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Purpose: The emergence of carbapenem-resistant P. aeruginosa (CRPA) harbouring acquired carbapenemase genes (blaVIM, blaIMP and blaNDM) has become a global public health threat. Three CRPA isolates included in the study had an extensively drug-resistant phenotype with susceptibility to colistin only and were positive for the blaNDM-1 gene. The current study aimed to investigate the genomic epidemiology and molecular characteristics of the blaNDM-1-positive CRPA isolates collected from the Gauteng region, South Africa. Methods: Short read whole genome sequencing (WGS) was performed to determine sequence types (STs), genetic relatedness, resistome, virulome and the genetic environment of the blaNDM-1 gene. Results: The WGS and phylogenetic analyses revealed that the study isolates belonged to an international high-risk clone ST773 and belonged to the same clade with eight blaNDM-1-positive ST773 isolates from Hungary, India, Nigeria, South Korea and USA. The study isolates harboured a wide repertoire of intrinsic and acquired antibiotic resistance genes (ARGs) related with mobile genetic elements, porins and efflux pumps, as well as virulence factor genes. The clade-specific ARGs (blaNDM-1, floR2/cmlA9, rmtB4, tetG) were found in a putative integrative and conjugative element (ICE) region similar to ICE6660-like. Conclusion: As ICE carrying the blaNDM-1 gene can easily spread to other P. aeruginosa isolates and other Gram-negative bacteria, the findings in this study highlight the need for appropriate management strategies and active surveillance of CRPA isolates in the Gauteng region, South Africa. [ABSTRACT FROM AUTHOR]

Background: Escherichia coli, a ubiquitous inhabitant of the gut microbiota, has been recognized as an indicator of fecal contamination and a potential reservoir for antibiotic resistance genes. Its prevalence in drinking water sources raises concerns about the potential dissemination of antibiotic resistance within aquatic ecosystems and the subsequent impact on public health. The ability of E. coli to acquire and transfer resistance genes, coupled with the constant exposure to low levels of antibiotics in the environment, underscores the need for comprehensive surveillance and rigorous antimicrobial stewardship strategies to safeguard the quality and safety of drinking water supplies, ultimately mitigating the escalation of antibiotic resistance and its implications for human well-being. Methods: WG5D strain, isolated from a drinking water distribution source in North-West Province, South Africa, underwent genomic analysis following isolation on nutrient agar, anaerobic cultivation, and DNA extraction. Paired-end Illumina sequencing with a Nextera XT Library Preparation kit was performed. The assembly, annotation, and subsequent genomic analyses, including phylogenetic analysis using TYGS, pairwise comparisons, and determination of genes related to antimicrobial resistance and virulence, were carried out following standard protocols and tools, ensuring comprehensive insights into the strain's genomic features. Results: This study explores the notable characteristics of E. coli strain WG5D. This strain stands out because it possesses multiple antibiotic resistance genes, encompassing tetracycline, cephalosporin, vancomycin, and aminoglycoside resistances. Additionally, virulence-associated genes indicate potential heightened pathogenicity, complemented by the identification of mobile genetic elements that underscore its adaptability. The intriguing possibility of bacteriophage involvement and factors contributing to pathogenicity further enriches our understanding. We identified E. coli WG5D as a potential human pathogen associated with a drinking water source in South Africa. The analysis provided several antibiotic resistance-associated genes/mutations and mobile genetic elements. It further identified WG5D as a potential human pathogen. The occurrence of E. coli WG5D raised the awareness of the potential pathogens and the carrying of antibiotic resistance in drinking water. Conclusions: The findings of this study have highlighted the advantages of the genomic approach in identifying the bacterial species and antibiotic resistance genes of E. coli and its potential as a human pathogen. [ABSTRACT FROM AUTHOR]

