Pre-referral Rectal Artesunate Treatment by Community-Based Treatment Providers in Ghana, Guinea-Bissau, Tanzania, and Uganda (Study 18): A Cluster-Randomized Trial.

BACKGROUND: If malaria patients who cannot be treated orally are several hours from facilities for injections, rectal artesunate prior to hospital referral can prevent death and disability. The goal is to reduce death from malaria by having rectal artesunate treatment available and used. How best to do this remains unknown. METHODS: Villages remote from a health facility were randomized to different community-based treatment providers trained to provide rectal artesunate in Ghana, Guinea-Bissau, Tanzania, and Uganda. Prereferral rectal artesunate treatment was provided in 272 villages: 109 through community-based health workers (CHWs), 112 via trained mothers (MUMs), 25 via trained traditional healers (THs), and 26 through trained community-chosen personnel (COMs); episodes eligible for rectal artesunate were established through regular household surveys of febrile illnesses recording symptoms eligible for prereferral treatment. Differences in treatment coverage with rectal artesunate in children aged <5 years in MUM vs CHW (standard-of-care) villages were assessed using the odds ratio (OR); the predictive probability of treatment was derived from a logistic regression analysis, adjusting for heterogeneity between clusters (villages) using random effects. RESULTS: Over 19 months, 54 013 children had 102 504 febrile episodes, of which 32% (31 817 episodes) had symptoms eligible for prereferral therapy; 14% (4460) children received treatment. Episodes with altered consciousness, coma, or convulsions constituted 36.6% of all episodes in treated children. The overall OR of treatment between MUM vs CHW villages, adjusting for country, was 1.84 (95% confidence interval [CI], 1.20-2.83; P = .005). Adjusting for heterogeneity, this translated into a 1.67 higher average probability of a child being treated in MUM vs CHW villages. Referral compliance was 81% and significantly higher with CHWs vs MUMs: 87% vs 82% (risk ratio [RR], 1.1 [95% CI, 1.0-1.1]; P < .0001). There were more deaths in the TH cluster than elsewhere (RR, 2.7 [95% CI, 1.4-5.6]; P = .0040). CONCLUSIONS: Prereferral episodes were almost one-third of all febrile episodes. More than one-third of patients treated had convulsions, altered consciousness, or coma. Mothers were effective in treating patients, and achieved higher coverage than other providers. Treatment access was low. CLINICAL TRIALS REGISTRATION: ISRCTN58046240.

Screening of selected ethnomedicinal plants from South Africa for larvicidal activity against the mosquito Anopheles arabiensis.

BACKGROUND: This study was initiated to establish whether any South African ethnomedicinal plants (indigenous or exotic), that have been reported to be used traditionally to repel or kill mosquitoes, exhibit effective mosquito larvicidal properties. METHODS: Extracts of a selection of plant taxa sourced in South Africa were tested for larvicidal properties in an applicable assay. Thirty 3rd instar Anopheles arabiensis larvae were exposed to various extract types (dichloromethane, dichloromethane/methanol) (1:1), methanol and purified water) of each species investigated. Mortality was evaluated relative to the positive control Temephos (Mostop; Agrivo), an effective emulsifiable concentrate larvicide. RESULTS: Preliminary screening of crude extracts revealed substantial variation in toxicity with 24 of the 381 samples displaying 100% larval mortality within the seven day exposure period. Four of the high activity plants were selected and subjected to bioassay guided fractionation. The results of the testing of the fractions generated identified one fraction of the plant, Toddalia asiatica as being very potent against the An. arabiensis larvae. CONCLUSION: The present study has successfully identified a plant with superior larvicidal activity at both the crude and semi pure fractions generated through bio-assay guided fractionation. These results have initiated further research into isolating the active compound and developing a malaria vector control tool.

Screening for adulticidal bioactivity of South African plants against Anopheles arabiensis.

