Roland N. Ndip

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.

Marked susceptibility of South African Helicobacter pylori strains to ciprofloxacin and amoxicillin: Clinical implications

OBJECTIVES Helicobacter pylori-associated infection is common in South Africa, as in other developing countries. Antibiotic resistance is recognised as a major cause of treatment failure. We studied the susceptibility and resistance patterns of H. pylori to guide empiric treatment and prevent the emergence of resistance. METHODS Two hundred H. pylori strains obtained from gastric biopsies of patients presenting with gastric-related morbidities attending Livingstone Hospital, Port Elizabeth, were evaluated for their susceptibility to seven antibiotics - metronidazole, clarithromycin, tetracycline, amoxicillin, gentamicin, ciprofloxacin and erythromycin. H. pylori was isolated following standard microbiology procedures, and susceptibility determined using the Kirby-Bauer disc diffusion and agar dilution methods. Comparisons of antimicrobial resistance rates with sex of the patients were determined using the chi-square test; a p-value of <0.05 was considered significant. RESULTS Marked susceptibility was observed for ciprofloxacin (100%) and amoxicillin (97.5%), and good activity for clarithromycin (80%) and gentamicin (72.5%). However, marked resistance (95.5%) was observed for metronidazole. The minimal inhibitory concentration (MIC) ranged from 0.0625 microg/ml to 8 microg/ml. The lowest MIC, with a range of 0.0625 - 1 microg/ml, was recorded for ciprofloxacin, while the highest (5 - 8 microg/ml) was noted for gentamicin. CONCLUSION Multidrug resistance was commonly encountered - a finding of clinical significance that calls for continuous surveillance of antibiograms to guide empiric treatment. We advocate the inclusion of ciprofloxacin in the treatment regimen of H. pylori infection in our study environment.

Selected South African Honeys and Their Extracts Possess In Vitro Anti-Helicobacter pylori Activity

BACKGROUND AND AIMS Eradication of Helicobacter pylori by triple therapy often results in a failure rate of 10-20%; thus, there is a need to seek alternative treatments. The aim of this study was to screen selected South African honeys for their anti-H. pylori activity, to extract the antimicrobial components using organic solvents and to determine the minimum inhibitory concentrations (MICs) of the extracts. METHODS Three locally produced honeys from different regions in South Africa were screened for anti-H. pylori activity at four different concentrations using the agar well diffusion technique. Subsequently, Pure honey was extracted using n-hexane, diethyl ether, chloroform and ethyl acetate; extracts were also examined for anti-H. pylori activity by agar well diffusion method. The MICs of the three most active extracts were determined both by visual inspection and spectrophotometric analysis at 620 nm using the broth microdilution method. The results were analyzed by one-way ANOVA at 95% significance level. RESULTS All honeys demonstrated anti-H. pylori activity and were most active at 75% v/v. The positive control (clarithromycin) recorded a zone diameter of 18.0 ± 7.4 mm not significantly different (p >0.05) from honeys at 75% v/v and solvent extracts. Chloroform extract recorded the lowest MIC(95) values that ranged from 0.156-5% v/v confirming this extract to be the most active. CONCLUSION All honeys demonstrated anti-H. pylori activity at concentrations ≥10%, as did the solvent extracts. Therefore, these honeys and solvent extracts possess potential compounds with therapeutic activity that could be further exploited as lead molecules in the treatment of H. pylori infections.

Phytochemical studies and antioxidant activity of two South African medicinal plants traditionally used for the management of opportunistic fungal infections in HIV/AIDS patients

BackgroundIt has been observed that perturbations in the antioxidant defense systems, and consequently redox imbalance, are present in many tissues of HIV-infected patients. Hence, the exogenous supply of antioxidants, as natural compounds that scavenge free radicals, might represent an important additional strategy for the treatment of HIV infection. The aim of this study was therefore to analyse the phytochemical constituents and antioxidant potential of Gasteria bicolor Haw and Pittosporum viridiflorum Sims., two South African plants traditionally used for the management of opportunistic fungal infections (OFIs) in AIDS patients.MethodsThe in vitro antioxidant properties of the two plants were screened through DPPH (1,1-diphenyl-2-picrylhydrazyl), NO (nitric oxide), H2O2 (hydrogen peroxide) radical scavenging effects and reducing power assays. Phytochemical studies were done by spectrophotometric techniques.ResultsThere were no significant differences in the flavonoid and proanthocyanidins contents between the leaves and bark extracts of Gasteria bicolor and Pittosporum viridiflorum respectively, while the total phenolic content of the bark extract of P. viridiflorum was significantly higher than that of G. bicolor leaf. The acetone extracts of both plants indicated strong antioxidant activities.ConclusionThe results from this study indicate that the leaves and stem extracts of Gasteria bicolor and Pittosporum viridiflorum respectively possess antioxidant properties and could serve as free radical inhibitors, acting possibly as primary antioxidants. Since reactive oxygen species are thought to be associated with the pathogenesis of AIDS, and HIV-infected individuals often have impaired antioxidant defenses, the inhibitory effect of the extracts on free radicals may partially justify the traditional use of these plants in the management of OFIs in HIV patients in South Africa.

