A single amplified genome catalog reveals the dynamics of mobilome and resistome in the human microbiome.

BACKGROUND: The increase in metagenome-assembled genomes (MAGs) has advanced our understanding of the functional characterization and taxonomic assignment within the human microbiome. However, MAGs, as population consensus genomes, often aggregate heterogeneity among species and strains, thereby obfuscating the precise relationships between microbial hosts and mobile genetic elements (MGEs). In contrast, single amplified genomes (SAGs) derived via single-cell genome sequencing can capture individual genomic content, including MGEs. RESULTS: We introduce the first substantial SAG dataset (bbsag20) from the human oral and gut microbiome, comprising 17,202 SAGs above medium-quality without co-assembly. This collection unveils a diversity of bacterial lineages across 312 oral and 647 gut species, demonstrating different taxonomic compositions from MAGs. Moreover, the SAGs showed cellular-level evidence of the translocation of oral bacteria to the gut. We also identified broad-host-range MGEs harboring antibiotic resistance genes (ARGs), which were not detected in the MAGs. CONCLUSIONS: The difference in taxonomic composition between SAGs and MAGs indicates that combining both methods would be effective in expanding the genome catalog. By connecting mobilomes and resistomes in individual samples, SAGs could meticulously chart a dynamic network of ARGs on MGEs, pinpointing potential ARG reservoirs and their spreading patterns in the microbial community. Video Abstract.

Intragenomic conflicts with plasmids and chromosomal mobile genetic elements drive the evolution of natural transformation within species.

Natural transformation is the only mechanism of genetic exchange controlled by the recipient bacteria. We quantified its rates in 786 clinical strains of the human pathogens Legionella pneumophila (Lp) and 496 clinical and environmental strains of Acinetobacter baumannii (Ab). The analysis of transformation rates in the light of phylogeny revealed they evolve by a mixture of frequent small changes and a few large quick jumps across 6 orders of magnitude. In standard conditions close to half of the strains of Lp and a more than a third in Ab are below the detection limit and thus presumably non-transformable. Ab environmental strains tend to have higher transformation rates than the clinical ones. Transitions to non-transformability were frequent and usually recent, suggesting that they are deleterious and subsequently purged by natural selection. Accordingly, we find that transformation decreases genetic linkage in both species, which might accelerate adaptation. Intragenomic conflicts with chromosomal mobile genetic elements (MGEs) and plasmids could explain these transitions and a GWAS confirmed systematic negative associations between transformation and MGEs: plasmids and other conjugative elements in Lp, prophages in Ab, and transposable elements in both. In accordance with the hypothesis of modulation of transformation rates by genetic conflicts, transformable strains have fewer MGEs in both species and some MGEs inactivate genes implicated in the transformation with heterologous DNA (in Ab). Innate defense systems against MGEs are associated with lower transformation rates, especially restriction-modification systems. In contrast, CRISPR-Cas systems are associated with higher transformation rates suggesting that adaptive defense systems may facilitate cell protection from MGEs while preserving genetic exchanges by natural transformation. Ab and Lp have different lifestyles, gene repertoires, and population structure. Nevertheless, they exhibit similar trends in terms of variation of transformation rates and its determinants, suggesting that genetic conflicts could drive the evolution of natural transformation in many bacteria.

Department-specific patterns of bacterial communities and antibiotic resistance in hospital indoor environments.

