Insecticide resistance in Culex quinquefasciatus from Zanzibar: implications for vector control programmes

Abstract

Background: Zanzibar has a long history of lymphatic filariasis (LF) caused by the filarial parasite Wuchereria bancrofti, and transmitted by the mosquito Culex quinquefasciatus Say. The LF Programme in Zanzibar has successfully implemented mass drug administration (MDA) to interrupt transmission, and is now in the elimination phase. Monitoring infections in mosquitoes, and assessing the potential role of interventions such as vector control, is important in case the disease re-emerges as a public health problem. Here, we examine Culex mosquito species from the two main islands to detect W. bancrofti infection and to determine levels of susceptibility to the insecticides used for vector control.

Methods: Culex mosquitoes collected during routine catches in Vitongoji, Pemba Island, and Makadara, Unguja Island were tested for W. bancrofti infection using PCR. Insecticide bioassays on Culex mosquitoes were performed to determine susceptibility to permethrin, deltamethrin, lambda-cyhalothrin, DDT and bendiocarb. Additional synergism assays with piperonyl butoxide (PBO) were used for lambda-cyhalothrin. Pyrosequencing was used to determine the kdr genotype and sequencing of the mitochondrial cytochrome oxidase I (mtCOI) subunit performed to identify ambiguous Culex species.

Results: None of the wild-caught Culex mosquitoes analysed were found to be positive for W. bancrofti. High frequencies of resistance to all insecticides were found in Wete, Pemba Island, whereas Culex from the nearby site of Tibirinzi (Pemba) and in Kilimani, Unguja Island remained relatively susceptible. Species identification confirmed that mosquitoes from Wete were Culex quinquefasciatus. The majority of the Culex collected from Tibirinzi and all from Kilimani could not be identified to species by molecular assays. Two alternative kdr alleles, both resulting in a L1014F substitution were detected in Cx. quinquefasciatus from Wete with no homozygote susceptible detected. Metabolic resistance to pyrethroids was also implicated by PBO synergism assays.

Conclusions: Results from the xenomonitoring are encouraging for the LF programme in Zanzibar. However, the high levels of pyrethroid resistance found in the principle LF vector in Pemba Island will need to be taken into consideration if vector control is to be implemented as part of the elimination programme.

Figure 1

Study sites on Unguja and Pemba Islands, Zanzibar.

Figure 2

Melt-curve profile for determining the presence or absence of W. bancrofti in Culex mosquitoes. The red asterix adjacent to specific peaks in the melt curve indicates W. bancrofti infected Culex (T_m = 74.3-74.8). The yellow asterix adjacent to non-specific peaks in the melt curve profile indicates Culex mosquitoes free from W. bancrofti infection.

Figure 3

Percentage mortality of Culex mosquitoes from Zanzibar following exposure to WHO insecticide treated papers. Percentage mortality and 95% confidence intervals from a) Wete (Pemba) b) Wete (Pemba) with pre-exposure to PBO (4%) c) Tibirinzi (Pemba) d) Kilimani (Unguja).

Figure 4

The frequency of Cx. quinquefasciatus from the study sites on Zanzibar. Cx. quinquefasciatus specific primers were used to discriminate this species from samples from Wete (N = 19), Tibirinzi (N = 24) and Kilimani (N = 20). Samples which failed to amplify were classed as unknown 'Culex spp'.