BACKGROUND: This study was conducted to evaluate whether a selection of South African ethnomedicinal plants included in this study displayed insecticidal properties when screened against adult stages of the mosquito. METHODS: 381 crude extracts of 80 plant taxa in 42 families were sprayed onto ceramic tiles and screened using the cone bio-assay method for insecticide efficacy testing. Blood-fed, female Anopheles arabiensis mosquitoes were exposed to the treated tiles for a period of sixty minutes. Mosquito mortality was monitored for twenty-four hours. RESULTS: Of all the extracts analysed, the highest activity was observed in Ptaeroxylon obliquum (Ptaeroxylaceae) and Pittosporum viridiflorum (Pittosporaceae), a single extract from each, exhibiting more than 50% mortality. A large proportion (81.63%) of the extracts tested displayed low levels of mosquitocidal activity. The remainder of the extracts (17.85%) exhibited no bioactivity (0% mortality). CONCLUSIONS: The screening results have shown that in accordance with WHO standards, none of the crude extracts tested had exhibited greater than 60% mortality against the adult stages of the malaria vector Anopheles arabiensis.

Evaluation of selected South African ethnomedicinal plants as mosquito repellents against the Anopheles arabiensis mosquito in a rodent model.

BACKGROUND: This study was initiated to establish whether any South African ethnomedicinal plants (indigenous or exotic), that have been reported to be used traditionally to repel or kill mosquitoes, exhibit effective mosquito repellent properties. METHODS: Extracts of a selection of South African taxa were tested for repellency properties in an applicable mosquito feeding-probing assay using unfed female Anopheles arabiensis. RESULTS: Although a water extract of the roots of Chenopodium opulifolium was found to be 97% as effective as DEET after 2 mins, time lag studies revealed a substantial reduction in efficacy (to 30%) within two hours. CONCLUSIONS: None of the plant extracts investigated exhibited residual repellencies >60% after three hours.

The use of placebo in a trial of rectal artesunate as initial treatment for severe malaria patients en route to referral clinics: ethical issues.

Placebo-controlled trials are controversial when individuals might be denied existing beneficial medical interventions. In the case of malaria, most patients die in rural villages without healthcare facilities. An artesunate suppository that can be given by minimally skilled persons might be of value when patients suddenly become too ill for oral treatment but are several hours from a facility that can give injectable treatment for severe disease. In such situations, by default, no treatment is (or can be) given until the patient reaches a facility, making the placebo control design clinically relevant; alternative bioequivalence designs at the facility would misrepresent reality and risk incorrect conclusions. We describe the ethical issues underpinning a placebo-controlled trial in severe malaria. To protect patients and minimise risk, all patients were referred immediately to hospital so that each had a higher chance of prompt treatment through participation. There was no difference between artesunate and placebo in patients who reached clinic rapidly; among those who could not, a single artesunate suppository significantly reduced death or permanent disability, a finding of direct and indirect benefit to patients in participating villages and elsewhere.

Efficacy of Artemisia afra phytotherapy in experimental tuberculosis.