The hospital indoor environment has a crucial impact on the microbial exposures that humans encounter. Resistance to antibiotics is a mechanism used by bacteria to develop resilience in indoor environments, and the widespread use of antibiotics has led to changes in the ecological function of resistance genes and their acquisition by pathogens. By integrating the 16S rRNA Illumina sequencing and high-throughput-quantitative PCR approaches with water and air dust samples across seven departments in Peking University Shenzhen Hospital, China, this study yields intriguing findings regarding the department-specific variations, correlations and source tracing of bacteria, antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) within the hospital indoor environment. A notable observation was the pivotal role played by seasonal variations in shaping the bacterial composition across the entire hospital indoor environment. Another department-specific finding was the correlation between ARGs and MGEs abundance, which was evident in the overall hospital indoor environment, but not found in the blood test room, ophthalmology, and gynecology departments. Notably, as an important source of bacteria and ARGs/MGEs for the blood test room, the gynecology department also presented a close link between bacterial communities and the presence of ARGs/MGEs. Additionally, the results reiterate the importance of surveillance and monitoring of antibiotic resistance, specifically in Legionella spp. in man-made water systems, and highlight the significance of understanding genetic elements like Tp614 involved in gene transfer and recombination, and their impact on antimicrobial treatment efficacy. KEY POINTS: • The department-specific variations, correlations and source tracing of bacteria, ARGs, and MGEs were uncovered in the hospital's indoor environment. • Although each department exhibited consistent seasonal impacts on bacterial compositions, the co-occurrence between the presence of ARGs and MGEs was exclusively evident in the emergency, surgery, pneumology and otolaryngology departments. • The gynecology department emerged as a crucial source of bacteria, ARGs and MGEs within the hospital. Additionally, it was found to exhibit a significant correlation between bacterial communities and the presence of ARGs and MGEs.

Contribution of the Mobilome to the Configuration of the Resistome of Corynebacterium striatum.

Corynebacterium striatum, present in the microbiota of human skin and nasal mucosa, has recently emerged as a causative agent of hospital-acquired infections, notable for its resistance to multiple antimicrobials. Its mobilome comprises several mobile genetic elements, such as plasmids, transposons, insertion sequences and integrons, which contribute to the acquisition of antimicrobial resistance genes. This study analyzes the contribution of the C. striatum mobilome in the transfer and dissemination of resistance genes. In addition, integrative and conjugative elements (ICEs), essential in the dissemination of resistance genes between bacterial populations, whose role in C. striatum has not yet been studied, are examined. This study examined 365 C. striatum genomes obtained from the NCBI Pathogen Detection database. Phylogenetic and pangenome analyses were performed, the resistance profile of the bacterium was recognized, and mobile elements, including putative ICE, were detected. Bioinformatic analyses identified 20 antimicrobial resistance genes in this species, with the Ermx gene being the most predominant. Resistance genes were mainly associated with plasmid sequence regions and class 1 integrons. Although an ICE was detected, no resistance genes linked to this element were found. This study provided valuable information on the geographic spread and prevalence of outbreaks observed through phylogenetic and pangenome analyses, along with identifying antimicrobial resistance genes and mobile genetic elements that carry many of the resistance genes and may be the subject of future research and therapeutic approaches.

The emergence of metronidazole-resistant Prevotella bivia harboring nimK gene in Japan.

We present the identification and characterization of the complete genome of metronidazole (MTZ)-resistant Prevotella bivia strain TOH-2715 [minimum inhibitory concentration (MIC): 8 mg/L], isolated from the urine of an elderly Japanese woman, as well as details of its mobile genetic elements (MGEs) containing antimicrobial resistance (AMR) genes and its relationship with other bacterial species determined using whole-genome sequencing (WGS) data. TOH-2715 possessed two chromosomes with putative MGEs containing AMR genes. Two AMR-related MGE regions were present in chromosome 2. MGE-region 1 (7,821 bp) included Tn6456, where nimK was located, and MGE-region 2 (58.8 Kbp) included the integrative and conjugative element (ICE), where tet(Q) and ermF were located. The genetic structure of the ICE of TOH-2715 was similar to that of CTnDOT-family transposons, where ermF and tet(Q) are located. A search of public databases revealed that nimK was present in Prevotella spp., including P. bivia, and was partially composed of a Tn6456-like element lacking the efflux transporter gene qacE and the Crp/Fnr family transcriptional regulator gene in some cases. Core ICE gene analysis showed that ICEs similar to that of TOH-2715 were present in Prevotella spp. and Bacteroides spp., suggesting horizontal gene transfer among anaerobes. This is the report of WGS analysis of an MTZ-resistant clinical strain of P. bivia (TOH-2715) with Tn6456 encoding nimK. Other submitted genomes have described the presence of nimK, but none of them have described MTZ resistance. Additionally, we described putative MGE regions containing the AMR gene within the genus Prevotella and among anaerobes, raising concerns about the future spread of nimK among anaerobes. IMPORTANCE: Metronidazole (MTZ) is an important antimicrobial agent in anaerobic infections and is widely used in clinical settings. The rate of MTZ resistance in anaerobic bacteria has been increasing in recent years, and the nim gene (nitro-imidazole reductase) is one of the resistance mechanisms. Prevotella bivia is found in humans in the urinary tract and vagina and is known to cause infections in some cases. One of the nim genes, nimK, has recently been discovered in this species of bacteria, but there are no reports of antimicrobial resistance (AMR)-related regions in its whole genome level. In this study, we analyzed the AMR region of nimK-positive P. bivia derived from clinical specimens based on comparisons with other anaerobic genomes. P. bivia was found to be engaged in horizontal gene transfer with other anaerobic bacteria, and the future spread of the nimK gene is a concern.

