The Comparative Effectiveness of Adult Mosquito Sampling Methods Dealing With Odor-baited Resting Box Traps in a Malaria-endemic Area in Southern Iran

BackgroundIran is under threat of a potential outbreak of mosquito-borne diseases, such as malaria, Dengue fever, Chikungunya, and Zika. The study aimed to determine the efficiency and sustainability of some adult mosquito sampling methods for designing effective entomological surveillance systems in a malaria endemic area.MethodsDifferent rates of tap water, sugar, and yeast (Saccharomyces cerevisiae) were mixed to provide CO2. Anopheles stephensi and Culex pipiens Bandar Abbas strains were reared in the insectary and used for tests. On a lab scale, CO2 orientation experiments were performed using a Y-tube olfactometer on the insectary mosquito strains. In the field trial, human landing catches (HLC), artificial pit shelter (APS), CO2-baited trap (CO2-BT), human and cow odor baited resting boxes (HOBT, COBT), cow urine baited trap (CUBT), and colored un-baited box (UB) were studied in Bandar Abbas, Hormozgan Province, south of Iran. Mean densities of An. stephensi and Cx. pipiens insectary strains, which oriented to CO2 as flowrate of 170, and 300 mL/minute was significantly higher compared to the control group (P < 0.05). The black un-baited inbox resting traps significantly more collected wild mosquito species compared to others colored UB (P ˂ 0.05). ResultsA total of 2722 collected mosquitoes comprises Culex pipiens (48.56%), Anopheles fluviatilis s.l.(14.21%), An. stephensi (11.68%), Cx. theileri (9.95%), Aedes caspius (7.01%), An. dthali (6.79%) An. culicifacies s.l. , An. pulcherrimus, An. sergentii,An. superpictus s.l., Cx. sitiens and, Ae.caspius less than 1%. Anopheles stephensi and, An.sergenti were most collected in CUBT. Anopheles fluviatilis s.l. and Ae. caspius were most found in HLC. Anopheles dthali, Cx. pipiens and, Cx. theileri were most abundant in APS. ConclusionsBlack CUBT and APS methods can be suggested as a perfect sampling strategy for malaria vectors surveillance. APS, and HLC methods were found useful to entomological surveillance systems for arboviral and filarial vector-borne diseases. Further modified sampling methods should be devoted to identify more effective sampling methods.

The Comparative Effectiveness of Adult Mosquito Sampling Methods Dealing With Odor-Baited Resting Box Traps in A Malaria-Endemic Area in Southern Iran

Background Iran is under threat of a potential outbreak of mosquito-borne diseases, such as malaria, Dengue fever, Chikungunya, and Zika. The study aimed to determine the efficiency and sustainability of some adult mosquito sampling methods for designing effective entomological surveillance systems in a malaria endemic area. Methods Different rates of tap water, sugar, and yeast (Saccharomyces cerevisiae) were mixed to provide CO2. Anopheles stephensi and Culex pipiens Bandar Abbas strains were reared in the insectary and used for tests. On a lab scale, CO2 orientation experiments were performed using a Y-tube olfactometer on the insectary mosquito strains. In the field trial, human landing catches (HLC), artificial pit shelter (APS), CO2-baited trap (CO2-BT), human and cow odor baited resting boxes (HOBT, COBT), cow urine baited trap (CUBT), and colored un-baited box (UB) were studied in Bandar Abbas, Hormozgan Province, south of Iran. Mean densities of An. stephensi and Cx. pipiens insectary strains, which oriented to CO2 as flowrate of 170, and 300 mL/minute was significantly higher compared to the control group (P < 0.05). The black un-baited inbox resting traps significantly more collected wild mosquito species compared to others colored UB (P ˂ 0.05). Results A total of 2722 collected mosquitoes comprises Culex pipiens (48.56%), Anopheles fluviatilis s.l.(14.21%), An. stephensi (11.68%), Cx. theileri (9.95%), Aedes caspius (7.01%), An. dthali (6.79%) An. culicifacies s.l., An. pulcherrimus, An. sergentii,An. superpictus s.l., Cx. sitiens and, Ae.caspius less than 1%. Anopheles stephensi and, An.sergenti were most collected in CUBT. Anopheles fluviatilis s.l. and Ae. caspius were most found in HLC. Anopheles dthali, Cx. pipiens and, Cx. theileri were most abundant in APS. Conclusions Black CUBT and APS methods can be suggested as a perfect sampling strategy for malaria vectors surveillance. APS, and HLC methods were found useful to entomological surveillance systems for arboviral and filarial vector-borne diseases. Further modified sampling methods should be devoted to identify more effective sampling methods.

