Sex-specific induction of CYP6 cytochrome P450 genes in cadmium and lead tolerant Anopheles gambiae.

BACKGROUND: Anopheles gambiae, one of the main Afro-tropical mosquito vector of malaria, has adapted to heavy metals in its natural habitat, and developed resistance to most conventional insecticides. Investigations were conducted to establish an association between tolerance to cadmium or lead-heavy metals, and expression of specific genes for cytochrome p450 enzymes associated with pyrethroid resistance in the mosquito. METHODS: Juvenile aquatic stages of the mosquito were selected for tolerance to cadmiun or lead through chronic exposure of the stages to maximum acceptable toxicant concentrations (MATCs) of the metals. Using real-time quantitative polymerase chain reaction (qPCR), three replicates each of male or female cadmium or lead-tolerant individuals and relevant controls were separately screened for expression of CYP6M2, CYP6P3 and CYP6Z1 genes. The variance in expression levels of the genes amongst the treatments was compared by ANOVA statistical tool. RESULTS: Expressions of all the genes were significantly lower (P <0.05) in females than in males. Within gender, there 1.3 - 2.3 or 3.1-4.2-fold reduction in expression of the genes in cadmium or lead selected than respective control populations. Expression of all the classes of gene was elevated in cadmium selected female populations relative to their respective controls. CONCLUSION: These findings suggest that tolerance to cadmium or lead in the mosquito can influence response in cytochrome p450 genes associated with metabolism of pyrethroids in the mosquito in a sex-specific manner. This can, in turn, affect sensitivity of the mosquito to pyrethroids and other xenobiotics associated with these genes, with potential implications in mosquito vector control operations.

Changes in genotypes of Plasmodium falciparum human malaria parasite following withdrawal of chloroquine in Tiwi, Kenya.

Chloroquine (CQ) drug was withdrawn in 1998 as a first-line treatment of uncomplicated malaria in Kenya. This was in response to resistance to the drug in Plasmodium falciparum malaria parasite. Investigations were conducted to determine prevalence of CQ resistance genotypes in the parasites in Tiwi, a malaria endemic town in Kenya, before and about a decade after the withdrawal of the drug. Blood samples were collected and spotted on filter papers in 1999 and 2008 from 75 and 77 out-patients respectively with uncomplicated malaria. The sampling was conducted using finger pricking technique. DNA was extracted from individual spots in the papers and screened for the presence of P. falciparum chloroquine resistance transporter (Pfcrt) and multi drug resistance (Pfmdr-1) markers using nested PCR. Nature of mutations (haplotypes) in the Pfcrt and Pfmdr-1 markers in the samples were confirmed using dot blot hybridization technique. Changes in pattern of CQ resistance in the parasite samples in 1999 and 2008 were assessed by Chi Square test. There was a significant (P<0.05) reduction in CQ resistant genotypes of the parasite between 1999 and 2008. Pfmdr and Pfcrt CQ resistant genotypes in 2008 reduced to 54.10 and 63.64% respectively, from 75.39 and 88.0% respectively in 1999. This reduction was accompanied by emergence of Pfcrt specific CQ sensitive (IEK) and intermediate/partially CQ resistant (MET) haplotypes. Results suggest significant reversal of the phenotype of the parasite from chloroquine resistant to wild/sensitive type. The novel haplotypes indicates transitional phase of the parasite to the wild type. Current prevalence of chloroquine resistant genotype is definitely above the threshold for efficacious re-introduction of chloroquine for treatment of malaria in Tiwi.