Genomic analysis of antibiotic resistance genes and mobile genetic elements in eight strains of nontyphoid Salmonella.

Nontyphoidal Salmonella (NTS) is the main etiological agent of human nontyphoidal salmonellosis. The aim of this study was to analyze the epidemiological characteristics and horizontal transfer mechanisms of antimicrobial resistance (AMR) genes from eight strains of NTS detected in Zhenjiang City, Jiangsu Province, China. Fecal samples from outpatients with food-borne diarrhea were collected in 2022. The NTS isolates were identified, and their susceptibility was tested with the Vitek 2 Compact system. The genomes of the NTS isolates were sequenced with the Illumina NovaSeq platform and Oxford Nanopore Technologies platform. The AMR genes and mobile genetic elements (MGEs) were predicted with the relevant open access resources. Eight strains of NTS were isolated from 153 specimens, and Salmonella Typhimurium ST19 was the most prevalent serotype. The AMR gene with the highest detection rate was AAC(6')-Iaa (10.5%) followed by TEM-1 (7.9%), sul2 (6.6%), and tet(A) (5.3%). Eleven MGEs carrying 34 AMR genes were identified on the chromosomes of 3 of the 8 NTS, including 3 resistance islands, 6 composite transposons (Tns), and 2 integrons. Eighteen plasmids carrying 40 AMR genes were detected in the 8 NTS strains, including 6 mobilizable plasmids, 3 conjugative plasmids, and 9 nontransferable plasmids, 7 of which carried 10 composite Tns and 3 integrons. This study provided a theoretical basis, from a genetic perspective, for the prevention and control of NTS resistance in Zhenjiang City. IMPORTANCE: Human nontyphoidal salmonellosis is one of the common causes of bacterial food-borne illnesses, with significant social and economic impacts, especially those caused by invasive multidrug-resistant nontyphoidal Salmonella, which entails high morbidity and mortality. Antimicrobial resistance is mainly mediated by drug resistance genes, and mobile genetic elements play key roles in the capture, accumulation, and dissemination of antimicrobial resistance genes. Therefore, it is necessary to study the epidemiological characteristics and horizontal transfer mechanisms of antimicrobial resistance genes of nontyphoidal Salmonella to prevent the spread of multidrug-resistant nontyphoidal Salmonella.

Hijackers, hitchhikers, or co-drivers? The mysteries of mobilizable genetic elements.

Mobile genetic elements shape microbial gene repertoires and populations. Recent results reveal that many, possibly most, microbial mobile genetic elements require helpers to transfer between genomes, which we refer to as Hitcher Genetic Elements (hitchers or HGEs). They may be a large fraction of pathogenicity and resistance genomic islands, whose mechanisms of transfer have remained enigmatic for decades. Together with their helper elements and their bacterial hosts, hitchers form tripartite networks of interactions that evolve rapidly within a parasitism-mutualism continuum. In this emerging view of microbial genomes as communities of mobile genetic elements many questions arise. Which elements are being moved, by whom, and how? How often are hitchers costly hyper-parasites or beneficial mutualists? What is the evolutionary origin of hitchers? Are there key advantages associated with hitchers' lifestyle that justify their unexpected abundance? And why are hitchers systematically smaller than their helpers? In this essay, we start answering these questions and point ways ahead for understanding the principles, origin, mechanisms, and impact of hitchers in bacterial ecology and evolution.

