Virulence Potential and Antimicrobial Resistance of Listeria monocytogenes Isolates Obtained from Beef and Beef-Based Products Deciphered Using Whole-Genome Sequencing.

Listeria monocytogenes is a ubiquitous bacterial pathogen that threatens the food chain and human health. In this study, whole-genome sequencing (WGS) was used for the genomic characterization of L. monocytogenes (n = 24) from beef and beef-based products. Multilocus Sequence Type (MLST) analysis revealed that ST204 of CC204 was the most common sequence type (ST). Other sequence types detected included ST1 and ST876 of CC1, ST5 of CC5, ST9 of CC9, ST88 of CC88, ST2 and ST1430 of CC2, and ST321 of CC321. Genes encoding for virulence factors included complete LIPI-1 (pfrA-hly-plcA-plcB-mpl-actA) from 54% (13/24) of the isolates of ST204, ST321, ST1430, and ST9 and internalin genes inlABC that were present in all the STs. All the L. monocytogenes STs carried four intrinsic/natural resistance genes, fosX, lin, norB, and mprF, conferring resistance to fosfomycin, lincosamide, quinolones, and cationic peptides, respectively. Plasmids pLGUG1 and J1776 were the most detected (54% each), followed by pLI100 (13%) and pLM5578 (7%). The prophage profile, vB_LmoS_188, was overrepresented amongst the isolates, followed by LP_101, LmoS_293_028989, LP_030_2_021539, A006, and LP_HM00113468. Listeria genomic island 2 (LGI-2) was found to be present in all the isolates, while Listeria genomic island 3 (LGI-3) was present in a subset of isolates (25%). The type VII secretion system was found in 42% of the isolates, and sortase A was present in all L. monocytogenes genomes. Mobile genetic elements and genomic islands did not harbor any virulence, resistance, or environmental adaptation genes that may benefit L. monocytogenes. All the STs did not carry genes that confer resistance to first-line antibiotics used for the treatment of listeriosis. The characterization of L. monocytogenes in our study highlighted the environmental resistance and virulence potential of L. monocytogenes and the risk posed to the public, as this bacterium is frequently found in food and food processing environments.

Resistome, mobilome, virulome analysis and phylogenomics of Enterococcus faecalis isolated from raw muscle foods of beef origin in Gauteng, 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, 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.

Genus-wide genomic characterization of Macrococcus: insights into evolution, population structure, and functional potential.

Introduction: Macrococcus species have been isolated from a range of mammals and mammal-derived food products. While they are largely considered to be animal commensals, Macrococcus spp. can be opportunistic pathogens in both veterinary and human clinical settings. This study aimed to provide insight into the evolution, population structure, and functional potential of the Macrococcus genus, with an emphasis on antimicrobial resistance (AMR) and virulence potential. Methods: All high-quality, publicly available Macrococcus genomes (n = 104, accessed 27 August 2022), plus six South African genomes sequenced here (two strains from bovine clinical mastitis cases and four strains from beef products), underwent taxonomic assignment (using four different approaches), AMR determinant detection (via AMRFinderPlus), and virulence factor detection (using DIAMOND and the core Virulence Factor Database). Results: Overall, the 110 Macrococcus genomes were of animal commensal, veterinary clinical, food-associated (including food spoilage), and environmental origins; five genomes (4.5%) originated from human clinical cases. Notably, none of the taxonomic assignment methods produced identical results, highlighting the potential for Macrococcus species misidentifications. The most common predicted antimicrobial classes associated with AMR determinants identified across Macrococcus included macrolides, beta-lactams, and aminoglycosides (n = 81, 61, and 44 of 110 genomes; 73.6, 55.5, and 40.0%, respectively). Genes showing homology to Staphylococcus aureus exoenzyme aureolysin were detected across multiple species (using 90% coverage, n = 40 and 77 genomes harboring aureolysin-like genes at 60 and 40% amino acid [AA] identity, respectively). S. aureus Panton-Valentine leucocidin toxin-associated lukF-PV and lukS-PV homologs were identified in eight M. canis genomes (≥40% AA identity, >85% coverage). Using a method that delineates populations using recent gene flow (PopCOGenT), two species (M. caseolyticus and M. armenti) were composed of multiple within-species populations. Notably, M. armenti was partitioned into two populations, which differed in functional potential (e.g., one harbored beta-lactamase family, type II toxin-antitoxin system, and stress response proteins, while the other possessed a Type VII secretion system; PopCOGenT p < 0.05). Discussion: Overall, this study leverages all publicly available Macrococcus genomes in addition to newly sequenced genomes from South Africa to identify genomic elements associated with AMR or virulence potential, which can be queried in future experiments.