Background: Citrobacter species are Gram-negative opportunistic pathogens commonly reported in nosocomial-acquired infections. This study characterised four Citrobacter species that were isolated from surface water in the North West Province, South Africa. Results: Phenotypic antimicrobial susceptibility profiles of the isolates demonstrated their ability to produce the extended-spectrum β-lactamase (ESBL). Whole genomes were sequenced to profile antibiotic resistance and virulence genes, as well as mobile genetic elements. In silico taxonomic identification was conducted by using multi-locus sequence typing and average nucleotide identity. A pangenome was used to determine the phylogenomic landscape of the Citrobacter species by using 109 publicly available genomes. The strains S21 and S23 were identified as C. braakii, while strains S24 and S25 were C. murliniae and C. portucalensis, respectively. Comparative genomics and sequenced genomes of the ESBL-producing isolates consisted of n = 91; 83% Citrobacter species in which bla-CMY−101 (n = 19; 32,2%) and bla-CMY−59 (n = 12; 38,7%) were prevalent in C. braakii, and C. portucalensis strains, respectively. Macrolide (acrAB-TolC, and mdtG) and aminoglycoside (acrD) efflux pumps genes were identified in the four sequenced Citrobacter spp. isolates. The quinolone resistance gene, qnrB13, was exclusive to the C. portucalensis S25 strain. In silico analysis detected plasmid replicon types IncHI1A, IncP, and Col(VCM04) in C. murliniae S24 and C. portucalensis S25, respectively. These potentially facilitate the T4SS secretion system in Citrobacter species. In this study, the C. braakii genomes could be distinguished from C. murliniae and C. portucalensis on the basis of gene encoding for cell surface localisation of the CPS (vexC) and identification of genes involved in capsule polymer synthesis (tviB and tviE). A cluster for the salmochelin siderophore system (iro-BCDEN) was found in C. murliniae S24. This is important when it comes to the pathogenicity pathway that confers an advantage in colonisation. Conclusions: The emerging and genomic landscapes of these ESBL-producing Citrobacter species are of significant concern due to their dissemination potential in freshwater systems. The presence of these ESBL and multidrug-resistant (MDR) pathogens in aquatic environments is of One Health importance, since they potentially impact the clinical domain, that is, in terms of human health and the agricultural domain, that is, in terms of animal health and food production as well as the environmental domain. [ABSTRACT FROM AUTHOR]

Enterococci are among the most common opportunistic hospital pathogens. -is study used whole-genome sequencing (WGS) and bioinformatics to determine the antibiotic resistome, mobile genetic elements, clone and phylogenetic relationship of Enterococcus faecalis isolated from hospital environments in South Africa. -is study was carried out from September to November 2017. Isolates were recovered from 11 frequently touched sites by patients and healthcare workers in di8erent wards at 4 levels of healthcare (A, B, C, and D) in Durban, South Africa. Out of the 245 identi:ed E. faecalis isolates, 38 isolates underwent whole-genome sequencing (WGS) on the Illumina MiSeq platform, following microbial identi:cation and antibiotic susceptibility tests. -e tet(M) (31/38, 82%) and erm(C) (16/38, 42%) genes were the most common antibiotic-resistant genes found in isolates originating from di8erent hospital environments which corroborated with their antibiotic resistance phenotypes. -e isolates harboured mobile genetic elements consisting of plasmids (n = 11) and prophages (n = 14) that were mostly clone-speci:c. Of note, a large number of insertion sequence (IS) families were found on the IS3 (55%), IS5 (42%), IS1595 (40%), and Tn3 transposons the most predominant. Microbial typing using WGS data revealed 15 clones with 6 major sequence types (ST) belonging to ST16 (n = 7), ST40 (n = 6), ST21 (n = 5), ST126 (n = 3), ST23 (n = 3), and ST386 (n = 3). Phylogenomic analysis showed that the major clones were mostly conserved within speci:c hospital environments. However, further metadata insights revealed the complex intraclonal spread of these E. faecalis major clones between the sampling sites within each speci:c hospital setting. -e results of these genomic analyses will o8er insights into antibioticresistantE. faecalis in hospital environments relevant to the design of optimal infection prevention strategies in hospital settings. [ABSTRACT FROM AUTHOR]

