Melvin Leteane

Background:HIV-1 drug resistance poses a major threat to the success of antiretroviral therapy. The high costs of available HIV drug resistance assays prohibit their routine usage in resource-limited settings. Pan-degenerate amplification and adaptation (PANDAA), a focused genotyping approach based on quantitative PCR (qPCR), promises a fast and cost-effective way to detect HIV drug resistance mutations (HIVDRMs). Given the high cost of current genotyping methods, we sought to use PANDAA for screening key HIVDRMs in antiretroviral-naïve individuals at codons 103, 106 and 184 of the HIV-1 reverse transcriptase gene. Mutations selected at these positions have been shown to be the most common driver mutations in treatment failure. Methods:A total of 103 samples from antiretroviral-naïve individuals previously genotyped by Sanger population sequencing were used to assess and verify the performance of PANDAA. PANDAA samples were run on the ABI 7500 Sequence Detection System to genotype the K103N, V106M and M184V HIVDRMs. In addition, the cost per sample and reaction times were compared. Results:Sanger population sequencing and PANDAA detected K103N mutation in three (2.9%) out of 103 participants. There was no evidence of baseline V106M and M184V mutations observed in our study. To genotype the six HIVDRMs it costs approximately 40 USD using PANDAA, while the reagents cost per test for Sanger population sequencing is approximately 100 USD per sample. PANDAA was performed quicker compared to Sanger sequencing, 2 hours for PANDAA versus 15 hours for Sanger sequencing. Conclusion:The performance of PANDAA and Sanger population sequencing demonstrated complete concordance. PANDAA could improve patient management by providing quick and relatively cheap access to drug-resistance information.

Background:HIV-1 drug resistance poses a major threat to the success of antiretroviral therapy. The high costs of available HIV drug resistance assays prohibit their routine usage in resource-limited settings. Pan-degenerate amplification and adaptation (PANDAA), a focused genotyping approach based on quantitative PCR (qPCR), promises a fast and cost-effective way to detect HIV drug resistance mutations (HIVDRMs). Given the high cost of current genotyping methods, we sought to use PANDAA for screening key HIVDRMs in antiretroviral-naïve individuals at codons 103, 106 and 184 of the HIV-1 reverse transcriptase gene. Mutations selected at these positions have been shown to be the most common driver mutations in treatment failure. Methods:A total of 103 samples from antiretroviral-naïve individuals previously genotyped by Sanger population sequencing were used to assess and verify the performance of PANDAA. PANDAA samples were run on the ABI 7500 Sequence Detection System to genotype the K103N, V106M and M184V HIVDRMs. In addition, the cost per sample and reaction times were compared. Results:Sanger population sequencing and PANDAA detected K103N mutation in three (2.9%) out of 103 participants. There was no evidence of baseline V106M and M184V mutations observed in our study. To genotype the six HIVDRMs it costs approximately 40 USD using PANDAA, while the reagents cost per test for Sanger population sequencing is approximately 100 USD per sample. PANDAA was performed quicker compared to Sanger sequencing, 2 hours for PANDAA versus 15 hours for Sanger sequencing. Conclusion:The performance of PANDAA and Sanger population sequencing demonstrated complete concordance. PANDAA could improve patient management by providing quick and relatively cheap access to drug-resistance information.