A metagenomics-based approach to decipher the resistome and mobilome of two seahorse species, Hippocampus barbouri and Hippocampus comes.

OBJECTIVE: This study aimed to explore the abundance and diversity of antibiotic resistance genes (ARGs) in seahorses (Hippocampus barbouri and Hippocampus comes) and their surrounding environment. METHODS: A combination of shotgun metagenomics and bioinformatics was used to investigate the resistome of both seahorse species. RESULTS: The analyses demonstrated a higher abundance of ARGs in seahorse-associated microbiomes, particularly in skin and gut samples, compared to those from water and sediment. Interestingly, genes conferring multidrug resistance (e.g., acrB, acrF, cpxA, msbA, and oqxB) were highly prevalent in all samples, especially in skin and gut samples. High levels of genes conferring resistance to fluoroquinolones (e.g., mfd and emrB), ß-lactam (e.g., blaCMY-71, blaOXA-55, and penA), aminocoumarin (e.g., mdtB and mdtC), and peptide antibiotics (arnA, pmrE, and rosA) were also observed in skin and gut samples. An enrichment of mobile genetic elements (MGEs) was also observed in the analysed samples, highlighting their potential role in facilitating the acquisition and spread of ARGs. In fact, the abundance of mobilisation (MOB) relaxases (e.g., MOBF, MOBP, MOBT, and MOBV) in gut and skin samples suggests a high potential for conjugation events. CONCLUSIONS: The occurrence of ARGs and MGEs in seahorses and the surrounding environment raises concerns about their transmission to humans, either through direct contact or the consumption of contaminated seafood. To the best of our knowledge, this study represents the first comprehensive analysis of ARGs in seahorse-associated microbiomes, and its results emphasise the need for monitoring and controlling the spread of ARGs in environmental settings.

The lactococcal ICE-ome encodes a repertoire of exchangeable traits with potential industrial relevance.

Dairy industries apply selected lactococcal strains and mixed cultures to produce diverse fermented products with distinctive flavor and texture properties. Innovation of the starter culture functionality in cheese applications embraces natural biodiversity of the Lactococcus species to identify novel strains with alternative flavor or texture forming capacities and/or increased processing robustness and phage resistance. Mobile genetic elements (MGE), like integrative conjugative elements (ICEs) play an important role in shaping the biodiversity of bacteria. Besides the genes involved in the conjugation of ICEs from donor to recipient strains, these elements also harbor cargo genes that encode a wide range of functions. The definition of such cargo genes can only be achieved by accurate identification of the ICE boundaries (delimiting). Here, we delimited 25 ICEs in lactococcal genome sequences with low contig numbers using insertion-sites flanking single-copy core-genome genes as markers for each of the distinct ICE-integrases we identified previously within the conserved ICE-core genes. For ICEs in strains for which genome information with large numbers of contigs is available, we exemplify that CRISPR-Cas9 driven ICE-curing, followed by resequencing, allows accurate delimitation and cargo definition of ICEs. Finally, we compare and contrast the cargo gene repertoire of the 26 delimited lactococcal ICEs, identifying high plasticity among the cargo of lactococccal ICEs and a range of encoded functions that is of apparent industrial interest, including restriction modification, abortive infection, and stress adaptation genes.

Emergence of a clinical Klebsiella pneumoniae harboring an acrAB-tolC in chromosome and carrying the two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65 in a plasmid.