Antimicrobial resistance is a growing public health problem and a threat to effective treatment and prevention of an array of infections caused by bacteria. Africa is already faced with many socio-economic and health crises. Many countries in Africa can seldom boast of a standardized health care facility comparable to those in developed countries. Yet, the non-therapeutic use of COL has been banned in developed countries. However, in Africa, except for South Africa, COL is an over-the-counter (OTC) medication sold and dispensed by non-professionals/without a veterinarian's supervision. The ban of non-therapeutic COL in developed countries has proven to reduce the development of mobile colistin resistance (MCR) in humans and animals. The unregulated use of COL has been proven to select pathogenic and commensal bacteria resistance. A transmissible plasmid-mediated colistin determinant, mobile COL resistance (mcr) gene, which is rapidly transferred/acquired horizontally or laterally intra/inter-species/genera, has been reported. A highly promiscuous mobile genetic element like plasmids containing transposons, insertion sequences, and integrons aid the carriage/rapid transfer and acquisition of these mcr genes. Hence, we highlight the danger posed by escalating colistin (COL) resistance in the continent and the impetus to halt the indiscriminate and non-therapeutic use of COL to protect public health. [ABSTRACT FROM AUTHOR]

Enterococcus faecalis is a ubiquitous bacterium found in various environments, including processed beef meat, and is known for its importance in both food safety and public health. This pivotal significance stems not solely from its virulence but also from its adeptness in eliciting multidrug-resistant infections in humans. The aim of this study was to investigate the population structure, resistome, mobilome, and virulome of E. faecalis obtained from processed beef meat sources in South Africa. A total of eight genomes sequenced in this study were examined, alongside 78 publicly available, high-quality genomes of E. faecalis , with a comprehensive analysis conducted to identify antimicrobial resistance (AMR) determinants, virulence factors, and mobile genetic elements (MGE). Six distinct sequence types (STs) (ST79, ST860, ST40, ST238, ST21, and ST700) and 41 core virulence factors were found across all the genomes. The virulence factors included genes encoding adherence (ace, asa1, Ef0485, ebpA, ebpB, ebpC, srtC); exoenzyme (Ef3023, Ef0818, gelE, sprE); immunomodulation (cpsA, cpsB, cpsC, cpsD, cpsE, cpsF, cpsG, cpsH, cpsI, cpsK), and biofilm formation (bopD, fsrA, fsrB, fsrC). In addition, AMR genes were identified across all genomes, which include aminoglycoside resistance (ant(6)-Ia), trimethoprim resistance (dfrA), drug and biocide resistance (efrA and efrB), multidrug efflux pump (emeA), clindamycin quinupristin-dalfopristin, dalfopristin resistance (lsaA), and tetracycline resistance (tetM). The genomes of E. faecalis sequenced here contained a variety of MGEs, including Insertion Sequences (ISs), transposons, prophages, and plasmids, which may have facilitated genetic exchange within and between these species. The results highlight that beef meat products act as a reservoir for virulent E. faecalis strains possessing antibiotic-resistance traits. This study provides insight into the genomic characteristics, antimicrobial resistance genes, virulence factors, and genetic mobile elements associated with eight E. faecalis isolates from processed beef meat in the Gauteng