Prevalence and Molecular Epidemiology of Human Coronaviruses in Africa Prior to the SARS-CoV-2 Outbreak: A Systematic Review.

Coronaviruses, re-emerging in human populations, cause mild or severe acute respiratory diseases, and occasionally epidemics. This study systematically reviewed human coronavirus (HCoVs) infections in Africa prior to the SARS-CoV-2 outbreak. Forty studies on the prevalence or molecular epidemiology of HCoVs were available from 13/54 African countries (24%). The first published data on HCoV was from South Africa in 2008. Eight studies (20%) reported on HCoV molecular epidemiology. Endemic HCoV prevalence ranged from 0.0% to 18.2%. The prevalence of zoonotic MERS-CoV ranged from 0.0% to 83.5%. Two studies investigated SARS-CoV infection, for which a prevalence of 0.0% was reported. There was heterogeneity in the type of tests used in determining HCoV prevalence. Two studies reported that risk factors for HCoV include exposure to infected animals or humans. The quantity of virologic investigations on HCoV on the African continent was scant, and Africa was not prepared for SARS-CoV-2.

Molecular epidemiology of SARS-CoV-2 in Northern South Africa: wastewater surveillance from January 2021 to May 2022.

Introduction: Wastewater-based genomic surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) provides a comprehensive approach to characterize evolutionary patterns and distribution of viral types in a population. This study documents the molecular epidemiology of SARS-CoV-2, in Northern South Africa, from January 2021 to May 2022. Methodology: A total of 487 wastewater samples were collected from the influent of eight wastewater treatment facilities and tested for SARS-CoV-2 RNA using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). SARS-CoV-2 positive samples with genome copies/mL ≥1,500 were subjected to allele-specific genotyping (ASG) targeting the Spike protein; 75 SARS-CoV-2 positive samples were subjected to whole genome sequencing (WGS) on the ATOPlex platform. Variants of concern (VoC) and lineages were assigned using the Nextclade and PangoLIN Software. Concordance for VoC between ASG and WGS analyses was determined. Sequence relationship was determined by phylogenetic analysis. Results: Seventy-five percent (365/487) of the influent samples were positive for SARS-CoV-2 RNA. Delta and Omicron VoC were more predominant at a prevalence of 45 and 32%, respectively, and they were detected as early as January and February 2021, while Beta VoC was least detected at a prevalence of 5%. A total of 11/60 (18%) sequences were assigned lineages and clades only, but not a specific VoC name. Phylogenetic analysis was used to investigate the relationship of these sequences to other study sequences, and further characterize them. Concordance in variant assignment between ASG and WGS was seen in 51.2% of the study sequences. There was more intra-variant diversity among Beta VoC sequences; mutation E484K was absent. Three previously undescribed mutations (A361S, V327I, D427Y) were seen in Delta VoC. Discussion and Conclusion: The detection of Delta and Omicron VoCs in study sites earlier in the outbreak than has been reported in other regions of South Africa highlights the importance of population-based approaches over individual sample-based approaches in genomic surveillance. Inclusion of non-Spike protein targets could improve the specificity of ASG, since all VoCs share similar Spike protein mutations. Finally, continuous molecular epidemiology with the application of sensitive technologies such as next generation sequencing (NGS) is necessary for the documentation of mutations whose implications when further investigated could enhance diagnostics, and vaccine development efforts.

Epidemiology of Campylobacter infections among children of 0-24 months of age in South Africa.

