Molecular analysis of DHFR and DHPS genes in P. falciparum clinical isolates from the Haut--Ogooué region in Gabon.

The main objective of this work was to determine the prevalence of mutations in genes coding for the dihydropteroate synthase (DHPS) and the dihydrofolate reductase (DHFR) enzymes which are implicated in resistance of P. falciparum to antifolate (pyrimethamine-sulfadoxine (P/S)). In this study, 117 human blood samples were collected at Franceville located in the region of Haut-Ogooué (South-eastern Gabon). In this area, a relatively low level of sensitivity of Plasmodium falciparum to P/S has been reported with 18.2% of RII and 12.1% of RI resistance. A nested polymerase chain reaction was used to amplify a fragment of the DHFR gene containing codon 108, where a point mutation causing a Serine (wild type) to Asparagine or to a Threonine (resistant types) change occurs in pyrimethamine resistant parasites. Eleven DHFR fragments were sequenced and mutations occurring at codons 51, 59 and 108 were analysed. The DHPS gene was amplified by polymerase chain reaction (PCR) and sequenced directly or after cloning. Variant amino acid residues 436, 437, 540, 581, 613 associated with sulfadoxine resistance were analysed. The analysis of codon 108 of the DHFR gene was undertaken for 81 isolates. More than one DHFR P. falciparum genotype was present in 64% of the samples. We showed that 47% of 141 DHFR gene PCR products had Serine (wild phenotype), and 52% had Asparagine. We found one isolate with the Thr-108 confirmed by sequencing of the PCR product. Triple, double and single DHFR mutant at positions 51, 59 and 108 were found. Only codons 436 and 437 of the 38 analysed sequences of the DHPS gene revealed point mutations. These results have been compared with those reported from different sites in Africa, Asia or South-America.

[Seroprevalence of rickettsial infections of the Boutonneuse fever group or its apparent group in Bangui (Central African Republic)]. / Séroprévalence des infections à rickettsies du groupe des fièvres boutonneuses ou apparentées à Bangui (République centrafricaine).

115 sera from African individuals, including 66 HIV-1-infected individuals and 49 HIV-seronegative age-matched healthy controls, living in Bangui, Central African Republic, were screened for Rickettsia conorii antibody by indirect immunofluorescent assay; 53 (46%) sera had significant IgG to R. conorii titer, and 5 (4.3%) had significant IgM to R. conorii titer, without significant difference between HIV-positive and negative individuals. These findings indicate a great exposure to spotted fever group rickettsial infections in Bangui.

[Evaluation of a simple and rapid method of Plasmodium falciparum DNA extraction using thick blood smears from Gabonese patients]. / Evaluation d'une méthode simple et rapide d'extraction de l'ADN de Plasmodium falciparum à partir de gouttes épaisses de patients gabonais.

In field-based studies, sometimes it is difficult to collect and store samples. We have evaluated a method of malaria parasite deoxyribonucleic (DNA) extraction from non-stained thick dried blood smears collected from 108 Gabonese patients. This method of DNA isolation was compared to those using phenol/chloroform. Patients parasitemia ranged from 0 to 240,000 parasites/microliter of blood. Both methods of DNA preparation gave similar results. Of the 108 slides, 57% were Plasmodium falciparum positive after PCR analysis of the MSA-2 gene and 34% were positive by microscopical examination. Thirty-six and seventy-two blood smears from patients were also tested after one and four weeks' storage respectively, at room temperature, and the parasite DNA was successfully extracted. We conclude that this simple method of collection and rapid procedure of parasite DNA isolation are adequate and convenient in the field when a large number of samples are required and in the case of repetitive samplings of patients.

Allelic family-specific humoral responses to merozoite surface protein 2 (MSP2) in Gabonese residents with Plasmodium falciparum infections.

Merozoite surface protein 2 (MSP2) expressed by Plasmodium falciparum asexual blood stages has been identified as a promising vaccine candidate. In order to explore allelic family-specific humoral responses which may be responsible for parasite neutralization during natural infections, isolates from individuals with either asymptomatic infections or uncomplicated malaria and residing in a Central African area where Plasmodium transmission is high and perennial, were analysed using MSP2 as polymorphic marker. The family-specific antibody responses were assessed by ELISA using MSP2 synthetic peptides. We observed an age-dependence of P. falciparum infection complexity. The decrease of infection complexity around 15 years of age was observed simultaneously with an increase in the mean number of MSP2 variants recognized. No significant difference in the P. falciparum genetic diversity and infection complexity was found in isolates from asymptomatic subjects and patients with uncomplicated malaria. The longitudinal follow-up showed a rapid development of immune responses to various regions of MSP2 variants within one week. Comparing humoral responses obtained with the other major antigen on the merozoite surface, MSP1, our findings suggest that different pathways of responsiveness are involved in antibody production to merozoite surface antigens.

Coxiella burnetii infection among subjects infected with HIV type 1 in the Central African Republic.

Sixty-six sera from HIV-1-seropositive adult African subjects and 49 sera from HIV-seronegative age and sex matched healthy African controls living in Bangui, Central African Republic, were screened for Coxiella burnetii antibody by an indirect immunofluorescent antibody test. 16.7% of HIV-infected patients and 16.3% of the HIV-negative controls had positive IgG titres, with no significant difference between the two groups. Two of the seven HIV-infected patients seropositive for Coxiella burnetii for whom clinical data was available had a medical history compatible with symptomatic Q fever. These findings indicate that there is a high degree of exposure to Coxiella burnetii infection in Bangui. In individuals co-infected with HIV and Coxiella burnetii, cellular immunosuppression could favour symptomatic Q fever. Physicians should be aware of the possibility of symptomatic Coxiella burnetii infection among HIV-infected people, particularly in endemic regions for both infections such as in sub-saharan Africa.