province of South Africa. • Enterococcus faecalis is a ubiquitous bacterium found in various environments, including processed beef meat, and is known for its importance in both food safety and public health. This pivotal significance stems not solely from its virulence but also from its adeptness in eliciting multidrug-resistant infections in humans. The aim of this study was to investigate the population structure, resistome, mobilome, and virulome of E. faecalis obtained from processed beef meat sources in South Africa. A total of eight genomes sequenced in this study were examined, alongside 78 publicly available, high-quality genomes of E. faecalis , with a comprehensive analysis conducted to identify antimicrobial resistance (AMR) determinants, virulence factors, and mobile genetic elements (MGE).Six distinct sequence types (STs) (ST79, ST860, ST40, ST238, ST21, and ST700) and 41 core virulence factors were found across all the genomes. The virulence factors included genes encoding adherence (ace, asa1, Ef0485, ebpA, ebpB, ebpC, srtC); exoenzyme (Ef3023, Ef0818, gelE, sprE); immunomodulation (cpsA, cpsB, cpsC, cpsD, cpsE, cpsF, cpsG, cpsH, cpsI, cpsK), and biofilm formation (bopD, fsrA, fsrB, fsrC). In addition, AMR genes were identified across all genomes, which include aminoglycoside resistance (ant(6)-Ia), trimethoprim resistance (dfrA), drug and biocide resistance (efrA and efrB), multidrug efflux pump (emeA), clindamycin quinupristin-dalfopristin, dalfopristin resistance (lsaA), and tetracycline resistance (tetM). The genomes of E. faecalis sequenced here contained a variety of MGEs, including Insertion Sequences (ISs), transposons, prophages, and plasmids replicons, which may have facilitated genetic exchange within and between these species. The results highlights that beef meat products act as a reservoir for virulent E. faecalis strains possessing antibiotic-resistance traits. This study provides insight into the genomic characteristics, antimicrobial resistance genes, virulence factors, and genetic mobile elements associated with eight E. faecalis isolates from processed beef meat in the Gauteng province of South Africa. [ABSTRACT FROM AUTHOR]

Food animals are considered reservoirs of methicillin‐resistant Staphylococcus aureus (MRSA) and are implicated in their zoonotic transmission in the farm‐to‐plate continuum. LA‐MRSA has been reported as a zoonotic agent that has the potential to spread to humans and may cause infections in at‐risk groups. In this study, whole genome sequencing was used to describe the genetic environment (resistance mechanisms, virulence factors and mobile genetic elements) and investigate the genetic lineages of MRSA isolates from pigs in Cameroonian and South African abattoirs. During March–October 2016, 288 nasal and rectal pooled samples from 432 pigs as well as nasal and hand swabs from 82 humans were collected. Genomic DNA was sequenced using an Illumina MiSeq platform. Generated reads were de novo‐assembled using the Qiagen CLC Genomics Workbench and SPAdes. The assembled contigs were annotated, and antibiotic resistance genes, virulence factors, plasmids, SCCmec and phage elements were identified with ResFinder, Virulence Finder, PlasmidFinder, SCCmec Finder and PHAST, respectively. Core genome single nucleotide analysis was undertaken to assess clonal relatedness among isolates. A lower MRSA prevalence was observed in pigs in Cameroon (n = 1/13; 0.07%) compared with South Africa (n = 4/22; 18.18%), and none of the workers were colonized by MRSA. Genome analysis identified various antibiotic resistance genes along with six virulence factors in all isolates. All MRSA isolates belonged to the clonal lineage ST398 (spa‐type t011) and harboured the type Vc SCCmec and several plasmids. Our study shows that the livestock‐associated MRSA clonal lineage ST398 is already present in both Cameroon and South Africa and is probably underestimated in the absence of molecular epidemiological studies. It reveals the serious food safety and public health threat associated with this animal strain and underscores the need for interventions to contain this resistant clone. [ABSTRACT FROM AUTHOR]