BACKGROUND: Campylobacter spp. are one of the most frequent causes of diarrhoeal disease in humans throughout the world. This study aimed at determining the prevalence and the genotypic distribution of Campylobacter spp. and their association with diarrhoea and child growth in children of less than the age of two in the Limpopo Province of South Africa. METHODS: A total of 4280 diarrheal and non-diarrheal stool samples were collected on a monthly basis from children recruited at birth and followed up to 24 months. All stool samples were screened for the presence Campylobacter antigen using ELISA technique after which CAH 16S primer was used on the positive samples to confirm the presence of Campylobacter. Subsequently, the PCR positive samples were further characterised using species specific primers for Campylobacter jejuni and Campylobacter coli. RESULTS: Campylobacter antigen was detected in 564/4280 (13.2%). Campylobacter was more commonly found in diarrheal stools (20.4%) compared to non-diarrheal stools (12.4%) with a statistically significant difference (χ2 = 7.345; p = 0.006). Throughout the year there were two main peaks of Campylobacter infection one in December- January and the second peak in June. The prevalence of Campylobacter increased with the age of the children up to 11 months after which the prevalence decreased. Out of 564 positive ELISA samples, 257 (45.6%) were confirmed to have 16S rRNA gene for Campylobacter spp. Furthermore, C. jejuni was found to be more prevalent (232/257) than C. coli (25/257) with a prevalence of 90.3% and 9.7%, respectively. Both C. jejuni and C. coli were significantly associated with diarrhea with statistical values of (χ2 = 22.224; p < 0.001) and (χ2 = 81.682; p < 0.001) respectively. Sequences generated from the analysis of hip gene confirmed the PCR positives samples were C. jejuni positive. CONCLUSIONS: This study has delineated a high prevalence of Campylobacter spp. in the study cohort. Moreover, C. jejuni was found to be more prevalent than C. coli both of which were associated with diarrhea. These findings are of clinical and epidemiological significance.

Development of a pseudovirus assay and evaluation to screen natural products for inhibition of HIV-1 subtype C reverse transcriptase.

ETHNOPHARMACOLOGICAL RELEVANCE: Medicinal plants are used in the management of Human Immunodeficiency Virus and Acquired Immunodeficiency Syndrome (HIV/AIDS) in many developing country settings where HIV-1 subtype C drives the epidemic. Efforts to identify plant derived molecules with anti-HIV properties require reproducible assay systems for routine screening of selected plant compounds. Although a number of standardized HIV-1 pseudoviruses have been generated to assess infectivity, replicability or reproducibility, HIV-1 subtype C (HIV-1-C) pseudoviruses have not been comprehensively characterized to identify inhibitory plant substances. AIM OF THE STUDY: The current study aimed at developing an HIV-1-C pseudovirus assay, and evaluate plant substances targeting reverse transcriptase (RT) activity. MATERIALS AND METHODS: HIV-1 subtype C pseudoviruses containing a luciferase reporter gene were generated by transfection of human 293T cells. HIV-1 subtype B (HIV-1-B) wild type pseudoviruses and mutants resistant to nucleoside and non-nucleoside RT inhibitors were also generated and used as controls. Selected plant substances and the RT inhibitors Zidovudine (AZT) and Nevirapine (NVP), were used to evaluate inhibition. Pseudovirus infectivity was determined by luciferase measurement in CF2/CD4+/CCR5 cells, and cytotoxicity was determined using the MTT assay. AZT and NVP inhibited wild type pseudoviruses in a dose dependent manner, with IC50 values in the nanomolar range. RESULTS: Pseudoviruses harbouring RT drug resistance mutations were poorly suppressed by AZT and NVP. Catechin, obtained from Peltophorum africanum inhibited HIV-1-C and HIV-1-B pseudoviruses with selective indices of 6304 µM (IC50: 0.49 µM, CC50: 3089 µM) and 1343 µM (IC50: 2.3 µM, CC50: 3089 µM), respectively; while the methanol root crude extract of Elaeodendron transvaalense gave IC50 values of 11.11 µg/ml and 16.86 µg/ml, respectively. CONCLUSION: The developed HIV-1-C pseudovirus assay can be used to screen plant substances for RT inhibition, and may have utility in settings with limited access to high level biosafety facilities.

Prevalence of torque viruses in HIV-infected and non-HIV-infected Nigerian subjects: analysis of near-full-length genome sequences.