Artemisia afra [Jacq] (Asteraceae) phytotherapy is widely used for its medicinal properties in traditional practices. In this study we investigated whether extracts of A. afra are capable of controlling mycobacterial replication. For Mycobacterium aurum cultured in the presence of aqueous-, methanol- and dichloromethane (DCM) extracts of A. afra we found that bacterial replication was inhibited by the dichloromethane extract only. Activity of the DCM extract was confirmed in dose-dependent studies against both M. aurum and M. tuberculosis with an IC(50) =270 microg/ml and IC(50) = 290microg/ml, respectively. Fractionation of the DCM extract and evaluation of its efficacy in vitro found that most of the antimycobacterial activity was associated with isolate fraction C8 that contained several sesquiterpene lactones, the most prominent of which are Artemin and Arsubin. Evaluation of the bactericidal efficacy in vitro showed that isolate fraction C8 reduced replication of M. aurum and M. tuberculosis in a dose-dependent manner with IC(50) =1.9 microg/ml and IC(50) = 2.0 microg/ml, respectively, and an MIC = 10 microg/ml. Further, isolate fraction C8 and the DCM extract was administered to M. tuberculosis-infected mice at a tolerated dose of 1000 microg/kg for up to 26 weeks and mycobacterial burdens compared to untreated-, INH/RIF treated- and aqueous-extract-treated animals to assess its bactericidal activity in vivo. Bacterial replication remained unaffected during treatment with either isolate fraction C8 or the DCM extract resulting in pulmonary and splenic bacilli burdens comparable to that of untreated mice. In contrast, INH/RIF treatment cleared M. tuberculosis infection after only 8 weeks to undetectable levels. Interestingly, treatment of M. tuberculosis-infected mice with aqueous extract of A. afra regulated pulmonary inflammation during early infection notwithstanding its inability to inhibit mycobacterial growth. This study clearly demonstrates that A. afra contains in vitro anti-mycobacterial activity, modulates pulmonary inflammation in early mycobacterial infection, and that the mouse experimental tuberculosis model may serve as a useful assay for evaluating the utility of phytotherapy.

Antidiabetic screening and scoring of 11 plants traditionally used in South Africa.

AIM: To investigate the traditional antidiabetic uses of indigenous or naturalised South African plants using an optimised screening and scoring method. MATERIALS AND METHODS: Eleven plant species were screened against Chang liver, 3T3-L1 adipose and C2C12 muscle cells measuring glucose utilisation in all three cell lines and toxicity in the hepatocytes and adipocytes only. A scoring system was devised to aid interpretation of results. RESULTS: Catharanthus roseus results correlated with previously reported in vivo results, with best stimulation of glucose utilisation in hepatocytes. Momordica foetida and Momordica balsamina extracts were active in myocytes but only the latter stimulated glucose utilisation in hepatocytes. Brachylaena discolor gave the best overall results, with all plant parts giving high activity scores and negligible toxicity. In vitro toxicity results for Catharanthus roseus, Vinca major, Momordica balsamina and some Sclerocarya birrea extracts raise concern for chronic use. CONCLUSION: This screening system increases the likelihood of identifying drug candidates using in vitro antidiabetic screening of crude plant extracts, whilst the scoring system aids data interpretation. ETHNOPHARMACOLOGICAL RELEVANCE: The multitude of metabolic steps affected by Type II diabetes offer many drug targets but they complicate in vitro screening to validate traditional uses or find new drug leads from plants.

Pharmacokinetics and tolerability of artesunate and amodiaquine alone and in combination in healthy volunteers.

OBJECTIVES: The WHO recommends artemisinin-based combination therapies for treatment of uncomplicated falciparum malaria. At least 15 African countries have adopted artesunate plus amodiaquine as treatment policy. As no pharmacokinetic data on this combination have been published to date, we investigated its pharmacokinetic interactions and tolerability in healthy volunteers in Africa. METHODS: In a randomized, three-phase, cross-over study, amodiaquine (10 mg/kg) and artesunate (4 mg/kg) were given as single oral doses to 15 healthy volunteers. Artesunate was given to all volunteers on day 0. On day 7 they received either amodiaquine or amodiaquine plus artesunate and the alternative regimen on day 28. The pharmacokinetics of artesunate and amodiaquine and their main active metabolites dihydroartemisinin and desethylamodiaquine were compared following monotherapy and combination therapy using analysis of variance. RESULTS: Thirteen volunteers completed the study, and pharmacokinetic parameters could be determined for twelve volunteers. When given in combination, the mean AUC was lower for dihydroartemisinin [ratio 67% (95% CI 51-88%); P = 0.008] and desethylamodiaquine [ratio 65% (95% CI 46-90%); P = 0.015] when compared with monotherapy. Adverse events of concern occurred in four volunteers (27%): grade 3 transaminitis (n = 1), neutropaenia (n = 2), and hypersensitivity (n = 1). CONCLUSION: The total drug exposure to both drugs was reduced significantly when they were given in combination. The clinical significance of these interactions is unclear and must be studied in malaria patients. The frequency and nature of adverse events among the healthy volunteers were of concern, and suggest laboratory monitoring would be needed in malaria patients treated with artesunate plus amodiaquine.

