Phenotypic and genome-based characterization of Klebsiella species from different One Health sources in South Africa reveals the presence of multidrug-resistant isolates

Mohamed Ezzat El Zowalaty^1*Bibek Lamichhane²Linda Falgenhauer³Dong-Yeop Lee⁴Emmanuel Chima Eze⁵Sean Gregory Young⁶Asmaa Saleh⁷Stephen Forsythe⁸Oliver Zishiri¹Dong-Hun Lee⁴Ji-Yeon Hyeon⁴Yosra A Helmy²

• ¹University of KwaZulu-Natal, Durban, South Africa
• ²University of Kentucky, Lexington, United States
• ³State Laboratory for Human Medicine, Dillenburg, Germany
• ⁴Konkuk University, Gwangjin-gu, Republic of Korea
• ⁵Louisiana State University, Baton Rouge, United States
• ⁶The University of Texas Southwestern Medical Center, Dallas, United States
• ⁷Princess Nourah bint Abdulrahman University, Riyadh, Saudi Arabia
• ⁸Foodmicrobe.com Ltd, Nottingham,, United Kingdom

The emergence of antimicrobial-resistant Klebsiella represents a serious public health concern. Bacterial strains were isolated from different domestic animals located in KwaZulu Natal and Eastern Cape Provinces in South Africa. Antibiotic susceptibility testing, biofilm formation and hypermucoviscosity were investigated. Genomic DNA of the isolates was extracted, and whole genome sequencing was performed using an Illumina MiSeq platform. Here, we report the isolation, molecular detection, virulence and antimicrobial resistance determinants, as well as phylogenetic analyses of 16 Klebsiella strains using whole genome sequence (WGS) analysis. Ten Klebsiella pneumoniae strains, three Klebsiella variicola and three Klebsiella aerogenes were detected. The assembled contigs were analyzed for AMR and virulence genes, K and O types, and multi-locus sequence type (MLST). To compare the sequenced strains to other previously sequenced Klebsiella species, a distance-based phylogeny method was used to confirm the taxonomic designation of the 16 Klebsiella genomes in an extended context of 169 genomes from the different Klebsiella species. Antibiotic resistance genes conferring resistance to beta-lactams (blaSHV-11, blaSHV-1, blaSHV-119, and blaLEN-16), quinolones (oqxA, oqxA -like, oqxA10, oqxA11, oqxB, oqxB-like, oqxB19, and oqxB25), tetracyclines (tet(A)), and fosfomycins (fosA-like) were detected. All isolates harbored genes conferring resistance to at least three antibiotic classes, thus classifying them as multi-drug-resistant (MDR). Nine multilocus sequence types (ST-76, ST-128, ST-494, ST-617, ST-2465, ST-4039, ST-5470, ST-5478, and ST-5497) were identified using the MLST K. pneumoniae scheme in the Klebsiella collection. The latter three STs being reported for the first time in the current study. MLST analysis revealed that all the three K. variicola isolates belonged to ST-696 using the MLST K. variicola scheme. All 16 Klebsiella isolates (100%) were in vitro biofilm producers. No presumptively hypervirulent strains were identified. All the 16 Klebsiella isolates analyzed contained intact prophages with each genome harboring between one and five prophages. The detection of MDR K. pneumoniae highlights the significance of genomic surveillance for monitoring the dissemination of antimicrobial resistance in livestock pathogens and within the food chain. The report of novel sequence types (ST-5470, ST-5478, and ST-5497) in K. pneumoniae present new allelic profiles, indicating the ongoing evolution and diversification within the species.