Torque teno virus (TTV), torque teno mini virus (TTMV) and torque teno midi virus (TTMDV) are members of the family Anelloviridae that are known to infect humans. Although no pathogenic roles have been associated with anelloviruses, their high prevalence and perceived ubiquitousness have provoked scientific interest in understanding their molecular and biological characteristics. We used nested PCR to determine the prevalence of anelloviruses among 130 human immunodeficiency virus (HIV)-infected patients and 130 healthy blood donors, and analyzed three near-full-length genome sequences of TTV isolates from HIV-infected and non-HIV infected Nigerians. Statistical analysis showed that the rate of TTV infection was significantly higher in the HIV-infected group (65%) than in the blood donor group (26%) (p < 0.05, χ2 = 40.3). TTMV and TTMDV infections were very high in both groups, ranging between 88 and 95%. No significant association was found between TTV infection and age, sex, CD4+ cell count, HIV viral load or alanine aminotransferase (ALT) level. Near-full-length genome sequences of TTV isolates FL100, FL08 and BD67 determined by next-generation sequencing were 3.6 kb, 3.2 kb and 2.9 kb, respectively, in size. Their GenBank accession numbers are MK820644, MK820645, MK820646, respectively. These isolates shared 59% sequence identity across the whole genome and clustered in two different phylogenetic groups. Our study established for the first time the circulation of TTV, TTMV and TTMDV in the Nigerian population, with a disproportionately higher prevalence of TTV in HIV-infected patients. The near-complete TTV genome sequences from Nigeria are similar to the sequences KT163879 and KT163916 (3748 and 3190 respectively), obtained from the plasma of HIV-infected subjects from the United States, and EU305675 (2919), identified in human plasma samples from France.

Aflatoxin exposure in children living in Mirpur, Dhaka: data from MAL-ED companion study.

Dietary exposure to aflatoxin is implicated in growth faltering of children. Despite the high burden of childhood stunting in urban Bangladesh, there are no data on long-term exposure to aflatoxin. This study aimed to explore aflatoxin exposure levels in a group of children followed longitudinally. The current study used data and biospecimens collected during 2010-2014 as part of the MAL-ED birth cohort study in an urban slum of Mirpur, Dhaka where children were followed from birth to 36 months. AFB1-lysine adduct concentrations were determined by isotope dilution mass spectrometry from plasma samples collected at 7, 15, 24, and 36 months of age. The limit of detection was 0.5 pg of AFB1-lys/mg albumin. In 744 plasma samples, the geometric mean of AFB1-lysine/mg albumin was 1.07 pg (range 0.04-123.5 pg/mg albumin). The proportion of children with detectable aflatoxin exposure was 10.1, 20.9, 17.9, and 61.7% for 7, 15, 24, and 36 months, respectively. Reduction in breastfeeding prevalence (80% at 24 months vs. 38% in 36 months) corresponded with the high-level detection of AFB1-lysine at the age of 36 months. AFB1-lysine concentrations were the highest at the end of monsoon. This study reveals that 62% of children in slum settlement were exposed to aflatoxin by the end of the third year of life. High aflatoxin exposure was detected at the end of rainy season and with the introduction of family food. These findings suggest interventions to ameliorate the problem of chronic aflatoxin exposure including childhood stunting.

A putative HIV-1 subtype C/CRF11_cpx unique recombinant from South Africa.

The HIV epidemic in South Africa is overwhelmingly driven by HIV-1 subtype C viruses. The HIV gag, pol, env (C2-V5) and nef sequences of virus 08MB26ZA, obtained from a 47 year old woman, were studied by phylogenetic analysis, REGA and the jumping Profile Hidden Markov Model (jPHMM) tools. The pol gene was further analyzed for recombination by Simplot. The pol and env sequences were examined for genetic drug resistance mutations and predicted co-receptor usage respectively. There was agreement in the assignment of the gag sequence as pure HIV-1 subtype C by phylogenetic, REGA and jPHMM analyses. The pol sequence clustered with CRF11_cpx and in the J-clade by phylogenetic analysis; and to a CRF11_cpx/subtype C recombinant by REGA. The assignment of pol to CRF11_cpx and subtype C was confirmed by Simplot. The recombinant was of the R5 biotype, with no important drug resistance mutations in the pol region. The epidemiologic and biologic significance of the virus are unknown. The finding suggests that complex viruses are being introduced into South Africa with potential implications for diagnosis. This is apparently the first report from South Africa of a putative unique recombinant involving CRF11_cpx and subtype C genomes.

Development of the Dzimauli community in Vhembe District, Limpopo province of South Africa, for the MAL-ED cohort study.

