Transmitted/Founder HIV-1 Subtype C Viruses Show Distinctive Signature Patterns in Vif, Vpr, and Vpu That Are Under Subsequent Immune Pressure During Early Infection.

Viral variants that predominate during early infection may exhibit constrained diversity compared with those found during chronic infection and could contain amino acid signature patterns that may enhance transmission, establish productive infection, and influence early events that modulate the infection course. We compared amino acid distributions in 17 patients recently infected with HIV-1C with patients with chronic infection. We found significantly lower entropy in inferred transmitted/founder (t/f) compared with chronic viruses and identified signature patterns in Vif and Vpr from inferred t/f viruses. We investigated sequence evolution longitudinally up to 500 days postseroconversion and compared the impact of selected substitutions on predicted human leukocyte antigen (HLA) binding affinities of published and predicted cytotoxic T-lymphocyte epitopes. Polymorphisms in Vif and Vpr during early infection occurred more frequently at epitope-HLA anchor residues and significantly decreased predicted epitope-HLA binding. Transmission-associated sequence signatures may have implications for novel strategies to prevent HIV-1 transmission.

tat Exon 1 exhibits functional diversity during HIV-1 subtype C primary infection.

Human immunodeficiency virus type 1 (HIV-1) Tat is a mediator of viral transcription and is involved in the control of virus replication. However, associations between HIV-1 Tat diversity and functional effects during primary HIV-1 infection are still unclear. We estimated selection pressures in tat exon 1 using the mixed-effects model of evolution with 672 viral sequences generated from 20 patients infected with HIV-1 subtype C (HIV-1C) over 500 days postseroconversion. tat exon 1 residues 3, 4, 21, 24, 29, 39, and 68 were under positive selection, and we established that specific amino acid signature patterns were apparent in primary HIV-1C infection compared with chronic infection. We assessed the impact of these mutations on long terminal repeat (LTR) activity and found that Tat activity was negatively affected by the Ala(21) substitution identified in 13/20 (65%) of patients, which reduced LTR activity by 88% (± 1%) (P < 0.001). The greatest increase in Tat activity was seen with the Gln(35)/Lys(39) double mutant that resulted in an additional 49% (± 14%) production of LTR-driven luciferase (P = 0.012). There was a moderate positive correlation between Tat-mediated LTR activity and HIV-1 RNA in plasma (P = 0.026; r = 0.400) after 180 days postseroconversion that was reduced by 500 days postseroconversion (P = 0.043; r = 0.266). Although Tat activation of the LTR is not a strong predictor of these clinical variables, there are significant linear relationships between Tat transactivation and patients' plasma viral loads and CD4 counts, highlighting the complex interplay between Tat mutations in early HIV-1C infection.

Viral diversity and diversification of major non-structural genes vif, vpr, vpu, tat exon 1 and rev exon 1 during primary HIV-1 subtype C infection.

To assess the level of intra-patient diversity and evolution of HIV-1C non-structural genes in primary infection, viral quasispecies obtained by single genome amplification (SGA) at multiple sampling timepoints up to 500 days post-seroconversion (p/s) were analyzed. The mean intra-patient diversity was 0.11% (95% CI; 0.02 to 0.20) for vif, 0.23% (95% CI; 0.08 to 0.38) for vpr, 0.35% (95% CI; -0.05 to 0.75) for vpu, 0.18% (95% CI; 0.01 to 0.35) for tat exon 1 and 0.30% (95% CI; 0.02 to 0.58) for rev exon 1 during the time period 0 to 90 days p/s. The intra-patient diversity increased gradually in all non-structural genes over the first year of HIV-1 infection, which was evident from the vif mean intra-patient diversity of 0.46% (95% CI; 0.28 to 0.64), vpr 0.44% (95% CI; 0.24 to 0.64), vpu 0.84% (95% CI; 0.55 to 1.13), tat exon 1 0.35% (95% CI; 0.14 to 0.56 ) and rev exon 1 0.42% (95% CI; 0.18 to 0.66) during the time period of 181 to 500 days p/s. There was a statistically significant increase in viral diversity for vif (p = 0.013) and vpu (p = 0.002). No associations between levels of viral diversity within the non-structural genes and HIV-1 RNA load during primary infection were found. The study details the dynamics of the non-structural viral genes during the early stages of HIV-1C infection.

Microbiological characterisation of southern African medicinal and cosmetic clays.