Objective: The emergence of clinical Klebsiella pneumoniae strains harboring acrAB-tolC genes in the chromosome, along with the presence of two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65 genes on a plasmid, has presented a significant clinical challenge. Methods: In order to study the detailed genetic features of K. pneumoniae strain SC35, both the bacterial chromosome and plasmids were sequenced using Illumina and nanopore platforms. Furthermore, bioinformatics methods were employed to analyze the mobile genetic elements associated with antibiotic resistance genes. Results: K. pneumoniae strain SC35 was found to possess a class A beta-lactamase and demonstrated resistance to all tested antibiotics. This resistance was attributed to the presence of efflux pump genes, specifically acrAB-tolC, on the SC35 chromosome. Additionally, the SC35 plasmid p1 carried the two repetitive tandem core structures for bla KPC-2 and bla CTX-M-65, as well as bla TEM-1 with rmtB, which shared overlapping structures with mobile genetic elements as In413, Tn3, and TnAs3. Through plasmid transfer assays, it was determined that the SC35 plasmid p1 could be successfully transferred with an average conjugation frequency of 6.85 × 10-4. Conclusion: The structure of the SC35 plasmid p1 appears to have evolved in correlation with other plasmids such as pKPC2_130119, pDD01754-2, and F4_plasmid pA. The infectious strain SC35 exhibits no susceptibility to tested antibioticst, thus effective measures should be taken to prevent the spread and epidemic of this strain.

Mobile genetic elements define the non-random structure of the Salmonella enterica serovar Typhi pangenome.

Bacterial relatedness measured using select chromosomal loci forms the basis of public health genomic surveillance. While approximating vertical evolution through this approach has proven exceptionally valuable for understanding pathogen dynamics, it excludes a fundamental dimension of bacterial evolution-horizontal gene transfer. Incorporating the accessory genome is the logical remediation and has recently shown promise in expanding epidemiological resolution for enteric pathogens. Employing k-mer-based Jaccard index analysis, and a novel genome length distance metric, we computed pangenome (i.e., core and accessory) relatedness for the globally important pathogen Salmonella enterica serotype Typhi (Typhi), and graphically express both vertical (homology-by-descent) and horizontal (homology-by-admixture) evolutionary relationships in a reticulate network of over 2,200 U.S. Typhi genomes. This analysis revealed non-random structure in the Typhi pangenome that is driven predominantly by the gain and loss of mobile genetic elements, confirming and expanding upon known epidemiological patterns, revealing novel plasmid dynamics, and identifying avenues for further genomic epidemiological exploration. With an eye to public health application, this work adds important biological context to the rapidly improving ways of analyzing bacterial genetic data and demonstrates the value of the accessory genome to infer pathogen epidemiology and evolution.IMPORTANCEGiven bacterial evolution occurs in both vertical and horizontal dimensions, inclusion of both core and accessory genetic material (i.e., the pangenome) is a logical step toward a more thorough understanding of pathogen dynamics. With an eye to public, and indeed, global health relevance, we couple contemporary tools for genomic analysis with decades of research on mobile genetic elements to demonstrate the value of the pangenome, known and unknown, annotated, and hypothetical, for stratification of Salmonella enterica serovar Typhi (Typhi) populations. We confirm and expand upon what is known about Typhi epidemiology, plasmids, and antimicrobial resistance dynamics, and offer new avenues of exploration to further deduce Typhi ecology and evolution, and ultimately to reduce the incidence of human disease.

Copy number variation introduced by a massive mobile element facilitates global thermal adaptation in a fungal wheat pathogen.

Copy number variation (CNV) can drive rapid evolution in changing environments. In microbial pathogens, such adaptation is a key factor underpinning epidemics and colonization of new niches. However, the genomic determinants of such adaptation remain poorly understood. Here, we systematically investigate CNVs in a large genome sequencing dataset spanning a worldwide collection of 1104 genomes from the major wheat pathogen Zymoseptoria tritici. We found overall strong purifying selection acting on most CNVs. Genomic defense mechanisms likely accelerated gene loss over episodes of continental colonization. Local adaptation along climatic gradients was likely facilitated by CNVs affecting secondary metabolite production and gene loss in general. One of the strongest loci for climatic adaptation is a highly conserved gene of the NAD-dependent Sirtuin family. The Sirtuin CNV locus localizes to an ~68-kb Starship mobile element unique to the species carrying genes highly expressed during plant infection. The element has likely lost the ability to transpose, demonstrating how the ongoing domestication of cargo-carrying selfish elements can contribute to selectable variation within populations. Our work highlights how standing variation in gene copy numbers at the global scale can be a major factor driving climatic and metabolic adaptation in microbial species.