The Dzimauli community is located in the Vhembe district in the northern part of Limpopo Province, South Africa. The district is bordered by Botswana and Zimbabwe to the north and Mozambique to the East. The study site population is entirely blacks and 53% female, with a mean household size of 6 persons. Through a consultative process, we engaged and prepared the Dzimauli, a community of low socioeconomic status, to participate in a longitudinal, observational study. In addition to contributing to the objectives of The Etiology, Risk Factors and Interactions of Enteric Infections and Malnutrition and the Consequences for Child Health and Development (MAL-ED) cohort study, we established a high degree of public trust and understanding of scientific research within the community and its leaders. This has resulted in creating an entirely new site suitable for potential future field-based intervention studies based on an improved understanding of the factors influencing child health in this community.

Genetic variants of APOC3 promoter and HLA-B genes in an HIV infected cohort in northern South Africa: a pilot study.

Metabolic disorders and hypersensitivities affect tolerability and impact adherence to highly active antiretroviral therapy (HAART). The aim of this study was to determine the prevalence of C-482T/T-455C variants in the Apolipoprotein C3 (APOC3) promoter gene and Human leukocyte antigen (HLA)-B*57:01, known to impact lipid metabolic disorders and hypersensitivity respectively; and to correlate genotypes with gender, CD4+ cell count and viral load in an HIV infected cohort in northern South Africa. Frequencies of C-482 and T-455 polymorphisms in APOC3 were determined by restriction fragment length polymorphism analysis. Allele determination for HLA-B was performed with Assign SBT software in an HLA library. Analysis of APOC3 C-482 site revealed a prevalence of 196/199 (98.5%) for CC, 1/199 (0.5%) for CT and 2/199 (1.0%) for TT genotype (p = 0.000 with 1° of freedom; χ2 = 126.551). For the T-455 site, prevalences were: 69/199 (35%) for TT and 130/199 (65%) for the CC genotype (p = 0.000 with 1° of freedom; χ2 = 199). There was no association between gender and the presence of -482 (p = 1; χ2 = 0.00001) or -455 genotypes (p = 0.1628; χ2 = 1.9842). There was no significant difference in the increase in CD4+ cell count irrespective of genotypes. Significant increases in CD4+ cell count were observed in males and females considering the -455C genotype, but not in males for the -455T genotype. Viral load decreases were significant with the -455C and -482C genotypes irrespective of gender. HLA-B*57:01 was not identified in the study cohort. The apparently high prevalence of APOC3 T-455CC genotype needs confirmation with a larger samples size and triglyceride measurements to support screening of patients to pre-empt HAART associated lipid disorders.

Genetic Analysis of HIV-1 Integrase Sequences from Treatment Naive Individuals in Northeastern South Africa.

Raltegravir, an integrase inhibitor, is not a component of the current South African antiretroviral treatment guidelines, but it could be introduced in the near future as cases of virological failures from current treatment regimens begin to occur. The aim of this study was to analyze the complete HIV integrase gene obtained from individuals at two treatment sites in northeastern South Africa for the presence of Raltegravir associated drug resistant mutations and viral subtypes based on the integrase gene. Examination for mutations against other integrase inhibitors, such as Elvitegravir and Dolutegravir, was also done. Viruses from 127 treatment naive individuals were analyzed. Genetic drug resistance mutations were determined using the Stanford HIV Drug Resistance Interpretation program and the International AIDS society-USA guidelines. Viral subtyping was done by phylogenetic analysis, and recombinants were determined using the REGA, jpHMM and RIP tools. No major resistance mutations were detected. However, 7% of the sequences had minor mutations and polymorphisms. The majority (99%) of the viruses were HIV-1 C. Recombination analysis showed that the polymerase gene of one virus was likely composed of HIV-1 subtype A1 and C sequences. The present study indicates that Raltegravir, Elvitegravir and Dolutegravir resistant mutations may be absent in the study communities and further indicates the presence of recombinant viruses in northeastern South Africa.

Low prevalence of transmitted genetic drug resistance in a cohort of HIV infected naïve patients entering antiretroviral treatment programs at two sites in northern South Africa.