Molecular characterization of Anopheles fluviatilis species complex in the Islamic Republic of Iran

A species-specific polymerase chain reaction [PCR] assay was used to identify the species composition of the Anopheles fluviatilis complex in the Islamic Republic of Iran. All the amplified DNA samples from specimens collected from different areas yielded a fragment of 450 bp size, a PCR product corresponding to that of the species denoted as Y. The sequence data from 21 ITS2 [second internal transcribed spacer] regions were compared with those publicly available in the GenBank database and confirmed that the specimens were 100% identical to species Y of India. Species Y is presumably the same as species T that has no role in transmission of malaria in India, whereas An. fluviatilis is known as a secondary vector of malaria in the Islamic Republic of Iran

Are members of the Anopheles fluviatilis complex conspecific?

Anopheles fluviatilis sensu lato, a primary malaria vector in India, was identified to be comprised of four cryptic species, provisionally designated as species S, T, U and V. However, Kumar et al. (Mol Ecol Resour, 2013;13:354-61) considered all of the then known three members of this species complex (S, T and U) conspecific. The specific status of species S and T was refuted based on the lack of sufficient barcode gap in mitochondrial-CO1 and the perceived presence of heterozygotes in populations as detected through one of the two species-specific PCR assays employed for the cryptic species identification. The existence of species U was refuted claiming that earlier investigations have already refuted their existence. This conclusion is concerning because of the differential public health implications of members of the Fluviatilis Complex. Here we discuss problems associated with the CO1-based barcode approach for delimitation of cryptic species, the perceived heterozygosity between species S and T based on a species-specific PCR assay, and interpretation of published reports. We demonstrated that fixed differences do exist in the ITS2-rDNA sequence of species S and T with no evidence of heterozygotes in sympatric populations and, that the observed heterozygosity by Kumar et al. in the ITS2-based species diagnostic PCR is due to the high mispriming tendency of the T-specific primer with species S. We infer that mitochondrial DNA-based barcoding-gap, an arbitrary threshold recommended for species delimitation, alone, is inadequate to delimit the members of An. fluviatilis complex.

Population Dynamics and Insecticide Susceptibility of Anopheles culicifacies in Malaria Endemic Districts of Chhattisgarh, India

A study was undertaken in the villages of Korea and Bastar district (Chhattisgarh) during the years 2012–2015 to investigate the bionomics of malaria vectors and the prevalence of their sibling species complexes. Entomological surveys carried out every month included indoor resting collections, pyrethrum spray catches, light trap catches, and insecticide susceptibility status of Anopheles culicifacies using World Health Organization (WHO) methods. Anopheles culicifacies and Anopheles fluviatilis species were assayed by polymerase chain reaction (PCR) for the detection of malaria parasite, and sibling species were identified using PCR and DNA sequencing. A total of 13,186 samples of Anopheles comprising 15 species from Bastar and 16 from Korea were collected. An. Culicifacies was recorded as the most dominant species and also the only active vector at both sites. This species was found to be resistant to dichlorodiphenyltrichloroethane (DDT) and Malathion, showing signs of emerging resistance against pyrethroids. Among the sibling species of An. culicifacies, the group BCE was found in maximum numbers, while sibling species T of the An. fluviatilis was recorded to be dominant among its complex. The study provides a comprehensive view of the vector bionomics in the highly malarious regions of India that may have importance in developing vector control strategies.

Ecology of Malaria Vectors in an Endemic Area, Southeast of Iran

Background: Malaria has long been regarded as one of the most important public health issues in Iran. Although the country is now in the elimination phase, some endemic foci of malaria are still present in the southeastern areas of the country. In some endemic foci, there are no data on the malaria vectors. To fill this gap, the present study was designed to provide basic entomological data on malaria vectors in the southeastern areas of Iran. Methods: Adult and larval stages of Anopheles mosquitoes were collected by using different catch methods. Resistance of the main malaria vector in the study area to selected insecticides was evaluated using diagnostic doses advised by the World Health Organization in 2013–2014. Results: A total of 3288 larvae and 1055 adult Anopheles mosquitoes were collected, and identified as: Anopheles stephensi (32.1%), Anopheles culicifacies s.l. (23.4%), Anopheles dthali (23.2%), Anopheles superpictus s.l. (12.7%), and Anopheles fluviatilis s.l. (8.6%). Anopheles stephensi was the most predominant mosquito species collected indoors at the study area, with two peaks of activity in May and November. This species was found to be resistant to DDT 4%, tolerant to malathion 5% and susceptible to other tested insecticides. Conclusion: All the five malaria vectors endemic to the south of Iran were collected and identified in the study area. Our findings on the ecology and resting/feeding habitats of these malaria vectors provide information useful for planning vector control program in this malarious area.