The potential antimicrobial properties of a tridentate polypyridyl ligand 4-butoxy-N,N-bis(pyridin-2-ylmethyl)aniline (BUT) 1 and its corresponding mixed ligand ruthenium complexes were investigated on drug-resistant and non-drug-resistant bacterial species. The ligand and its complexes were synthesized and successfully characterized by¹H NMR, UV/Vis, and FTIR spectra; ESI-MS; and magnetic susceptibility. Electronic spectra and magnetic susceptibility of these Ru(II)/(III) complexes suggest that they are of a low spin crystal field split, where the Ru(III) is ad⁵and Ru(II)d⁶low spin. These compounds were tested for antibacterial activity on two bacterial species:Staphylococcus aureus(S. aureus) andKlebsiella pneumoniae(K. pneumoniae), as well as their drug-resistant strains methicillin-resistantStaphylococcus aureus(MRSA) and multidrug resistantKlebsiella pneumoniae(MDRK. pneumoniae). All the compounds inhibited growth of the two non-drug-resistant bacteria and only one drug-resistant strain MRSA. However, only the ligands BUT and 2,2-dipyridylamine showed activity against MRSA, while all complexes did not show any antibacterial activity on MRSA. We observed large zones of inhibition for the Gram-positiveS. aureusand MRSA bacteria, compared to the Gram-negativeK. pneumoniaebacteria. DNA cleavage studies with gel electrophoresis showed denatured bacterial DNA on the gel from all the complexes, with the exception of the ligand, suggesting DNA nuclease activity of the complexes in the bacterial DNA.

Foot-and-mouth disease (FMD) is considered to be endemic in Ghana. However, our knowledge of the local epidemiology of the disease is restricted by a lack of serological information and data for characterized viruses causing field outbreaks. In order to improve our understanding of the prevailing situation, this study was initiated to establish the FMD viruses (FMDV) circulating in the country. During 2016, sera (n = 93) and epithelia/oral swab (n = 20) samples were collected from cattle from four districts in Southern Ghana that experienced FMD outbreaks. Sera were analyzed using the PrioCHECK® FMDV non-structural protein (NSP) ELISA whereas the epithelia/oral swab samples were examined by virus isolation, antigen ELISA, reverse transcription polymerase chain reaction (RT-PCR), and sequencing of VP1 followed by phylogenetic analysis. Assay for antibodies against FMDV NSPs provided evidence of exposure to FMDV in 88.2% (82/93) of the sera tested. Serotypes O and A viruses were detected from clinical samples by RT-PCR and sequencing of VP1. Phylogenetic analysis of VP1 coding sequences revealed that the serotype O viruses belonged to the West Africa (WA) topotype and were most closely related to viruses from Niger and Benin, while the serotype A viruses clustered within genotype IV (G-IV) of the Africa topotype and were most closely related to viruses from Nigeria. This study provides useful information on FMDV serotypes and viral lineages that circulate in Ghana and West Africa that may aid in the formulation of effective FMD control strategies.

The hepatitis B virus (HBV) is a global problem; however, the burden of HBV infection in pregnant women in Botswana is unknown. We sought to determine the prevalence of chronic and occult HBV infection in human immunodeficiency virus (HIV)-infected and -uninfected pregnant women in Botswana. Samples from 752 pregnant women were tested for hepatitis B surface antigen (HBsAg), and HBsAg-positive samples were tested for hepatitis B e antigen (HBeAg) and HBV DNA load. Samples that were HBsAg negative were screened for occult HBV infection by determining the HBV DNA load. HBV genotypes were determined based on a 415-base-pair fragment of the surface gene. Among the 752 women tested during pregnancy or early postpartum, 16 (2.1%) (95% confidence interval (CI): 2.0–2.2) were HBsAg-positive. The prevalence of chronic HBV infection was higher (3.1%) among HIV-infected (95% CI: 3.0–3.2) compared with HIV-uninfected women (1.1%) (95% CI: 1.07–1.1, p = 0.057). Among the 622 HBsAg-negative women, the prevalence of occult HBV infection was 6.6% (95% CI: 6.5–6.7). Three of thirteen HBsAg-positive participants were HBeAg-positive, and all were HIV-negative. Of the 11 maternal samples successfully genotyped, five (45.5%) were genotype D3, five (45.5%) were genotype A1, and one was genotype E (9%). Low and similar proportions of HIV-infected and -uninfected pregnant women in Botswana had occult or chronic HBV infection. We identified a subset of HIV-negative pregnant women who had high HBV DNA levels and were HBeAg-positive, and thus likely to transmit HBV to their infants.