The effects of traditionally used medicinal and cosmetic clays in southern Africa on selected microorganisms were studied using microbiological media. The clay pH, microchemical composition, kind of associated microorganisms and antimicrobial activity of clays against test microorganisms were determined. The clays contained varying numbers of microorganisms which ranged from 0 up to 105 CFU/g. Clay pH ranged from 2.3-8.9. Neither Escherichia coli, nor other faecal coliforms were detected. Clays of pH value of <4 displayed antimicrobial activities. Clays which were active against test microorganisms had Na(2)O, Al(2)O(3), SiO(2), SO(3), CuO or Cl(2)O as major components. Microbial activity of clays was attributed mainly to low pH but cations such as Cu, Al, S or Cl and various anions might have contributed to the microbicidal effects. No antimicrobial activity was established for many of the clays commonly used in the treatment of common ailments of microbial origin.

Presence of Bacillus cereus in street foods in Gaborone, Botswana.

The purpose of this study was to evaluate the microbiological safety and quality of street foods sold in Gaborone, Botswana. A total of 148 point-of-sale composite street food samples were bought and analyzed between June 2001 and May 2002. The analysis focused on the level of contamination of various street foods with Bacillus cereus. The B. cereus (vegetative and spores), total spore, and total viable counts were determined on all the samples. Also B. cereus isolates from 444 individual point-of-sale food samples were characterized with respect to their biochemical profiles and enterotoxigenic properties. The B. cereus contamination rate for point-of-sale foods was 65%. The B. cereus counts ranged from not detectable to levels as high as 9.1 log CFU/g. Despite the high rate of contamination of some samples, generally, most samples had B. cereus counts of less than 4 log CFU/g; hence, they were of acceptable microbiological quality. Bacillus diarrheal enterotoxin was detected from 52 isolates from individual portions of meals using the B. cereus enterotoxin reversed passive latex agglutination kit. Results of the assay revealed that 59.6% of the B. cereus isolates were enterotoxigenic. Most of the enterotoxigenic isolates were obtained from vegetable samples.

Prevalence and antibiotic resistance profiles of Escherichia coli O157:H7 in beef products from retail outlets in Gaborone, Botswana.

Four hundred meat samples (134 meat cubes, 133 minced meat, 133 fresh sausages) were collected from 15 supermarkets and butcheries in Gaborone, Botswana, between the summer months of October 2002 and March 2003. Samples were assayed for Escherichia coli O157 by selective enrichment in modified E. coli broth containing novobiocin, followed by immunomagnetic separation and plating onto sorbitol MacConkey agar supplemented with potassium tellurite. The isolates were biochemically and serologically confirmed by API 20E and O157 antisera, respectively. The prevalence rates for E. coli O157 were 5.22% in meat cube samples, 3.76% in minced meat samples, and 2.26% in fresh sausages. The isolates showed single, double, and triple antibiotic resistance. Fifty-three percent of them were resistant to cephalothin. Resistance was also recorded for sulphatriad (33%), colistin sulphate (26%), streptomycin (0.7%), and tetracycline (26%). It is recommended that the cause for antibiotic resistance be investigated using a larger number of samples from cattle, especially from ranching areas of the country.

Prevalence of Salmonella in two Botswana abattoir environments.

A 1-year study was carried out to investigate the prevalence of Salmonella in two abattoir environments coded "A" and "B" in Gaborone, Botswana. The total number of environmental samples collected from abattoirs A and B was 250 and 300, respectively. The samples were taken from soils in the corrals, knife blades, saw blades, cattle-drinking water, cattle feces, and feed. Preenrichment, enrichment, and selective/differential media, which enabled the favorable growth of Salmonella, were used in the study. Salmonellae were present in all sampled environments. The most common serotypes found in the environment at abattoir A were E1, C1, C2, and B. Serotypes B, C1, C2, C3, and E1 were common in abattoir B. Antigenic characterization of the salmonellae isolates showed that Salmonella Anatum, Salmonella Azteca, Salmonella Saintpaul, Salmonella Cerro, and Salmonella Westhampton were predominant in abattoir A, whereas Salmonella Anatum, Salmonella Mbandaka, Salmonella Molade, Salmonella Reading, and Salmonella Oranienburg were dominant in abattoir B. Implementing hazard analysis critical control point principles in work procedures would definitely reduce the gross contamination taking place in abattoirs.

Antimicrobial flavonoids from Bolusanthus speciosus.

