Clonality and genetic profiles of drug-resistant Mycobacterium tuberculosis in the Eastern Cape Province, South Africa.

In this study, we investigated the diversity of drug-resistant Mycobacterium tuberculosis isolates from families who own cattle in the Eastern Cape Province of South Africa using spoligotyping and mycobacterial interspersed repetitive-unit-variable number tandem repeat (MIRU-VNTR) typing. The Mycobacterium tuberculosis was investigated using MIRU-VNTR and the Mycobacterium tuberculosis families were evaluated using spoligotyping. Spoligotyping grouped 91% of the isolates into seven clusters, while 9% of the deoxyribonucleic acid (DNA) from TB isolates were unclustered from a total of 154 DNA used. Previously described shared types were observed in 89.6% of the isolates, with the Beijing family, SIT1, the principal genotype in the province, while the families T, SIT53 and X1, SIT1329 were the least detected genotypes. MIRU-VNTR grouped 81% of the isolates in 23 clusters while 19% were unclustered. A combination of the VNTR and spoligotyping grouped 79% of the isolates into 23 clusters with 21% unclustered. The low level of diversity and the clonal spread of drug-resistant Mycobacterium tuberculosis isolates advocate that the spread of TB in this study may be instigated by the clonal spread of Beijing genotype. The results from this study provide vital information about the lack of TB control and distribution of Mycobacterium tuberculosis complex strain types in the Eastern Cape Province of South Africa.

The Limpopo Non-Metropolitan Drinking Water Supplier Response to a Diagnostic Tool for Technical Compliance.

Water services providers should supply water that is fit for human consumption, taking into account multi-barrier approaches and technical aspects such as design aspects, operation monitoring, final water quality compliance monitoring, plant monitoring practices, maintenance, and risk management practices. Against this background, this study focused on applying the diagnostic tool for technical compliance as well as assessing the compliance of water treatment plants with management norms. Six plants in the Vhembe District Municipality were selected; the Vondo, Malamulele, Mutshedzi, and Mutale plants (conventional), and the Dzingahe and Tshedza package plants. During the first assessment, four (Malamulele, Mutshedzi, Mutale and Dzingahe) plants scored between 44% and 49% and achieved Class 3 certification, revealing serious challenges requiring immediate intervention. Two water plants (Vondo and Tshedza, scoring 53% and 63%, respectively) were in the Class 2 category, revealing serious challenges requiring attention and improvement. During the second assessment, all plants scored between 63% and 87% (Class 2 category). The greatest improvement (30%) was noted for the Dzingahe and Tshedza plants, followed by the Malamulele plant, while the Mutale, Vondo, and Mutshedzi plants improved their scores by 20%, 17% and 14%, respectively. After corrective actions and re-measurement, no plant complied. It is recommended that Water Services Providers (WSPs) regularly apply the diagnostic tools and water safety plans as developed in order to comply with applicable standards.

Antibacterial activity of crude extracts of some South African medicinal plants against multidrug resistant etiological agents of diarrhoea.

BACKGROUND: This study evaluated the antibacterial activity of some plants used in folklore medicine to treat diarrhoea in the Eastern Cape Province, South Africa. METHODS: The acetone extracts of Acacia mearnsii De Wild., Aloe arborescens Mill., A. striata Haw., Cyathula uncinulata (Schrad.) Schinz, Eucomis autumnalis (Mill.) Chitt., E. comosa (Houtt.) Wehrh., Hermbstaedtia odorata (Burch. ex Moq.) T.Cooke, Hydnora africana Thunb, Hypoxis latifolia Wight, Pelargonium sidoides DC, Psidium guajava L and Schizocarphus nervosus (Burch.) van der Merwe were screened against Staphylococcus aureus, Escherichia coli, Enterococcus faecalis, multi-resistant Salmonella enterica serovar Isangi, S. typhi, S. enterica serovar Typhimurium, Shigella flexneri type 1b and Sh. sonnei phase II. A qualitative phytochemical screening of the plants extracts was by thin layer chromatography. Plants extracts were screened for antibacterial activity using serial dilution microplate technique and bioautography. RESULTS: The TLC fingerprint indicated the presence of terpenoids and flavonoids in the herbs. Most of the tested organisms were sensitive to the crude acetone extracts with minimum inhibitory concentration (MIC) values ranging from 0.018-2.5 mg/mℓ. Extracts of A. striata, C. uncinulata, E. autumnalis and P. guajava were more active against enteropathogens. S. aureus and Sh. flexneri were the most sensitive isolates to the crude extracts but of significance is the antibacterial activity of A. arborescens and P. guajava against a confirmed extended spectrum betalactamase positive S. enterica serovar Typhimurium. CONCLUSION: The presence of bioactive compounds and the antibacterial activity of some of the selected herbs against multidrug resistant enteric agents corroborate assertions by traditional healers on their efficacies.