Molecular Detection of Antibiotic Resistance in South African Isolates of Helicobacter pylori

Rapid diagnosis and treatment of Helicobacter pylori (H. pylori) presents a challenge. We aimed at investigating the presence of H. pylori, susceptibility profile, and associated mutations in an effort to validate the effectiveness of GenoType HelicoDR assay in H. pylori typing in our environment. Two hundred and fifty-four biopsy specimens were cultured and DNA extracted from seventy-eight positive cultures using the Qiagen DNA extraction kit. The GenoType Helico DR which employs reverse hybridisation was used to confirm the presence of H. pylori, determination of its susceptibility to antimicrobials, and detection of mutations conferring resistance to clarithromycin and fluoroquinolones. The organism was isolated from 168/254 (66.1 %) of the specimens by culture. Of the 78 strains used for further investigation, 12/78 (15.38%) were resistant to clarithromycin while 66/78 (84.61%) were susceptible. For fluoroquinolone, 70/78 (89.74%) strains were susceptible while 8 (10.26%) were resistant. Mutations were observed in 17 strains with A2147G being the most prevalent; A2146C and D91N were the least. The reverse hybridisation assay is an easy and fast technique in confirming the presence of H. pylori, its antimicrobial profile, and associated mutations. Analysis regarding the suitability of this assay for H. pylori typing is warranted in other regions.

Prevalence of Helicobacter pylori vacA, cagA and iceA genotypes in South African patients with upper gastrointestinal diseases.

Clinical response to Helicobacter pylori infection may be determined by specific virulence-associated genotypes which varies geographically. The aim of this study was to investigate the diversity of putative virulence markers of H. pylori; cagA, vacA and iceA in the Eastern Cape Province of South Africa. One hundred H. pylori strains obtained from dyspeptic patients were used. Gastric biopsies were obtained from 254 dyspeptic patients. H. pylori was cultured and strains were studied. Bacterial genotypes cagA, vacA (s and m subtypes) and iceA were analysed by PCR using specific primers. CagA was identified in 90% of the strains investigated. Fifty-eight of the 100 strains had the vacA signal sequence genotype s1 and 26 had subtype s2. Combined vacA s1/s2 was detected in 16 of the strains. VacA middle region analysis showed that 8 (8%) strains were m1 while 50 were m2. Combined vacA m1/m2 was detected in 36 of the strains. s1m2 (20%) and s2m2 (20%) genotypes were the most common allelic combinations of the vacA gene among the strains. Multiple vacA genotypes were detected in this study. Twenty-six percent of the strains identified had both iceA1 and iceA2. All our strains tested positive for the ureC (glmM) gene. This study reveals a high prevalence of vacA, cagA and iceA2.

Identification and Antibacterial Evaluation of Bioactive Compounds from Garcinia kola (Heckel) Seeds

We assessed the bioactivity of G. kola seeds on Streptococcus pyogenes, Staphylococcus aureus, Plesiomonas shigelloides and Salmonella typhimurium. The crude ethyl acetate, ethanol, methanol, acetone and aqueous extracts were screened by the agar-well diffusion method and their activities were further determined by Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) assays. The extracts were fractionated by Thin Layer Chromatography (TLC). Bioautography was used to assess the activity of the possible classes of compounds present in the more active extracts. Column chromatography was used to purify the active compounds from the mixture, while GC-MS was used to identify the phytocomponents of the fractions. The inhibition zone diameters of the extracts ranged from 0–24 ± 1.1 mm, while MIC and MBC values ranged between 0.04–1.25 mg/mL and 0.081–2.5 mg/mL, respectively. The chloroform/ethyl acetate/formic acid (CEF) solvent system separated more active compounds. The MIC of the fractions ranged between 0.0006–2.5 mg/mL. CEF 3 (F3), CEF 11 (F11) and CEF 12 (F12) revealed the presence of high levels of linoleic acid, 1,2-benzenedicarboxylic acid and 2,3-dihydro-3,5-dihydroxy-6-methyl ester, respectively. The results obtained from this study justify the use of this plant in traditional medicine and provide leads which could be further exploited for the development of new and potent antimicrobials.