A new isoflavanone namely 3,5,7,2',4'-pentahydroxy-8,3'-di(gamma,gamma-dimethylallyl)isoflavanone (bolusanthin II) and four new pterocarpans identified as 3-hydroxy-6',6'-dimethylpyrano[2',3':1,2] [6a R,11a R]-8,9-methylenedioxypterocarpan (bolucarpan A), 3-hydroxy-6',6'-dimethyl-4',5'-dihydropyrano[2',3':1,2][6a R,11a R]- 8,9-methylenedioxypterocarpan (bolucarpan B), 3-hydroxy-9-methoxy-6',6'-dimethylpyrano-[2',3':1,2][6a R,11a R]-pterocarpan (bolucarpan C) and 3-hydroxy-9-methoxy-6',6'-dimethyl-4',5'-dihydropyrano[2',3':1,2][6a R,11a R]-pterocarpan (bolucarpan D) and three known isoflavonoids were isolated from the methanolic extracts of the root bark, while eight known isoflavonoids were isolated from the stem bark of Bolusanthus speciosus. These compounds showed antimicrobial activity against Bacillus subtilis, Staphylococcus aureus, Escherichia coli, Saccharomyces cerevisiae and Candida mycoderma using the agar overlay technique.

Erythrinaline alkaloids and antimicrobial flavonoids from Erythrina latissima.

The seed pods of Erythrina latissima yielded erysotrine, erysodine, syringaresinol, vanillic acid, a new erythrina alkaloid, (+)-10,11-dioxoerysotrine, which was lethal to brine shrimp and 2-(5'-hydroxy-3'-methoxy phenyl)-6-hydroxy-5-methoxybenzofuran, which showed strong antimicrobial activity against the yeast spores, Gram-positive and Gram-negative bacteria. The root bark gave four known pterocarpans which showed moderate to strong antifungal activity against the yeast spores and three known flavonoids showed antimicrobial activity against all test microorganisms.

The Microbiology of Phane, an Edible Caterpillar of the Emperor Moth, Imbrasia belina.

The larvae of Imbrasia belina (Westwood) are cooked and sun dried to make a product known as phane, which is consumed as a delicacy. A study was conducted to determine the sanitary quality of phane and the kinds of microorganisms associated with it. It also looked into the potential for the existence of health risk associated with its consumption. Laboratory- and field-processed phane and that from open markets were subjected to microbiological analyses. The total microbial population for the larvae was in the range of 3 × 105 to 2 × 107 CFU/g. Species belonging to seven genera of bacteria and five genera of fungi were isolated from the larvae. About 50% of the identified bacteria were gram-positive, yet their combined population was much lower than that of the gram-negative bacteria. Cooking (89 to 93°C) under both sets of conditions (laboratory and field) reduced the microbial population to less than 9 × 103 CFU/g. The survivors were mostly sporeformers. Laboratory-processed phane was contaminated during drying, but none of the isolates were coliforms and the population increment was marginal. Field-processed phane, on the other hand, had a population of 4 × 104 to 1 ×108 CFU/g after 24 h of drying. The high moisture content of phane (55%) and a high degree of contamination from the soil and air appeared to have contributed to the increased population. Aspergilli including A. flavus and phycomycetes were frequently isolated from the samples. Coliforms were present in 30% and 50% of the phane processed in the field and in market phane, respectively. Escherichia coli and Klebsiella pneumoniae were found in 33% and 21%, respectively, of samples acquired from the market. The presence of K. pneumoniae . E. coli , a toxin-producing sporeformer ( Bacillus cereus ), and mycotoxin-producing fungi ( A. flavus , Penicillium sp., and Fusarium sp.) all point to the possible existence of health risks associated with its consumption.

Prevalence of Coliforms in Traditionally Dried Leafy Vegetables Sold in Open Markets and Food Stores in Gaborone, Botswana.

The aerobic mesophilic count, coliform count, and prevalence of Escherichia coli on two vegetables commonly consumed by a majority of people in Botswana were determined. The aerobic count was found to be >105 CFU/g in both kinds of vegetables and as many as 27 to 30% of the samples had counts >107 CFU/g. Escherichia coli was present in 82% of the Amaranthus leaves sold as morogo in food stores and 50% of the Phaseolus leaves purchased from open markets. Nonfecal coliforms were present in all samples. The nonfecal coliforms belong to species of Klebsiella and Enterobacter and were present in the samples at values of 1 × 101 to 4 × 104 CFU/g.