Absence of hybridization with the wild-type and mutant rpoB probes in the Genotype MTBDRplus assay detects 'disputed' rifampicin mutations.

OBJECTIVE: Mycobacterium tuberculosis isolates that fail to hybridize to at least one rpoB wild-type or any mutation probe on the Genotype MTBDRplus strip are assumed to be rifampicin-resistant. However, the precise mutation(s) are unknown. We sought to identify the mutations in isolates with such hybridization patterns and determine if the mutations are associated with resistance to rifampicin. METHODS: In this study, 275 M. tuberculosis isolates were screened with the Genotype MTBDRplus assay to identify isolates with the hybridization pattern. These isolates were sequenced and their minimum inhibitory concentrations (MIC) determined using the Bactec MGIT 960 system. RESULTS: Among the 275 isolates tested, 15 (6%) isolates with the hybridization pattern were identified. Sequencing showed that failure to hybridize to rpoB wild-type probes resulted from the presence of 'disputed' rifampicin mutations, which are mutations not always associated with a rifampicin-resistant phenotype. All, except 3/15, isolates had a rifampicin-resistant phenotype (MIC > 1 µg/mL). One of the three isolates with a rifampicin-susceptible phenotype had the same mutation at position 526 (His526Leu) as another isolate that had a rifampicin-resistant phenotype. CONCLUSION: The recommendation of the Genotype MTBDRplus assay to assume rifampicin resistance based solely on failure to hybridize to rpoB wild-type probe allows the identification of important RIF-resistant isolates. About 20% (3/15) of such isolates could be missed by relying only on the standard MGIT 960 DST assay for drug susceptibility testing.

Mentoring for early-career women in health research: the HIGHER Women Consortium approach.

Attracting and retaining women in health research is crucial as it will maximize creativity and innovation as well as increase gender competency and expertise in the field. To help address the gender gap in the research for health field in Cameroon, some women research scientists formed the Higher Institute for Growth in HEalth Research for Women (HIGHER Women) consortium to support and encourage the growth of women research scientists through a training institute with a Mentor-Protégé Program (MPP). The consortium set up a MPP aiming at providing professional guidance to facilitate protégés' growth and emergence in health research. The consortium has conducted two workshops aiming at increasing the early-career women's skills needed to launch their career and focusing on proposal writing with the aim of producing a fundable project. Since 2015, the consortium has brought together approximately 100 women comprising of 80 protégés. The most significant outcome is in the protégés' feedback from their annual evaluations. The protégés are now more likely to submit abstracts and attend international conferences. Some grants have been obtained as a result of the working relationship with mentors. The HIGHER women consortium works to develop a pipeline of women leaders in health research by fostering growth and leadership culture through their MPP.

Extensive multiple test centre evaluation of the VecTest malaria antigen panel assay.

To determine which species and populations of Anopheles transmit malaria in any given situation, immunological assays for malaria sporozoite antigen can replace traditional microscopical examination of freshly dissected Anopheles. We developed a wicking assay for use with mosquitoes that identifies the presence or absence of specific peptide epitopes of circumsporozoite (CS) protein of Plasmodium falciparum and two strains of Plasmodium vivax (variants 210 and 247). The resulting assay (VecTest Malaria) is a rapid, one-step procedure using a 'dipstick' test strip capable of detecting and distinguishing between P. falciparum and P. vivax infections in mosquitoes. The objective of the present study was to test the efficacy, sensitivity, stability and field-user acceptability of this wicking dipstick assay. In collaboration with 16 test centres world-wide, we evaluated more than 40 000 units of this assay, comparing it to the standard CS ELISA. The 'VecTest Malaria' was found to show 92% sensitivity and 98.1% specificity, with 97.8% accuracy overall. In accelerated storage tests, the dipsticks remained stable for > 15 weeks in dry conditions up to 45 degrees C and in humid conditions up to 37 degrees C. Evidently, this quick and easy dipstick test performs at an acceptable level of reliability and offers practical advantages for field workers needing to make rapid surveys of malaria vectors.

The state of immunology in Africa: HIV/AIDS and malaria.

HIV and malaria are two major infections that are responsible for the greatest burden of diseases, morbidity and mortality in the African population. Successful research has been undertaken in Africa into novel means of monitoring HIV disease progression and in identifying vaccine candidates. The role of IgG isotypes in malaria has been investigated, as have parasite adhesion molecules important for pathogenesis. It is hoped that vaccines for malaria will soon prove successful. However, many problems still face immunology research in Africa.