Infection with drug resistant viruses influences the outcome of antiretroviral therapy (ART). This study was carried out to determine the transmitted genetic drug resistance profile in a cohort of patients prior to initiation of treatment at two treatment sites in northern South Africa. These study sites were among the first to benefit from antiretroviral drugs in this region. Data on HIV drug resistance are also limited in northern South Africa; and resistance testing prior to initiation of treatment is not undertaken. In 2008, 80 protease and 80 reverse transcriptase nucleotide sequences obtained from 80 patients were analyzed for genetic drug resistance using the calibrated population resistance tool for transmitted drug resistance. Viral genetic subtypes were determined by phylogenetic analysis. Two drug resistance mutations (M41L and K103N) were detected in two different patients (2.5%; 95% CI: 0.0077-0.0863). Twenty-three sequences (29%) harbored at least one secondary mutation in the reverse transcriptase gene; while all sequences had at least one minor mutation in the protease gene. Phylogenetic analysis of the protease and reverse transcriptase genes showed that 79 out of 80 viruses were HIV-1 subtype C, and one was an A1/C recombinant. The observations suggest that after 4 and 8 years access to ART in Mankweng and the Bela Bela communities respectively, drug resistance mutations in the naïve population was low. Regular studies are needed to update information on drug resistant viruses in treatment naïve patients to inform better treatment policies.

Prevalence of MDR1 C3435T and CYP2B6 G516T polymorphisms among HIV-1 infected South African patients.

Data on genetic polymorphisms associated with response to anti-HIV drugs has accumulated over the years. Information on how polymorphisms influence drug metabolism and transport to target sites is important in guiding dosage or selection of appropriate alternative therapies. This study determined the frequency of MDR1 C3435T and CYP2B6 G516T polymorphisms associated with the transport and metabolism of efavirenz and nevirapine, in a population of South African HIV infected patients. In addition, association of polymorphisms with immunologic and virologic factors was investigated. A 207bp of MDR1 exon 26 and a 161bp of CYP2B6 exon 4 were obtained from patients by polymerase chain reaction. Analysis of population-based sequences of MDR1 revealed a frequency of 89% and 11% of C and T alleles respectively (n=197; X^{2} = 0.974; p=0.324). Restriction fragment length polymorphism (RFLP) analysis of the CYP2B6 gene revealed a prevalence of 9.5% of GG, 78.4% of GT and 12.1% of TT genotype (n= 199; X^{2} = 65.204; p=0.00). There was no significant difference between immune recovery and decline in viral load (n=53), with genotype after repeated calculations of analysis of variance (ANOVA).

Molecular epidemiology of HIV in two highly endemic areas of northeastern South Africa.

There is paucity of data on the genetic landscape of HIV-1 viruses circulating in the Limpopo Province of northeastern South Africa. Here, we examine the genetic diversity of viruses from Bela-Bela and Musina, two towns with high HIV prevalence. Between June 2007 and March 2008, blood samples were collected from antiretroviral-drug-naïve individuals. Viruses were analyzed for genetic subtypes and drug resistance mutations. All of the viruses in these samples were shown by phylogenetic analysis based on gag p17, gag p24, reverse transcriptase, protease and envelope C2-C3 gene regions to belong to HIV-1 subtype C. Two of 44 reverse transcriptase sequences (4.5%) contained N rather than the consensus K at position 103. The K103N mutation is normally associated with resistance to NNRTIs. No major mutations were observed in the protease gene. However, several polymorphisms and amino acid changes normally considered to be minor drug resistance mutations were observed in the protease sequences. These results suggest that HIV-1 subtype C remains the predominant variant responsible for the epidemic in northeastern South Africa and that the prevalence of drug-resistant viruses among the naïve population is low.

Peltophorum africanum, a traditional South African medicinal plant, contains an anti HIV-1 constituent, betulinic acid.