Whole Genome-Based Characterization of Listeria monocytogenes Isolates Recovered From the Food Chain in South Africa.

Listeria monocytogenes is an important foodborne pathogen which has the ability to adapt and survive in food and food processing facilities where it can persist for years. In this study, a total of 143 L. monocytogenes isolates in South Africa (SA) were characterized for their strain's genetic relatedness, virulence profiles, stress tolerance and resistance genes associated with L. monocytogenes. The Core Genome Multilocus Sequence Typing (cgMLST) analysis revealed that the most frequent serogroups were IVb and IIa; Sequence Types (ST) were ST204, ST2, and ST1; and Clonal Complexes (CC) were CC204, CC1, and CC2. Examination of genes involved in adaptation and survival of L. monocytogenes in SA showed that ST1, ST2, ST121, ST204, and ST321 are well adapted in food processing environments due to the significant over-representation of Benzalkonium chloride (BC) resistance genes (bcrABC cassette, ermC, mdrL and Ide), stress tolerance genes (SSI-1 and SSI-2), Prophage (φ) profiles (LP_101, vB LmoS 188, vB_LmoS_293, and B054 phage), plasmids profiles (N1-011A, J1776, and pLM5578) and biofilm formation associated genes. Furthermore, the L. monocytogenes strains that showed hyper-virulent potential were ST1, ST2 and ST204, and hypo-virulent were ST121 and ST321 because of the presence and absence of major virulence factors such as LIPI-1, LIPI-3, LIPI-4 and the internalin gene family members including inlABCEFJ. The information provided in this study revealed that hyper-virulent strains ST1, ST2, and ST204 could present a major public health risk due to their association with meat products and food processing environments in SA.

Population Structure of Non-ST6 Listeria monocytogenes Isolated in the Red Meat and Poultry Value Chain in South Africa.

Meat products have been implicated in many listeriosis outbreaks globally, however there is a dearth of information on the diversity of L. monocytogenes isolates circulating in food products in South Africa. The aim of this study was to investigate the population structure of L. monocytogenes isolated in the meat value chain within the South African market. Based on whole-genome sequence analysis, a total of 217 isolates were classified into two main lineage groupings namely lineages I (n = 97; 44.7%) and II (n = 120; 55.3%). The lineage groups were further differentiated into IIa (n = 95, 43.8%), IVb (n = 69, 31.8%), IIb (n = 28, 12.9%), and IIc (n = 25, 11.5%) sero-groups. The most abundant sequence types (STs) were ST204 (n = 32, 14.7%), ST2 (n = 30, 13.8%), ST1 (n = 25, 11.5%), ST9 (n = 24, 11.1%), and ST321 (n = 21, 9.7%). In addition, 14 clonal complex (CCs) were identified with over-representation of CC1, CC3, and CC121 in "Processed Meat-Beef", "RTE-Poultry", and "Raw-Lamb" meat categories, respectively. Listeria pathogenic islands were present in 7.4% (LIPI-1), 21.7% (LIPI-3), and 1.8% (LIPI-4) of the isolates. Mutation leading to premature stop codons was detected in inlA virulence genes across isolates identified as ST121 and ST321. The findings of this study demonstrated a high-level of genomic diversity among L. monocytogenes isolates recovered across the meat value chain control points in South Africa.