RESUMO
BACKGROUND: Africa has one of the highest incidences of gonorrhea. Neisseria gonorrhoeae is gaining resistance to most of the available antibiotics, compromising treatment across the world. Whole-genome sequencing (WGS) is an efficient way of predicting AMR determinants and their spread in the population. Recent advances in next-generation sequencing technologies like Oxford Nanopore Technology (ONT) have helped in the generation of longer reads of DNA in a shorter duration with lower cost. Increasing accuracy of base-calling algorithms, high throughput, error-correction strategies, and ease of using the mobile sequencer MinION in remote areas lead to its adoption for routine microbial genome sequencing. To investigate whether MinION-only sequencing is sufficient for WGS and downstream analysis in resource-limited settings, we sequenced the genomes of 14 suspected N. gonorrhoeae isolates from Nairobi, Kenya. METHODS: Using WGS, the isolates were confirmed to be cases of N. gonorrhoeae (n = 9), and there were three co-occurrences of N. gonorrhoeae with Moraxella osloensis and N. meningitidis (n = 2). N. meningitidis has been implicated in sexually transmitted infections in recent years. The near-complete N. gonorrhoeae genomes (n = 10) were analyzed further for mutations/factors causing AMR using an in-house database of mutations curated from the literature. RESULTS: We observe that ciprofloxacin resistance is associated with multiple mutations in both gyrA and parC. Mutations conferring tetracycline (rpsJ) and sulfonamide (folP) resistance and plasmids encoding beta-lactamase were seen in all the strains, and tet(M)-containing plasmids were identified in nine strains. Phylogenetic analysis clustered the 10 isolates into clades containing previously sequenced genomes from Kenya and countries across the world. Based on homology modeling of AMR targets, we see that the mutations in GyrA and ParC disrupt the hydrogen bonding with quinolone drugs and mutations in FolP may affect interaction with the antibiotic. CONCLUSION: Here, we demonstrate the utility of mobile DNA sequencing technology in producing a consensus genome for sequence typing and detection of genetic determinants of AMR. The workflow followed in the study, including AMR mutation dataset creation and the genome identification, assembly, and analysis, can be used for any clinical isolate. Further studies are required to determine the utility of real-time sequencing in outbreak investigations, diagnosis, and management of infections, especially in resource-limited settings.
RESUMO
OBJECTIVE: To evaluate initial reported willingness to participate in a hypothetical HIV vaccine clinical trial and actual participation of volunteers in a longitudinal observational study. METHODS: We recruited HIV negative male and female volunteers aged 18-45 years into a longitudinal observational study at KAVI-ICR Kangemi in Kenya, to serve as a pool from which to draw participants into a phase I HIV vaccine clinical trial. A structured questionnaire was used to collect information regarding willingness to join a HIV vaccine clinical trial in the future. Study follow-up visits were every 6 months. RESULTS: A total of 105 participants were screened and 100 (M46:F54) were enrolled into the observational study. Ninety- four per cent of those enrolled expressed willingness to participate in a future HIV vaccine trial. Altruism and desire to learn the body's response to the vaccine were the most motivating factors at 40% and 25% respectively. At the onset of a 40-person phase I HIV vaccine trial, 86 observational study participants who had previously expressed willingness to participate were contacted but only 26 (30%) came for information. All 26 consented to participate and after screening for eligibility, 24 were eligible. Of the 24, 15 were enrolled. These numbers were not adequate; hence the vaccine trial employed other recruitment methods to meet the deficit. CONCLUSION: Observational "pools" of cohorts may not provide adequate number of participants into vaccine clinical trials even if they report willingness; therefore supplementary recruitment methods such as direct community recruitment, passive approach, and snowballing need to be in place.