Horizontal acquisition of mobile genetic elements is a powerful evolutionary driving force that can profoundly affect pathogens epidemiology and their interactions with the environment and host. In the last decade, the role of the epidemic megaplasmid, pESI was demonstrated in the global emergence of multi-drug resistant (MDR) Salmonella enterica serovar Infantis strains, but it was unknown if this was a one-time phenomenon, or that pESI can drive the emergence of other pathogens. Epidemiological, molecular, whole genome sequencing, de-novo assembly, bioinformatics and genetic approaches were used to analyze the emergence of a pESI-positive Salmonella enterica serovar Muenchen strain in Israel. Since 2018, we report the emergence and high prevalence of S. Muenchen in Israel, which consisted at 2020, 40% (1055/2671) of all clinical Salmonella isolates. We show that the emergence of S. Muenchen is dominated by a clonal MDR strain, report its complete assembled genome sequence, and demonstrate that in contrast to preemergent strains, it harbors the epidemic megaplasmid, pESI, which can be self-mobilized into E. coli and other Salmonella serovars. Additionally, we identified bioinformatically highly similar genomes of clinical isolates that were recently collected in South Africa, UK and USA. This is a second documented case of a pathogen emergence associated with pESI acquisition. Considering the genetic cargo of pESI that enhances resistance, stress tolerance and virulence, and its ability to conjugate into prevalent Salmonella serovars, we provide further support that pESI facilities the emergence and spreading of new Salmonella strains. [ABSTRACT FROM AUTHOR]

[Display omitted] • Chicken litter is a major reservoir of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs). • Twenty-one different ARGs were detected in the antibiotic-resistant enterococci isolates from the soil and the chicken litter. • The identified ARGs were mobilised on diverse mobile genetic elements. • The litter-amended soil isolates harboured new ARB and ARGs that were not detected in the soil before the amendment. • Phylogenomic analysis showed that a few isolates from the litter-amended soil were related with the chicken litter isolates. Manure from food animals exposed to antibiotics is often used as soil fertiliser, potentially releasing antibiotic-resistant bacteria (ARB) with diverse antibiotic-resistance genes (ARGs) into the soil. To determine the impact of chicken litter application on the soil resistome, Enterococcus spp. isolated from chicken litter and soil samples collected before and after the soil amendment were characterised, using whole-genome sequencing and bioinformatics tools. Nineteen Enterococcus spp. isolates from the three sources were sequenced on Illumina Miseq platform to ascertain the isolates' resistome, mobilome, virulome, clonality, and phylogenomic relationships. Multilocus sequence typing (MLST) analysis revealed eight novel sequence types (STs) (ST1700, ST1752, ST1753, ST1754, ST1755, ST1756, ST1004, and ST1006). The isolates harboured multiple resistance genes including those conferring resistance to inter alia macrolides-lincosamide-streptogramin (erm(B) , lnu(B), lnu(G) , lsaA , lsaE , eat(A) , msr(C)), tetracycline (tet(M) , tet(L) , tet(S)), aminoglycosides (aac(6')-Ii , aac(6')-Ii h, ant(6)-Ia , aph(3')-III , ant(9)-Ia), fluoroquinolones (efmA, and emeA), vancomycin (VanC {VanC-2 , VanXY , VanXY C-3, VanXY C-4, VanRC }), and chloramphenicol (cat). The litter-amended soil harboured new ARB (particularly E. faecium) and ARGs (ant(6)-Ia , aac(6')-Ii , aph(3')-III), lnu(G) , msr(C), and eat(A), efmA) that were not previously detected in the soil. The identified ARGs were associated with diverse mobile genetic elements (MGEs) such as insertion sequences (IS6, ISL3, IS256, IS30), transposons (Tn 3 and Tn 916) and plasmids (repUS43, repUS1, rep9b, and rep 22). Twenty-eight virulence genes encoding adherence/biofilm formation (ebpA , ebpB , ebpC), antiphagocytosis (elrA) and bacterial sex pheromones (Ccf10, cOB1 , cad , and camE), were detected in the genomes of the isolates. Phylogenomic analysis revealed a close relationship between a few isolates from litter-amended soil and the chicken litter isolates. The differences in the ARG and ARB profiles in the soil before and after the litter amendment and their association with diverse MGEs indicate the mobilisation and transmission of ARGs and ARB from the litter to the soil. [ABSTRACT FROM AUTHOR]