The biodiversity of medicinal plants in South Africa makes them rich sources of leading compounds for the development of novel drugs. Peltophorum africanum (Fabaceae) is a deciduous tree widespread in South Africa. The stem bark has been traditionally employed to treat diarrhoea, dysentery, sore throat, wounds, human immunodeficiency virus/ acquired immune deficiency syndrome (HIV/AIDS), venereal diseases and infertility. To evaluate these ethnobotanical clues and isolate lead compounds, butanol and ethyl acetate extracts of the stem bark were screened for their inhibitory activities against HIV-1 using MAGI CCR5+ cells, which are derived from HeLa cervical cancer cells and express HIV receptor CD4, a chemokine receptor CCR5 and HIV-LTR-beta- galactosidase. Bioassay-guided fractionation using silica gel chromatography was also conducted. The ethyl acetate and butanol extracts of the stem bark of Peltophorum africanum showed inhibitory activity against HIV-1, CXCR4 (X4) and CCR5 (R5) tropic viruses. The ethyl acetate and butanol extracts yielded previously reported anti-HIV compounds, (+)-catechin, a flavonoid, and bergenin, a C-galloylglycoside, respectively. Furthermore, we identified betulinic acid from the ethyl acetate fraction for the first time. The fractions, which contained betulinic acid, showed the highest selective index. We therefore describe the presence of betulinic acid, a not well-known anti-HIV compound, in an African medicinal herb, which has been used for therapy, and claim that betulinic acid is the predominant anti-HIV-1 constituent of Peltophorum africanum. These data suggest that betulinic acid and its analogues could be used as potential therapeutics for HIV-1 infection.

Polymorphisms in HIV-1 subtype C proteases and the potential impact on protease inhibitors.

OBJECTIVES: (i) To review data on the genetic profile of the protease (PR) gene of human immunodeficiency virus (HIV)-1-C primary isolates relative to HIV-1-B; (ii) to examine data on the susceptibility of HIV-1-C isolates harbouring polymorphisms to PR inhibitors (PI) and the development of resistance; and (iii) to identify gaps required for an improved understanding of the role of polymorphisms in resistance development of HIV-1-C to PI. METHOD: Literature review. RESULTS: Significant differences exist between the baseline nucleotide and amino acid sequences of PR of HIV-1-B and HIV-1-C. Some of the amino acid substitutions seen in HIV-1-B when exposed to PI occur naturally in HIV-1-C isolates. Studies used different methodologies and interpretation systems to evaluate the phenotypic significance of polymorphisms seen in subtype C viruses, with conflicting outcomes. The evolutionary path to the resistance of HIV-1-C to PI may be different from that of HIV-1-B. CONCLUSIONS: Infection with HIV-1-C is driving the AIDS epidemic in regions of the world with the most urgent needs for the management of the disease. More and more individuals will require PR inhibitors in second-line therapies, as access to antiretrovirals progresses. It is proposed that a standardized protocol be adopted to evaluate the phenotypic significance of the highly polymorphic HIV-1-C PR to PR inhibitors with the aim of better informing the tailoring of treatment regimens for optimal clinical benefit.

Issues surrounding the use of herbal therapies for AIDS in endemic regions.

Herbal preparations are rampantly used in the treatment of AIDS in endemic regions. Despite beneficial effects from the use of some plant preparations, the issues that still have to be addressed comprise the interaction between herbal preparations and antiretrovirals, lack of conclusive clinical trials of widely used plants, and ethical issues surrounding the practices of traditional healers. The burden presented by AIDS requires an urgent need for closer collaboration among scientists, medical practitioners and traditional systems of medicine to evaluate promising herbal preparations rigorously, and develop policies and guidelines for the judicious use of herbs with proven clinical benefit.

Bacterial contamination of Vhuswa--a local weaning food and stored drinking-water in impoverished households in the Venda region of South Africa.

Bacterial contaminants of Vhuswa--a traditional maize-based weaning food, and domestic drinking-water stored in impoverished rural households in Venda of Limpopo province, South Africa, were determined. One hundred and twenty-five samples of Vhuswa fed to children aged less than five years were assessed for Escherichia coli, Campylobacter jejuni, Salmonella, and Shigella. The microbiological quality of 125 drinking-water samples was also evaluated using total coliforms, faecal coliforms, and faecal streptococci as indicators. The frequency of isolation of E. coli, Salmonella, Shigella, and C. jejuni from the Vhuswa samples was 70%, 5%, 5%, and 2% respectively. The geometric mean counts of total coliforms, faecal coliforms, and faecal streptococci in tap-water stored in household containers ranged from 4.9x10(2) to 5.8x10(3) cfu 100 mL(-1), 2.6x10(2) to 3.7x10(3) cfu 100 mL(-1), and 3.1x10(3) to 5.8x10(3) cfu 100 mL(-1) respectively, and for stored spring water it was 5.1x10(3) cfu 100 mL(-1), 3.2x10(3) cfu 100 mL(-1), and 5.1x10(3) cfu 100 mL(-1) respectively. The frequent contamination of water and food samples in this study has important implications for the health of children from impoverished communities.