A comparison of antibiotic resistance genes and mobile genetic elements in wild and captive Himalayan vultures.

As the most widely distributed scavenger birds on the Qinghai-Tibetan Plateau, Himalayan vultures (Gyps himalayensis) feed on the carcasses of various wild and domestic animals, facing the dual selection pressure of pathogens and antibiotics and are suitable biological sentinel species for monitoring antibiotic resistance genes (ARGs). This study used metagenomic sequencing to comparatively investigate the ARGs and mobile genetic elements (MGEs) of wild and captive Himalayan vultures. Overall, the resistome of Himalayan vultures contained 414 ARG subtypes resistant to 20 ARG types, with abundances ranging from 0.01 to 1,493.60 ppm. The most abundant resistance type was beta-lactam (175 subtypes), followed by multidrug resistance genes with 68 subtypes. Decreases in the abundance of macrolide-lincosamide-streptogramin (MLS) resistance genes were observed in the wild group compared with the zoo group. A total of 75 genera (five phyla) of bacteria were predicted to be the hosts of ARGs in Himalayan vultures, and the clinical (102 ARGs) and high-risk ARGs (35 Rank I and 56 Rank II ARGs) were also analyzed. Among these ARGs, twenty-two clinical ARGs, nine Rank I ARG subtypes, sixteen Rank II ARG subtypes were found to differ significantly between the two groups. Five types of MGEs (128 subtypes) were found in Himalayan vultures. Plasmids (62 subtypes) and transposases (44 subtypes) were found to be the main MGE types. Efflux pump and antibiotic deactivation were the main resistance mechanisms of ARGs in Himalayan vultures. Decreases in the abundance of cellular protection were identified in wild Himalayan vultures compared with the captive Himalayan vultures. Procrustes analysis and the co-occurrence networks analysis revealed different patterns of correlations among gut microbes, ARGs, and MGEs in wild and captive Himalayan vultures. This study is the first step in describing the characterization of the ARGs in the gut of Himalayan vultures and highlights the need to pay more attention to scavenging birds.

Spatial mapping of mobile genetic elements and their bacterial hosts in complex microbiomes.

The exchange of mobile genetic elements (MGEs) facilitates the spread of functional traits including antimicrobial resistance within bacterial communities. Tools to spatially map MGEs and identify their bacterial hosts in complex microbial communities are currently lacking, limiting our understanding of this process. Here we combined single-molecule DNA fluorescence in situ hybridization (FISH) with multiplexed ribosomal RNA-FISH to enable simultaneous visualization of both MGEs and bacterial taxa. We spatially mapped bacteriophage and antimicrobial resistance (AMR) plasmids and identified their host taxa in human oral biofilms. This revealed distinct clusters of AMR plasmids and prophage, coinciding with densely packed regions of host bacteria. Our data suggest spatial heterogeneity in bacterial taxa results in heterogeneous MGE distribution within the community, with MGE clusters resulting from horizontal gene transfer hotspots or expansion of MGE-carrying strains. Our approach can help advance the study of AMR and phage ecology in biofilms.

The mobilome of Staphylococcus aureus from wild ungulates reveals epidemiological links at the animal-human interface.

