Kanoi, B. | Search bioRxiv

bioRxiv2023

Identification of conserved cross-species B-cell linear epitopes in human malaria: A subtractive proteomics and immuno-informatics approach targeting merozoite stage proteins

Human malaria, caused by five Plasmodium species (P. falciparum, P. vivax, P. malariae, P. ovale, and P. knowlesi), remains a significant global health burden. While most interventions target P. falciparum, the species associated with high mortality rates and severe clinical symptoms, non-falciparum species exhibit different transmission dynamics, remain hugely neglected, and pose a significant challenge to malaria elimination efforts. Recent studies have reported the presence of antigens associated with cross-protective immunity, which can potentially disrupt the transmission of various Plasmodium species. With the sequencing of the Plasmodium genome and the development of immunoinformatic tools, in this study, we sought to exploit the evolutionary history of Plasmodium species to identify conserved cross-species B-cell linear epitopes in merozoite proteins. We retrieved Plasmodium proteomes associated with human malaria and applied a subtractive proteomics approach focusing on merozoite stage proteins. Bepipred 2.0 and Epidope were used to predict B-cell linear epitopes using P. falciparum as the reference species. The predictions were further compared against human and non-falciparum databases and their antigenicity, toxicity, and allergenicity assessed. Subsequently, epitope conservation was carried out using locally sequenced isolates from a malaria-endemic region in western Kenya (n=27) and Kenyan isolates from MalariaGEN version 6 (n=131). Finally, physiochemical characteristics and tertiary structure of the B-cell linear epitopes were determined. The analysis revealed eight epitopes that showed high similarity (70-100%) between falciparum and non-falciparum species. These epitopes were highly conserved when assessed across local isolates and those from the MalariaGEN database and showed desirable physiochemical properties. Our results show the presence of conserved cross-species B-cell linear epitopes that could aid in targeting multiple Plasmodium species. Nevertheless, validating their efficacy in-vitro and in-vivo experimentally is essential.

immunology↗

bioRxiv2023

Application of nanopore sequencing to identify antimicrobial resistance genes, mobile genetic elements and virulence factors in clinical isolates

The global health challenge posed by the emergence of antibiotic resistance pathogen is further exacerbated in African countries by the indiscriminate use of antibiotics, poor surveillance and lack of stewardship programs. To address this issue, we employed the Oxford Nanopore Technologies (ONT) to sequence 17 clinical isolates from a referral hospital in Kenya. Our comprehensive bioinformatics approach facilitated the assembly, identification of sequence types and prediction of antimicrobial resistance genes, mobile genetic elements (plasmids and integrons) and virulence genes. Of the 17 isolates, five were A. baumannii, four E. coli, three S. haemolyticus, three were E. cloacae, while S. aureus and E. faecalis were single isolates. For the detection of AMR genes, A. baumannii isolates harbored genes such as blaOXA-23 which mediates resistance to carbapenems, E. coli and E. cloacae carried blaCTX-M-15 which confers resistance to cephalosporins and S. haemolyticus harbored blaZ, responsible for resistance against penicillins. S. aureus co-haboured mecA and blaZ genes. In addition,, various other different AMR genes to chloramphenicol, macrolides, aminoglycosides, tetracycline were also observed. For plasmid replicons, E. coli carried the most number of plasmids and shared ColRNAI_1 and IncFIB(pB171)_1_pB171 with A. baumannii and IncR_1 with E. cloacae. Many genes encoding various virulence factors including fimA-I and ompA, senB were identified in E. coli, hlgA-C and hla/hly, hlb, hld in S. aureus and efaA, ebpA-C in E. faecalis. In conclusion, most isolates contained a combination of different AMR genes harbored in plasmids and integrons and virulence genes. This study provides significant information on genetic determinants of antibiotic resistant pathogens in clinical isolates and could assist in developing strategies that improve patient treatment. Author SummaryAntimicrobial resistance remains a major health challenge across the globe. The continued misuse and lack of proper monitoring has worsened the problem of antibiotic resistant infections. In this study, we sought to use nanopore sequencing to identify antibiotic resistance genes, mobile genetic elements and virulence factors from clinical isolates which showed resistance against commonly used antibiotics. We found the presence of resistance genes to multiple different antibiotics including beta-lactams, macrolides, tetracycline and aminoglycosides across multiple bacterial species. We also identified plasmid replicons and class I integrons which facilitate the spread of antimicrobial resistant genes. Furthermore, several virulence factors that help resistant bacteria to survive were identified. Overall, this study highlights the widespread issue of antibiotic resistance, factors contributing to its persistence in clinical isolates and utility of nanopore sequencing for monitoring genetic determinants of antimicrobial resistance.

microbiology↗