Evaluation of selected South African medicinal plants for inhibitory properties against human immunodeficiency virus type 1 reverse transcriptase and integrase.

Seventeen aqueous and methanol extracts from nine South African medicinal plants, ethnobotanically selected, were screened for inhibitory properties against HIV-1 reverse transcriptase (RT). Isolated compounds were additionally evaluated on HIV-1 integrase (IN). The strongest inhibition against the RNA-dependent-DNA polymerase (RDDP) activity of RT was observed with the methanol extract of the stem-bark of Peltophorum africanum Sond. (Fabaceae) (IC(50) 3.5 microg/ml), while the methanol extract of the roots of Combretum molle R.Br. ex G. Don (Combretaceae) was the most inhibitory on the ribonuclease H (RNase H) activity (IC(50) 9.7 microg/ml). The known compounds bergenin and catechin, and a red coloured gallotannin composed of meta-depside chains of gallic and protocatechuic acids esterified to a 1-O-isobutyroly-beta-D-glucopyranose core, were isolated from the methanol extract of the roots and stem-bark of Peltophorum africanum. The gallotannin inhibited the RDDP and RNase H functions of RT with IC(50) values of 6.0 and 5.0 microM, respectively, and abolished the 3'-end processing activity of IN at 100 microM. Catechin showed no effect on RT but had a moderate activity on HIV-1 IN. Bergenin was inactive on both enzymes. The aqueous and methanol extracts were non-toxic in a HeLaP4 cell line at a concentration of 400 microg/ml.

Characterization of human immunodeficiency virus type 1 from a previously unexplored region of South Africa with a high HIV prevalence.

HIV prevalence in the Limpopo Province has increased rapidly within the past 10 years, as in other parts of South Africa. Little is known about the genetic and biological properties of HIV circulating in this region including the baseline drug resistance profiles. We therefore collected blood samples from 42 HIV-1-infected patients residing in this region for analysis. All samples were shown to belong to HIV-1 subtype C by env and gag heteroduplex mobility assay (HMA). Viral isolates from 14 of these patients were shown to use the CCR5 coreceptor exclusively and had gp120 V3 loop sequences consistent with this phenotype. Sequence analysis of both protease and reverse transcriptase genes showed that none of 13 isolates harbored primary resistance mutations. These data suggest that HIV-1 subtype C is the predominant subtype circulating in the Limpopo Province, and that viral strains from this region are indistinguishable from those found in other parts of South Africa.

Diarrhoeagenic bacterial pathogens in HIV-positive patients with diarrhoea in rural communities of Limpopo Province, South Africa.

Potential enteric bacterial pathogens in 60 HIV-positive patients with chronic diarrhoea in rural communities of the Limpopo Province, South Africa, were identified using standard microbiological methods. The Kirby-Bauer disk-diffusion method was employed to determine antibiograms of isolated bacteria. Results revealed that diarrhoeagenic bacterial agents were isolated from 48 (80%) of the 60 HIV-positive patients with diarrhoea. Forty-four (73.3%) and 16 (26.7%) of the 60 patients were female and male respectively in the age range of 17-55 years with a mean of 34 years. Bacterial pathogens isolated comprised Campylobacter species (20.0%), Plesiomonas shigelloides (16.6%), Aeromonas species (13.3%), and Escherichia coli, Shigella and Salmonella species (10.0% each). No attempts were made to isolate parasites, fungi, or viruses. Antibiotic susceptibility profiles revealed resistance of the isolates to ampicillin, cephalothin, chloramphenicol, erythromycin, and streptomycin. However, all (100%) of P. shigelloides and Salmonella species were sensitive to nalidixic acid and ciprofloxacin. Most isolates were susceptible to nalidixic acid, ciprofloxacin, and gentamicin, indicating the usefulness of these drugs, although antibiograms may not always correlate with clinical usefulness.