Staphylococcus aureus thrives at animal-human-environment interfaces. A large-scale work from our group indicated that antimicrobial resistance (AMR) in commensal S. aureus strains from wild ungulates is associated with agricultural land cover and livestock farming, raising the hypothesis that AMR genes in wildlife strains may originate from different hosts, namely via exchange of mobile genetic elements (MGE). In this work, we generate the largest available dataset of S. aureus draft genomes from wild ungulates in Portugal and explore their mobilome, which can determine important traits such as AMR, virulence, and host specificity, to understand MGE exchange. Core genome multi-locus sequence typing based on 98 newly generated draft genomes and 101 publicly available genomes from Portugal demonstrated that the genomic relatedness of S. aureus from wild ungulates assigned to livestock-associated sequence types (ST) is greater compared to wild ungulate isolates assigned to human-associated STs. Screening of host specificity determinants disclosed the unexpected presence in wildlife of the immune evasion cluster encoded in φSa3 prophage, described as a human-specific virulence determinant. Additionally, two plasmids, pAVX and pETB, previously associated with avian species and humans, respectively, and the Tn553 transposon were detected. Both pETB and Tn553 encode penicillin resistance through blaZ. Pangenome analysis of wild ungulate isolates shows a core genome fraction of 2133 genes, with isolates assigned to ST72 and ST3224 being distinguished from the remaining by MGEs, although there is no reported role of these in adaptation to wildlife. AMR related gene clusters found in the shell genome are directly linked to resistance against penicillin, macrolides, fosfomycin, and aminoglycosides, and they represent mobile ARGs. Altogether, our findings support epidemiological interactions of human and non-human hosts at interfaces, with MGE exchange, including AMR determinants, associated with putative indirect movements of S. aureus among human and wildlife hosts that might be bridged by livestock.

Chromosomal and plasmid localization of ileS2 in high-level mupirocin-resistant Staphylococcus pseudintermedius and Staphylococcus aureus isolated from canine and feline origins.

OBJECTIVES: To characterize the mobile genetic elements and genetic localization of ileS2 in high-level mupirocin-resistant (Hi-MupR) methicillin-resistant Staphylococcus pseudintermedius (MRSP) and MRSA isolates recovered from canine and feline clinical samples. METHODS: The identification of bacterial species and presence of mecA and ileS2 genes in MRSP and MRSA isolates were performed using MALDI-TOF MS and PCR, respectively. Antimicrobial resistance (AMR) phenotypes were determined by broth microdilution assays. The genome characteristics, ileS2-containing elements and staphylococcal cassette chromosome mec (SCCmec) were illustrated using complete circular genomes obtained from hybrid assembly of Illumina short-reads and Oxford Nanopore Technologies long-reads. These were analysed through phylogenetic and bioinformatics approaches. RESULTS: A total of 18 MRSP clinical isolates and four MRSA clinical isolates exhibited the Hi-MupR phenotype and carried multiple AMR genes, including mecA and ileS2 genes. MRSP ST182-SCCmec V (n = 6) and ST282-ΨSCCmec57395-t10 (n = 4) contained the ileS2 transposable unit associated with IS257 on the chromosome. Three MRSA ST398-SCCmec V-t034/t4652 isolates carried ∼42 kb pSK41-like ileS2 plasmids, whereas similar ileS2 plasmids lacking tra genes were found in MRSP ST282-ΨSCCmec57395-t72/t21 isolates. Furthermore, a new group of ileS2 plasmids, carried by MRSP ST45-ΨSCCmec57395, ST433-ΨSCCmecKW21-t05 and ST2165-SCCmec IV-t06, and by one MRSA ST398-SCCmec V-t034 strain, shared the plasmid backbone with the cfr/fexA-carrying plasmid pM084526_1 in MRSA ST398. CONCLUSIONS: This study provides the first evidence of ileS2 integration into the S. pseudintermedius chromosome, which is a rare occurrence in staphylococcal species, and plasmids played a pivotal role in dissemination of ileS2 in both staphylococcal species.

Characterization and genetic analysis of extensively drug-resistant hospital acquired Pseudomonas aeruginosa isolates.