[Display omitted] • Chicken litter is a major reservoir of antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs). • Twenty-one different ARGs were detected in the antibiotic-resistant enterococci isolates from the soil and the chicken litter. • The identified ARGs were mobilised on diverse mobile genetic elements. • The litter-amended soil isolates harboured new ARB and ARGs that were not detected in the soil before the amendment. • Phylogenomic analysis showed that a few isolates from the litter-amended soil were related with the chicken litter isolates. Manure from food animals exposed to antibiotics is often used as soil fertiliser, potentially releasing antibiotic-resistant bacteria (ARB) with diverse antibiotic-resistance genes (ARGs) into the soil. To determine the impact of chicken litter application on the soil resistome, Enterococcus spp. isolated from chicken litter and soil samples collected before and after the soil amendment were characterised, using whole-genome sequencing and bioinformatics tools. Nineteen Enterococcus spp. isolates from the three sources were sequenced on Illumina Miseq platform to ascertain the isolates' resistome, mobilome, virulome, clonality, and phylogenomic relationships. Multilocus sequence typing (MLST) analysis revealed eight novel sequence types (STs) (ST1700, ST1752, ST1753, ST1754, ST1755, ST1756, ST1004, and ST1006). The isolates harboured multiple resistance genes including those conferring resistance to inter alia macrolides-lincosamide-streptogramin (erm(B) , lnu(B), lnu(G) , lsaA , lsaE , eat(A) , msr(C)), tetracycline (tet(M) , tet(L) , tet(S)), aminoglycosides (aac(6')-Ii , aac(6')-Ii h, ant(6)-Ia , aph(3')-III , ant(9)-Ia), fluoroquinolones (efmA, and emeA), vancomycin (VanC {VanC-2 , VanXY , VanXY C-3, VanXY C-4, VanRC }), and chloramphenicol (cat). The litter-amended soil harboured new ARB (particularly E. faecium) and ARGs (ant(6)-Ia , aac(6')-Ii , aph(3')-III), lnu(G) , msr(C), and eat(A), efmA) that were not previously detected in the soil. The identified ARGs were associated with diverse mobile genetic elements (MGEs) such as insertion sequences (IS6, ISL3, IS256, IS30), transposons (Tn 3 and Tn 916) and plasmids (repUS43, repUS1, rep9b, and rep 22). Twenty-eight virulence genes encoding adherence/biofilm formation (ebpA , ebpB , ebpC), antiphagocytosis (elrA) and bacterial sex pheromones (Ccf10, cOB1 , cad , and camE), were detected in the genomes of the isolates. Phylogenomic analysis revealed a close relationship between a few isolates from litter-amended soil and the chicken litter isolates. The differences in the ARG and ARB profiles in the soil before and after the litter amendment and their association with diverse MGEs indicate the mobilisation and transmission of ARGs and ARB from the litter to the soil. [ABSTRACT FROM AUTHOR]

The Human Immunodeficiency Virus type 1 (HIV-1) subtype C is currently the predominant subtype worldwide. Cell culture studies of Sub-Saharan African subtype C proviral plasmids are hampered by the low replication capacity of the resulting viruses, although viral loads in subtype C infected patients are as high as those from patients with subtype B. Here, we describe the sequencing and construction of a new HIV-1 subtype C proviral clone (pZAC), replicating more than one order of magnitude better than the previous subtype C plasmids. We identify the env-region for being the determinant for the higher viral titers and the pZAC Env to be M-tropic. This higher replication capacity does not lead to a higher cytotoxicity compared to previously described subtype C viruses. In addition, the pZAC Vpu is also shown to be able to down-regulate CD4, but fails to fully counteract CD317. [ABSTRACT FROM AUTHOR]