Antimycobacterial Activity and Low Cytotoxicity of Leaf Extracts of Some African Anacardiaceae Tree Species.

Treatment of tuberculosis (TB) is a challenge because of multidrug-resistant and extremely drug-resistant strains of Mycobacterium tuberculosis. Plant species contain antimicrobial compounds that may lead to new anti-TB drugs. Previous screening of some tree species from the Anacardiaceae family revealed the presence of antimicrobial activity, justifying further investigations. Leaf extracts of 15 Anacardiaceae tree species were screened for antimycobacterial activity using a twofold serial microdilution assay against the pathogenic Mycobacterium bovis and multidrug resistant M. tuberculosis and rapidly growing mycobacteria, Mycobacterium smegmatis, Mycobacterium fortuitum and Mycobacterium aurum. The vaccine strain, M. bovis and an avirulent strain, H37Ra M. tuberculosis, were also used. Cytotoxicity was assessed using a colorimetric assay against Vero kidney, human hepatoma and murine macrophage cells. Four out of 15 crude acetone extracts showed significant antimycobacterial activity with minimum inhibitory concentration varying from 50 to 100 µg/mL. Searsia undulata had the highest activity against most mycobacteria, followed by Protorhus longifolia. M. fortuitum was the strongest predictor of activity against multidrug-resistant TB (correlation coefficient = 0.65). Bioautography against M. aurum and M. fortuitum worked well as indicators of the Rf values of active compounds yielding strong zones of inhibition. The leaf extracts of S. undulata and P. longifolia had more than ten different antimycobacterial compounds and had low cytotoxicity with LC50 values above 100 µg/mL. Copyright © 2016 John Wiley & Sons, Ltd.

Molecular basis of virulence in clinical isolates of Escherichia coli and Salmonella species from a tertiary hospital in the Eastern Cape, South Africa.

BACKGROUND: Apart from localized gastrointestinal infections, Escherichia coli and Salmonella species are major causes of systemic disease in both humans and animals. Salmonella spp. cause invasive infections such as enteric fever, septicemia, osteomyelitis and meningitis while certain types of E. coli can cause systemic infections, includingpyelonephritis, meningitis and septicemia. These characteristic requires the involvement of a myriad of virulence factors. METHODS: This study investigated the virulence factors of Escherichia coli and Salmonella species in clinical specimens from patients with diarrhoea presenting to health care centres in Oliver R. Tambo District Municipality, Eastern Cape Province, Republic of South Africa. Microbiology analysis involved the use of cultural and molecular techniques. RESULTS: Out of a total of 315 samples screened, Salmonella isolates were obtained in 119 (37.8%) of cases and these comprised: S. choleraesuis (6%), S. enteritidis (4%), S. eppendorf (1%), S. hadar (1%), S. isangi (8%), S. panama (1%), S. typhi (52%), S. typhimurium (25%) and untyped Salmonella spp. (2%). Among the Salmonella species 87 (73.1%) were invasive. Using molecular diagnostic methods, diarrheagenic E. coli were detected in 90 cases (28.6%): the greater proportion of this were enteroaggregative E. coli (EAEC) 37 (41.1%), enteropathogenic E. coli (EPEC) 21 (23.3%) and enterohemorrhagic E. coli (EHEC) 21 (23.3%). The predominant virulence gene among the diarrheagenic E. coli was EAEC heat-stable enterotoxin astA genes while the virulence genes identified in the Salmonella strains were 15 (12.6%) flic and 105 (88.2%) inv genes. The amino acid identity of the representative genes showed 95-100% similarity to corresponding blast searched sequence. CONCLUSIONS: This study showed the diversity of virulence gene expression in two major enteric pathogens. S. typhi and enteroaggregative E. coli were the predominant enteropathogens in our study area with an indication that EAEC is endemic within our study population. It was observed among other things that some diarrheagenic E. coli isolated from apparently asymptomatic subjects expressed some virulence genes at frequency as high as seen in diarrheagenic cases. This study underlines the importance of understanding the virulence composition and diversity of pathogens for enhanced clinico-epidemiological monitoring and health care delivery.