Rifampin levels, interferon-gamma release and outcome in complicated pulmonary tuberculosis.

Factors that relate to medium-term outcome in patients with pulmonary tuberculosis (PTB) who have completed the 2-month intensive phase of treatment are incompletely understood. The relationship between in vitro production of interferon-gamma (IFN-gamma), interleukins (ILs)-5 and -10 and drug levels determined after 2 months of drug therapy, to outcome at 24 months was studied prospectively. Cytokine concentrations were determined from culture supernatants after stimulation of whole blood with purified protein derivative (PPD) of Mycobacterium tuberculosis. Plasma concentrations of rifampin, isoniazid, pyrazinamide and ethambutol were determined by high-performance liquid chromatography. The treatment failure and relapse free survival probability was 0.54 (95% CI: 0.40-0.67) at 24 months. In multivariate analysis of parameters at 2 months the strongest positive associations with disease free survival were IFN-gamma response to PPD (p=0.002) and serum creatinine (p=0.001). Drug concentrations were not associated with outcome although rifampin exposure correlated with IFN-gamma response to PPD (p=0.0132). These data suggest that the ability to mount a recall immune response to M. tuberculosis may influence treatment outcome. The data support the idea to identify persons at risk of a poor treatment outcome by monitoring of the in vitro response to tuberculosis antigens.

Combination of tunicamycin with anticancer drugs synergistically enhances their toxicity in multidrug-resistant human ovarian cystadenocarcinoma cells.

BACKGROUND: The pharmacologic modulatory effects of the antibiotic, tunicamycin (TM), on multidrug-resistant human UWOV2 ovarian cancer cells are reported. The UWOV2 cell line was derived from a cystadenocarcinoma in a patient refractory to combination chemotherapy with actinomycin D, vincristine (VCR), cis-diaminedichloroplatinum (II) (CDDP) and doxorubicin (DXR). In an attempt to explain drug resistance in this cell line, we examined the effects of TM on their sensitivity to various anticancer drugs, the uptake, efflux and retention of [3H]VCR, and their ability to bind [14C]DXR and [3H]azidopine (AZD), a photoaffinity label of the multidrug transporter, P-glycoprotein (Pgp). RESULTS: TM effectively decreased the EC50 for DXR, EXR, VCR and CDDP, thus enhancing their cytotoxicity. The antibiotic also prolonged the intracellular retention time of [3H]VCR and increased the binding of both [14C]DXR and [3H]AZD to the cells. CONCLUSION: It is concluded that the pharmacomodulatory effects of TM in these cells are mediated by global inhibition of protein and glycoprotein synthesis and synergistic interaction with antineoplastic drugs. The ability of TM to enhance the sensitivity of drug resistant tumour cells may have impact on the design and optimization of novel resistance modifiers to improve the efficacy of combination treatment of intractable neoplasms.

Population pharmacokinetics of artesunate and dihydroartemisinin following intra-rectal dosing of artesunate in malaria patients.