The epidemiology of Plasmodium falciparum malaria in two Cameroonian villages: Simbok and Etoa.

In support of ongoing immunologic studies on immunity to Plasmodium falciparum, demographic, entomologic, parasitologic, and clinical studies were conducted in two Cameroonian villages located 3 km apart. Simbok (population = 907) has pools of water present year round that provide breeding sites for Anopheles gambiae, whereas Etoa (population = 485) has swampy areas that dry up annually in which A. funestus breed. Results showed that individuals in Simbok receive an estimated 1.9 and 1.2 infectious bites per night in the wet and dry season, respectively, whereas individuals in Etoa receive 2.4 and 0.4 infectious bites per night, respectively. Although transmission patterns differ, the rate of acquisition of immunity to malaria appears to be similar in both villages. A prevalence of 50-75% was found in children < 10 years old, variable levels in children 11-15 years old, and 31% in adults. Thus, as reported in other parts of Africa, individuals exposed to continuous transmission of P. falciparum slowly acquired significant, but not complete, immunity.

Correlation between serum levels of antibodies to the 96-kD antigen of Plasmodium falciparum and protective immunity in Cameroon: a longitudinal study.

A longitudinal study was conducted in the Yaounde area of Cameroon that involved 211 individuals in June 1990, and 70 individuals for the follow-up study in December 1990. Sera from these subjects were tested against the recombinant 96-thermoresistant antigen of Plasmodium falciparum and the kinetics of antibody production to this protein show that titers tend to increase with age and are also related to antigen exposure. The increase in antibody titers with age correlates positively with the ability of the individual to prevent development of a high parasitemia. Adults who maintained stable high titers generally did not experience clinical attacks during the study period. The data suggest that antibodies against the 96-kD antigen participate in conferring some immunity to falciparum malaria.

Identification of anti-Plasmodium falciparum antibodies in human breast milk.

Malarial infections are rarely observed in neonates. It has been postulated that some immunity may be passively transferred during nursing, but anti-malarial antibodies (Abs) have not been detected in human milk. In this study, milk samples, collected 2-14 days after parturition from women at the Central Maternity Hospital, Yaounde, were evaluated for total IgG and IgA antibody levels by radial diffusion, protein composition by SDS-PAGE, anti-malarial antibodies using an isotype-specific immunofluorescence assay, and the ability to immunoprecipitate Plasmodium falciparum antigens metabolically labelled with 35S-methionine. Results showed that anti-P. falciparum antibodies were present in breast milk, and that paired milk and serum samples from individual women contained Abs that recognized similar malarial antigens.

Comparative studies of clinical groups of patients in an onchocerciasis endemic area for evidence of immune-mediated protection.

Four clinical groups of persons from an area endemic for onchocerciasis were compared using certain immunological parameters. The groups were: generalised onchocerciasis, patients with restricted distribution of onchocercal skin lesions, microfilaredermia patients with no clinical manifestations, and a group which clinically, had successfully resisted the infection. Specific serum antibodies to O. volvulus antigens were found in all groups. The IgG specific antibodies were highest in patients with generalised onchocerciasis and lowest in the group who had apparently contained the infection. The sera of persons from the latter group mediated leukocyte adherence to and immobilised microfilariae of O. volvulus. Using the Western blot technique, there were no onchocercal protein antigens that reacted exclusively with sera from the "protected group". However, the staining of the reaction bands was most intense when sera from this patient group reacted with low molecular weight specific onchocercal antigens (M. W. 10-57 KD).

Immunity to Onchocerca volvulus. Serum mediated leucocyte adherence to infective larvae in vitro.

Leucocyte adherence to infective larvae of Onchocerca volvulus in the presence of serum was evaluated using sera from four clinically distinct groups of patients with Onchocerciasis from an area hyperendemic for the disease. Significant cellular adherence to infective larvae occurred for the most part in the presence of sera obtained from subjects with either no microfilaridemia and few or no palpable nodules. These patients had, as well, the highest serum titres of specific anti-O. volvulus IgG antibodies. In contrast, sera from subjects with many palpable nodules and heavy skin infiltration with microfilariae (generalised disease) did not mediate significant adherence of leucocytes to infective larvae targets. Further, this group had the lowest serum levels of specific anti-O. volvulus antibodies of the IgG isotype. The findings are in keeping with the hypothesis that specific protective immunity may occur in O. volvulus infections.