Crude Ethanolic Extracts of Garcinia kola Seeds Heckel (Guttiferae) Prolong the Lag Phase of Helicobacter pylori: Inhibitory and Bactericidal Potential

Problems associated with current treatment regimens have generated a considerable interest in alternative approaches for the eradication of Helicobacter pylori infections using phytochemical compounds. In an attempt to identify potential sources of such compounds, the antimicrobial activity of five solvent extracts of Garcinia kola seeds were investigated against 30 clinical strains of H. pylori and a standard control strain, NCTC 11638, using standard microbiological techniques. Metronidazole and amoxicillin were included in these experiments as positive control antibiotics. All the extracts tested exhibited anti-H. pylori activity with zone diameters of inhibition between 0 and 25 mm. The ethanol extract demonstrated considerable anti-H. pylori activity with a percentage susceptibility of 53.3% and minimum inhibitory concentration for 50% susceptibility (MIC₅₀) values ranging from 0.63 to 5.0 mg/mL. Ranges of MIC₅₀ values for amoxicillin and metronidazole were 0.01-0.63 mg/mL and 0.04-5.0 mg/mL, respectively. The inhibitory activity of the ethanol extract was similar to that of metronidazole (P > .05) as opposed to amoxicillin (P < .05). The extract caused a 12-hour extension of the lag phase of H. pylori at 1.25 mg/mL. The same observations were recorded when this concentration was doubled and quadrupled alongside a killing rate of 80.1% and 93.7%, respectively, after 24 hours and of 100% after 30 hours. These results demonstrate that the ethanol extract of G. kola may contain therapeutically useful compounds against H. pylori.

Risk Factors for African Tick-Bite Fever in Rural Central Africa

African tick-bite fever is an emerging infectious disease caused by the spotted fever group Rickettsia, Rickettsia africae, and is transmitted by ticks of the genus Amblyomma. To determine the seroprevalence of exposure to R. africae and risk factors associated with infection, we conducted a cross-sectional study of persons in seven rural villages in distinct ecological habitats of Cameroon. We examined 903 plasma samples by using an indirect immunofluorescence assay for antibodies to R. africae and analyzed demographic and occupational data collected from questionnaires. Of the 903 persons tested, 243 (26.9%) had IgG/IgM/IgA reactive with R. africae. Persons from four of the seven village sites were significantly more likely to be seropositive (P < 0.05), and lowland forest sites tended to have higher seroprevalences. These results suggest that African tick-bite fever is common in adults in rural areas of Cameroon and that ecological factors may play a role in the acquisition of R. africae infection.

Preliminary Phytochemical Screening and In Vitro Anti-Helicobacter pylori Activity of Extracts of the Stem Bark of Bridelia micrantha (Hochst., Baill., Euphorbiaceae)

Helicobacter pylori is a major risk factor for gastritis, ulcers and gastric cancer. This study was aimed to determine the antimicrobial activity of the stem bark of Bridelia. micrantha on H. pylori isolated in South Africa. Extracts and clarithromycin were tested against 31 clinical strains, including a standard strain (NCTC 11638) of H. pylori, by measuring the diameters of the corresponding inhibition zones, followed by determination of the Minimum Inhibitory Concentration (MIC) (using metronidazole, and amoxicillin as control antibiotics) and the rate of kill. Preliminary phytochemical screening was also done. Inhibition zone diameters which ranged from 0–23 mm were observed for all five of the extracts and 0–35 mm for clarithromycin. Marked susceptibility of strains (100%) was noted for the acetone extract (P < 0.05), followed by ethyl acetate extract (93.5%). The MIC50 values ranged from 0.0048 to 0.156 mg/mL for the ethyl acetate extract and 0.0048 to 0.313 mg/mL for the acetone extract. The MIC90 values ranged from 0.0048 to 2.5 mg/mL for the ethyl acetate extract and 0.078 to >0.625 mg/mL for the acetone extract, respectively. Insignificant statistical difference in potency was observed when comparing the crude ethyl acetate extract to metronidazole and amoxicillin (P > 0.05). Complete killing of strain PE430C by the ethyl acetate extract was observed at 0.1 mg/mL (2 × MIC) and 0.2 mg/mL (4 × MIC) at 66 and 72 h. For strain PE369C, 100% killing was observed at 0.1 mg/mL (2 × MIC) in 66 and 72 h. The ethyl acetate extract could thus be a potential source of lead molecules for the design of new anti-Helicobacter pylori therapies as this study further confirmed the presence of phytochemicals including alkaloids, flavonoids, steroids, tannins and saponins.