria, Bacteroides, Firmicutes, and Actinobacteria. At the species level, ESKAPEE bacteria such as E. coli relative abundance decreased between raw and treated wastewater, but S. aureus, A. baumannii, and P. aeruginosa increased, as did the persistence of K. pneumoniae in both raw and treated wastewater. A total of 172 different ARGs were detected; bla_OXA, bla_VEB, bla_KPC, bla_GES, mphE, mef, erm, msrE, AAC(6′), ant(3″), aadS, lnu, PBP-2, dfrA, vanA-G, tet, and sul were found at the highest abundance and persistence. This study demonstrates the ability of ESKAPEE bacteria to survive tertiary treatment processes of hospital wastewater, as well as the persistence of clinically important antimicrobial resistance genes that are spreading in the environment. Full article

Freshwater environments are primary receiving systems of wastewater and effluents, which carry low concentrations of antibiotics and antimicrobial-resistant (AMR) bacteria and genes. Aquatic microbial communities are thus exposed to environmentally relevant concentrations of antibiotics (ERCA) that presumably influence the acquisition and spread of environmental AMR. Here, we analyzed ERCA exposure with and without the additional presence of municipal wastewater treatment plant effluent (W) and swine manure run-off (M) on aquatic biofilm resistomes. Microscopic analyses revealed decreased taxonomic diversity and biofilm structural integrity, while metagenomic analysis revealed an increased abundance of resistance, virulence, and mobile element-related genes at the highest ERCA exposure levels, with less notable impacts observed when solely exposed to W or M effluents. Microbial function predictions indicated increased gene abundance associated with energy and cell membrane metabolism and heavy metal resistance under ERCA conditions. In silico predictions of increased resistance mechanisms did not correlate with observed phenotypic resistance patterns when whole communities were exposed to antimicrobial susceptibility testing. This reveals important insight into the complexity of whole-community coordination of physical and genetic responses to selective pressures. Lastly, the environmental AMR risk assessment of metagenomic data revealed a higher risk score for biofilms grown at sub-MIC antibiotic conditions. Full article

The use of reclaimed wastewater to irrigate crops is a valuable option due to water scarcity. However, the presence of antibiotics residues that are not removed in the tertiary treatments reaches crop soils when irrigated and it poses a serious concern for human health. Crops rhizosphere is considered a hotspot of antibiotic resistant genes (ARG) being in addition a link to plant phyllosphere and human microbiome. Understanding the structure of the soil microbiota is crucial before applying any bioremediation or biostimulation strategy. The aim of this work was firstly to confirm the presence of antibiotics residues in soil and fruits in two greenhouses at the south of Spain irrigated with reclaimed water. Secondly, to characterize the rhizosphere microbiome of three crops (Capsicum annuum, Cucumis melo and Solanum melongena) cultured in those greenhouses. Finally, a predictive functional analysis was done using PICRUSt2 to figure out the rhizosphere resistome. The presence of residues of antibiotics was confirmed both soil and fruits. Antibiotics absorbed by plants correlated with those on soil. The most abundant resistance gene was the multidrug in all the three crops tested. Compatibility of basidiomycete fungi (i.e., Pleurotus) with soil bacteria could be indicative of their possible use for restoration of agricultural soils polluted with antibiotics. Full article