Seasonal abundance and distribution of Vibrio species in the treated effluent of wastewater treatment facilities in suburban and urban communities of Eastern Cape Province, South Africa.

We assessed the seasonal abundance and distribution of Vibrio species as well as some selected environmental parameters in the treated effluents of two wastewater treatment plants (WWTP), one each located in a suburban and urban community of Eastern Cape Province, South Africa. Vibrio population density ranged from 2.1 × 10(5) to 4.36 × 10(4) CFU/ml in the suburban community and from 2.80 × 10(5) to 1.80 × 10(5) CFU/ml in the urban community. Vibrio species associated with 180 µ, 60 µ, and 20 µ plankton sizes were observed at densities of 0-136 × 10(3) CFU/ml, 0-8.40 × 10(2) CFU/ml, and 0-6.80 × 10(2) CFU/ml, respectively at the suburban community's WWTP. In the urban community, observed densities of culturable Vibrio were 0-2.80 × 10(2) CFU/ml (180 µ), 0-6.60 × 10(2) CFU/ml (60 µm), and 0-1.80 × 10(3) CFU/ml (20 µm). The abundance of free-living Vibrio species ranged from 0 to 1.0 × 10(2) and 1.0 × 10(3) CFU/ml in the suburban and urban communities' WWTPs, respectively. Molecular confirmation of the presumptive Vibrio isolates revealed the presence of V. fluvialis (41.38%), V. vulnificus (34.48%), and V. parahaemolyticus (24.14%) in the suburban community effluents. In the urban community molecular confirmation revealed that the same species were present at slightly different percentages, V. fluvialis (40%), V. vulnificus (36%), and V. parahaemolyticus (24%). There was no significant correlation between Vibrio abundance and season, either as free-living or plankton-associated entities, but Vibrio species abundance was positively correlated with temperature (r=0.565; p<0.01), salinity, and dissolved oxygen (p<0.05). Turbidity and pH showed significant seasonal variation (p<0.05) across the seasons in both locations. This study underscores the potential of WWTPs to be sources of Vibrio pathogens in the watershed of suburban and urban communities in South Africa.

Evaluation of the antibacterial and anticancer activities of some South African medicinal plants.

BACKGROUND: Several herbs are traditionally used in the treatment of a variety of ailments particularly in the rural areas of South Africa where herbal medicine is mainly the source of health care system. Many of these herbs have not been assessed for safety or toxicity to tissue or organs of the mammalian recipients. METHODS: This study evaluated the cytotoxicity of some medicinal plants used, inter alia, in the treatment of diarrhoea, and stomach disorders. Six selected medicinal plants were assessed for their antibacterial activities against ampicillin-resistant and kanamycin-resistant strains of Escherichia coli by the broth micro-dilution methods. The cytotoxicities of methanol extracts and fractions of the six selected plants were determined using a modified tetrazolium-based colorimetric assay (3-(4, 5-dimethylthiazol)-2, 5-diphenyl tetrazolium bromide (MTT) assay). RESULTS: The average minimum inhibitory concentration (MIC) values of the plants extracts ranged from 0.027 mg/mℓ to 2.5 mg/mℓ after 24 h of incubation. Eucomis autumnalis and Cyathula uncinulata had the most significant biological activity with the least MIC values. The in vitro cytotoxicity assay on human hepatocarcinoma cell line (Huh-7) revealed that the methanol extract of E. autumnalis had the strongest cytotoxicity with IC(50) of 7.8 µg/mℓ. Ethyl acetate and butanol fractions of C. uncinulata, Hypoxis latifolia, E. autumnalis and Lantana camara had lower cytotoxic effects on the cancer cell lines tested with IC(50) values ranging from 24.8 to 44.1 µg/mℓ; while all the fractions of Aloe arborescens and A. striatula had insignificant or no cytotoxic effects after 72 h of treatment. CONCLUSIONS: Our results indicate that the methanol fraction of E. autumnalis had a profound cytotoxic effect even though it possessed very significant antibacterial activity. This puts a query on its safety and hence a call for caution in its usage, thus a product being natural is not tantamount to being entirely safe. However, the antibacterial activities and non-cytotoxic effects of A. arborescens and A. striatula validates their continuous usage in ethnomedicine.