BACKGROUND: Intra-rectal artesunate has been developed as a potentially life-saving treatment of severe malaria in rural village settings where administration of parenteral antimalarial drugs is not possible. We studied the population pharmacokinetics of intra-rectal artesunate and the relationship with parasitological responses in patients with moderately severe falciparum malaria. METHODS AND FINDINGS: Adults and children in Africa and Southeast Asia with moderately severe malaria were recruited in two Phase II studies (12 adults from Southeast Asia and 11 children from Africa) with intensive sampling protocols, and three Phase III studies (44 children from Southeast Asia, and 86 children and 26 adults from Africa) with sparse sampling. All patients received 10 mg/kg artesunate as a single intra-rectal dose of suppositories. Venous blood samples were taken during a period of 24 h following dosing. Plasma artesunate and dihydroartemisinin (DHA, the main biologically active metabolite) concentrations were measured by high-performance liquid chromatography with electrochemical detection. The pharmacokinetic properties of DHA were determined using nonlinear mixed-effects modelling. Artesunate is rapidly hydrolysed in vivo to DHA, and this contributes the majority of antimalarial activity. For DHA, a one-compartment model assuming complete conversion from artesunate and first-order appearance and elimination kinetics gave the best fit to the data. The mean population estimate of apparent clearance (CL/F) was 2.64 (l/kg/h) with 66% inter-individual variability. The apparent volume of distribution (V/F) was 2.75 (l/kg) with 96% inter-individual variability. The estimated DHA population mean elimination half-life was 43 min. Gender was associated with increased mean CL/F by 1.14 (95% CI: 0.36-1.92) (l/kg/h) for a male compared with a female, and weight was positively associated with V/F. Larger V/Fs were observed for the patients requiring early rescue treatment compared with the remainder, independent of any confounders. No associations between the parasitological responses and the posterior individual estimates of V/F, CL/F, and AUC0-6h were observed. CONCLUSIONS: The pharmacokinetic properties of DHA were affected only by gender and body weight. Patients with the lowest area under the DHA concentration curve did not have slower parasite clearance, suggesting that rectal artesunate is well absorbed in most patients with moderately severe malaria. However, a number of modelling assumptions were required due to the large intra- and inter-individual variability of the DHA concentrations.

Determinants of rifampin, isoniazid, pyrazinamide, and ethambutol pharmacokinetics in a cohort of tuberculosis patients.

Evaluation of sources of pharmacokinetic variation can facilitate optimization of tuberculosis treatment regimens by identification of avoidable sources of variation and of risk factors for low or high drug concentrations in patients. Our objective was to describe the pharmacokinetics of rifampin, isoniazid, pyrazinamide, and ethambutol in a cohort of tuberculosis patients established on first-line treatment regimens and to evaluate the determinants of pharmacokinetic variation. Plasma concentration-time profiles were determined for each of the drugs in 142 patients with drug-sensitive pulmonary tuberculosis after 2 months of daily treatment in hospital. Pharmacokinetic measures were described by noncompartmental analysis. Multiple linear regression was used to evaluate the patient and the treatment factors associated with variation of the area under the concentration-time curve from 0 to 8 h. Several factors independently associated with variations in antituberculosis drug concentrations were identified: human immunodeficiency virus infection was associated with 39% and 27% reductions for rifampin and ethambutol, respectively; formulation factors were determinants of rifampin and isoniazid bioavailability; female patients had increased rifampin and isoniazid concentrations but reduced ethambutol concentrations; older patients had higher levels of isoniazid and ethambutol; patients with a history of previous antituberculosis treatment had lower ethambutol concentrations; and the dose per kilogram of body weight was associated with the concentrations of all four agents. Further studies are required to assess the implications of variations in antituberculosis drug concentrations for efficacy and safety before decisions are made to change the dosing strategy in patients at risk.

A global perspective on vaccine safety and public health: the Global Advisory Committee on Vaccine Safety.

Established in 1999, the Global Advisory Committee on Vaccine Safety advises the World Health Organization (WHO) on vaccine-related safety issues and enables WHO to respond promptly, efficiently, and with scientific rigor to issues of vaccine safety with potential global importance. The committee also assesses the implications of vaccine safety for practice worldwide and for WHO policies. We describe the principles on which the committee was established, its modus operandi, and the scope of the work undertaken, both present and future. We highlight its recent recommendations on major issues, including the purported link between the measles-mumps-rubella vaccine and autism and the safety of the mumps, influenza, yellow fever, BCG, and smallpox vaccines as well as that of thiomersal-containing vaccines.