BACKGROUND: The incidence of hospital-acquired infections in extensively drug-resistant Pseudomonas aeruginosa (XDR-PA) has been increasing worldwide and is frequently associated with an increase in mortality and morbidity rates. The aim of this study was to characterize clinical XDR-PA isolates recovered during six months at three different hospitals in Egypt. RESULTS: Seventy hospital-acquired clinical isolates of P. aeruginosa were classified into multidrug-resistant (MDR), extensively drug-resistant (XDR) and pandrug-resistant (PDR), according to their antimicrobial resistance profile. In addition, the possession of genes associated with mobile genetic elements and genes encoding antimicrobial resistance determinants among isolates were detected using polymerase chain reaction. As a result, a significant percentage of the isolates (75.7%) were XDR, while 18.5% were MDR, however only 5.7% of the isolates were non-MDR. The phenotypic detection of carbapenemases, extended-spectrum ß-lactamases (ESBLs) and metallo ß-lactamase (MBL) enzymes showed that 73.6% of XDR-PA isolates were carbapenemases producers, whereas 75.5% and 88.7% of XDR-PA isolates produced ESBLs and MBL respectively. In addition, PCR screening showed that oxa gene was the most frequently detected gene of carbapenemases (91.4%), while aac(6')-lb gene was mostly detected (84.3%) among the screened aminoglycosides-resistance genes. Furthermore, the molecular detection of the colistin resistance gene showed that 12.9% of isolates harbored mcr-1 gene. Concerning mobile genetic element markers (intI, traA, tnp513, and merA), intI was the highest detected gene as it was amplified in 67 isolates (95.7%). Finally, phylogenetic and molecular typing of the isolates via ERIC-PCR analysis revealed 10 different ERIC fingerprints. CONCLUSION: The present study revealed a high prevalence of XDR-PA in hospital settings which were resistant to a variety of antibiotics due to several mechanisms. In addition, 98% of the XDR-PA clinical isolates contained at least one gene associated with movable genetic elements, which could have aided the evolution of these XDR-PA strains. To reduce spread of drug resistance, judicious use of antimicrobial agents and strict infection control measures are therefore essential.

Detection and Quantification of Conjugative Transfer of Mobile Genetic Elements Carrying Antibiotic Resistance Genes.

Multidrug resistance, due to acquired antimicrobial resistance genes, is increasingly reported in the zoonotic pathogen Streptococcus suis. Most of these resistance genes are carried by chromosomal Mobile Genetic Elements (MGEs), in particular, Integrative and Conjugative Elements (ICEs) and Integrative and Mobilizable Elements (IMEs). ICEs and IMEs frequently form tandems or nested composite elements, which make their identification difficult. To evaluate their mobility, it is necessary to (i) select the suitable donor-recipient pairs for mating assays, (ii) do PCR excision tests to confirm that the genetic element is able to excise from the chromosome as a circular intermediate, and (iii) evaluate the transfer of the genetic element by conjugation by doing mating assays. In addition to a dissemination of resistance genes between S. suis strains, MGEs can lead to a spreading of resistance genes in the environment and toward pathogenic bacteria. This propagation had to be considered in a One Health perspective.

Exploration of mobile genetic elements in the ruminal microbiome of Nellore cattle.

Metagenomics has made it feasible to elucidate the intricacies of the ruminal microbiome and its role in the differentiation of animal production phenotypes of significance. The search for mobile genetic elements (MGEs) has taken on great importance, as they play a critical role in the transfer of genetic material between organisms. Furthermore, these elements serve a dual purpose by controlling populations through lytic bacteriophages, thereby maintaining ecological equilibrium and driving the evolutionary progress of host microorganisms. In this study, we aimed to identify the association between ruminal bacteria and their MGEs in Nellore cattle using physical chromosomal links through the Hi-C method. Shotgun metagenomic sequencing and the proximity ligation method ProxiMeta were used to analyze DNA, getting 1,713,111,307 bp, which gave rise to 107 metagenome-assembled genomes from rumen samples of four Nellore cows maintained on pasture. Taxonomic analysis revealed that most of the bacterial genomes belonged to the families Lachnospiraceae, Bacteroidaceae, Ruminococcaceae, Saccharofermentanaceae, and Treponemataceae and mostly encoded pathways for central carbon and other carbohydrate metabolisms. A total of 31 associations between host bacteria and MGE were identified, including 17 links to viruses and 14 links to plasmids. Additionally, we found 12 antibiotic resistance genes. To our knowledge, this is the first study in Brazilian cattle that connect MGEs with their microbial hosts. It identifies MGEs present in the rumen of pasture-raised Nellore cattle, offering insights that could advance biotechnology for food digestion and improve ruminant performance in production systems.