Inhibitory properties of selected South African medicinal plants against Mycobacterium tuberculosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB) is the most commonly notified disease and the fifth largest cause of mortality. One in 10 cases is resistant to treatment in some areas. Several plants are used locally to treat TB-related disease. AIMS OF THE STUDY: The aim was to screen selected South African medicinal plants used to treat TB and related symptoms by traditional healers for antimycobacterial activity. MATERIALS AND METHODS: Ethnobotanical information on these plants was obtained. Crude acetone, methanol, hexane and ethanol extracts of 21 selected medicinal plants obtained in Venda, South Africa were screened for their ability to inhibit MTB H(37)Ra and a clinical strain resistant to first-line drugs and one second-line drug using tetrazolium microplate assay to determine the minimum inhibitory concentration (MIC). Results were analyzed using Microsoft Excel 2007 and One way ANOVA; p<0.05 was considered for statistical significance. RESULTS: Few acetone extracts were active against MTB with MIC under 100 microg/mL. Four plants showed lower MIC values; Berchemia discolor Klotzsch Hemsl 12, 5 microg/mL on H(37)Ra and 10.5 microg/mL on the clinical isolate, Bridelia micrantha Hochst. Baill (25 microg/mL), Warbugia salutaris Bertol. F Chiov (25 microg/mL), and Terminalia sericea Burch ex D. F (25 microg/mL) on both H(37)Ra and clinical isolate. However, the roots of Ximenia caffra Sond. Var. caffra, barks of Sclerocarya birrea (A Rich) Hochst, Asclepias fruticosa L, tubers of Allium sativum L, leaves of Carica papaya L, Solanum panduriforme E. Mey C, and roots of Securidaca longepedunculata Fresen gave MIC greater than 100 microg/mL. CONCLUSION: The acetone extracts of Berchemiadiscolor, Bridelia micrantha, Terminalia sericea and Warbugia salutaris could be important sources of mycobactericidal compounds against multidrug-resistant MTB.

PCR detection of Clostridium difficile triose phosphate isomerase (tpi), toxin A (tcdA), toxin B (tcdB), binary toxin (cdtA, cdtB), and tcdC genes in Vhembe District, South Africa.

Specific polymerase chain reaction (PCR) protocols were used to determine the prevalence of toxigenic Clostridium difficile in Vhembe, South Africa. Of 322 stool samples collected, toxigenic C. difficile was found in 23 (7.1%) cases and was significantly associated with diarrhea 20 (11.4%) compared with 3 (2%) in non-diarrheal samples (chi(2) = 426, P = 0.001), intestinal inflammation in 18 (12.1%) compared with 5 (2.9%) in lactoferrin-negative samples (chi(2) = 10.194, P = 0.001), and occult blood in 19 (16%) compared with 4 (2%) in occult blood-negative samples (chi(2) = 22.157, P < 0.001). Toxigenic C. difficile was more common among individuals > 50 years of age (20%), followed by those between 30 and 39 years of age (19%) and was not associated with HIV infections (chi(2) = 0.289, P = 0.591). Co-infection with other pathogens was common. Multivariate analysis indicated that toxigenic C. difficile was associated with E. bieneusi (P = 0.028), C. parvum (P = 0.007), and Enteroaggregative Escherichia coli (EAEC) (P = 0.007) in diarrheal samples. This study confirms the usefulness of PCR methodologies in the detection of toxigenic C. difficile and suggests that C. difficile is responsible for a small, but underappreciated, proportion of diarrheal cases in the region, and further study is warranted in this area.

Importance of anti-Plesiomonas antibody in the protection of rats against infection by Plesiomonas Shigelloides.

Rabbit anti-antibody raised against Plesiomonas shigelloides infections using standard immunological procedures. Ten (50%) of 20 rats died when only P. shigelloidess was injected while 2(10%) died [P < 0.05]; following injection of another group of 20 rats with P. shigelloides 24hours after administration of rabbit-Plesiomonas anti-antibody. In another scheme, 14(70%) of 20 rats died when p.shigelloides and colloidal carbon particles (CCP) were administered while only 4(20%) died (p<0.05) following injection of P. shigelloides and CCP 24 hours after administration of rabbit-Plesiomonas anti-antibody. No death was recorded for control rats which received only CCP. This study highlights a novel approach in the management or prevention of P. shigelloides infection in our environment.