In vitro antiplasmodial activity of medicinal plants native to or naturalised in South Africa.

The increasing prevalence and distribution of malaria has been attributed to a number of factors, one of them being the emergence and spread of drug resistant parasites. Efforts are now being directed towards the discovery and development of new chemically diverse antimalarial agents. The present study reports on the in vitro antiplasmodial activity of 134 plant taxa native to or naturalised in South Africa, representing 54 families, which were selected semi-quantitatively using weighted criteria. The plant extracts were tested for in vitro activity against a Plasmodium falciparum strain D10 using the parasite lactate dehydrogenase (pLDH) assay. Of the 134 species assayed, 49% showed promising antiplasmodial activity (IC(50)< or = 10 microg/ml), while 17% were found to be highly active (IC(50)< or = 5 microg/ml). Several plant species and genera were shown for the first time to possess in vitro antiplasmodial activity. These results support a rational rather than random approach to the selection of antiplasmodial screening candidates, and identify a number of promising taxa for further investigation as plant-based antimalarial agents.

Oral artesunate prevents Plasmodium berghei Anka infection in mice.

Artesunate, a semi-synthetic derivative of a naturally occurring anti-malarial artemisinin was compared with chloroquine in C57BL/6 mice infected with Plasmodium berghei Anka (PbA). A 7-day oral administration of artesunate prevented parasitaemia at 10 mg/kg/day. However, recrudescence of parasitaemia and cerebral malaria occurred upon cessation of treatment followed by death within 28 days. However, a 14-day course of artesunate (100 mg/kg/day) prevented completely the development of parasitaemia and cerebral malaria with a survival of more than 60-days as did 10 mg/kg/day chloroquine. These data demonstrate that oral artesunate inhibits PbA and prevents cerebral malaria, but needs to be administered at high dose and for prolonged time to eradicate PbA infection in mice.

The role of professionals in the South African chemical and biological warfare programme.

This paper provides a short account of the South African Defence Force's chemical and biological warfare programme during apartheid, specifically during the period 1980 to 1994. It examines the circumstances of recruitment of the scientists and physicians and their retention in the programme; details the 'scientific efforts' of the programme and its aberrations; and explores ethical issues in relation to the involvement of scientists in the programme.

The Effects of Cyclophosphamide on Alkaline Phosphatase Activity and on in vitro Post-Implantation Murine Blastocyst Development† : (preimplantation embryo/cyclophosphamide/teratogenicity/alkaline phosphatase/differentiation).

Cyclophosphamide, administered in doses of 20 and 40 mg/kg body weight to pregnant inbred CBA mice on the third day of gestation (60 h after the estimated time of ovulation) reduced the activity of non-specific alkaline phosphatase (E.C. 3.1.3.1.) in 84 h-old blastocysts in a dose-related fashion compared with controls (p < 0.02 in each case; comparison of groups by Kruskal Wallis analysis of variance). This effect was not demonstrated with 4 mg/kg body weight (p < 0.1). Forty mg/kg body weight, but not the lower doses of cyclophosphamide, significantly retarded the hatching of embryos from the zona pellucida, attachment to the culture dish, and the formation of trophoblast outgrowths when the blastocysts were subsequently cultured in vitro for 120 h. The growth of an expanded inner cell mass was impaired in the 20 and 40 mg/kg groups. The differentiation of the inner cell mass into endoderm and ectoderm was significantly affected in the 4 mg, 20 mg and 40 mg/kg groups (p < 0.05, p < 0.01 and p < 0.001 respectively). The possible relationships of these various findings are discussed in the text. These results suggest that alkaline phosphatase may be a useful indicator of impaired growth and differentiation of the inner cell mass after exposure of preimplantation embryos to teratogens.