A study of malaria vector surveillance as part of the Malaria Elimination Demonstration Project in Mandla, Madhya Pradesh

Background Understanding of malaria vector density, distribution, insecticide resistance, vector incrimination, infection status, and identification of sibling species are some of the essential components of vector control measures for achieving malaria elimination goals. Methods As part of the malaria elimination demonstration project, entomological surveillance was carried out from October 2017 to October 2019 by collecting indoor resting mosquitoes using hand catch method. Susceptibility test was done for determining the insecticide resistance status of vector mosquito Anopheles culicifacies using standard protocols by the World Health Organization. The cone bioassay method was used for determining the efficacy and quality of insecticide sprayed. Mosquitoes collected from different ecotypes were identified and processed for parasite identification, vector incrimination and sibling species determination. Results The two known malaria vector species (Anopheles culicifacies and Anopheles fluviatilis) were found in the study area, which have been previously reported in this and adjoining areas of the State of Madhya Pradesh. The prevalence of An. culicifacies was significantly higher in all study villages with peak in July while lowest number was recorded in May. Proportion of vector density was observed to be low in foothill terrains. The other anopheline species viz, Anopheles subpictus, Anopheles annularis, Anopheles vagus, Anopheles splendidus, Anopheles pallidus, Anopheles nigerrimus and Anopheles barbirostris were also recorded in the study area, although their prevalence was significantly less compared to the An. culicifacies. In 2017, An. culicifacies was found to be resistant to dichloro-diphenyl-trichloroethane (DDT) and malathion, with possible resistance to alphacypermethrin and susceptible to deltamethrin. However, in 2019, the species was found to be resistant to alphacypermethrin, DDT, malathion, with possible resistance to deltamethrin. The bioassays revealed 82 to > 98% corrected % mortality of An. culicifacies on day-one post-spraying and 35 to 62% on follow-up day-30. Anopheles culicifacies sibling species C was most prevalent (38.5%) followed by A/D and E while B was least pre-dominant (11.9%). Anopheles fluviatilis sibling species T was most prevalent (74.6%) followed by U (25.4%) while species S was not recorded. One An.culicifacies (sibling species C) was found positive for Plasmodium falciparum by PCR tests in the mosquitoes sampled from the test areas. Conclusion Based on the nine entomologic investigations conducted between 2017–2019, it was concluded that An. culicifacies was present throughout the year while An. fluviatilis had seasonal presence in the study areas. Anopheles culicifacies was resistant to alphacypermethrin and emerging resistance to deltamethrin was observed in this area. Anopheles culicifacies was confirmed as the malaria vector. This type of information on indigenous malaria vectors and insecticide resistance is important in implementation of vector control through indoor residual spraying (IRS) and use of insecticide-impregnated bed nets for achieving the malaria elimination goals.

Climate Change and the Risk of Malaria Transmission in Iran

Climate change is an important factor affecting the dynamics of the vectors population and, hence, the risk of vector-borne diseases. This study aimed to predict the environmental suitability for malaria vectors in Iran under climate change scenarios in 2030s and 2050s. Literature search was performed to find documents on the spatial distribution of Anopheles stephensi Liston, 1901, Anopheles culicifacies s.l. Giles, 1901, Anopheles fluviatilis s.l. James, 1902, Anopheles superpictus s.l. Grassi, 1899, Anopheles dthali Patton, 1905, Anopheles maculipennis s.l. Meigen, 1818, and Anopheles sacharovi Favre, 1903 (Diptera: Culicidae) published between 1970 and 2017. The bioclimatic data under three climate change scenarios (representative concentration pathway 2.6 [RCP2.6], RCP4.5, and RCP8.5) and MaxEnt model were used to predict the ecological niches for each species. Comparison between the two study periods under the three scenarios for each species revealed that RCP8.5 would reduce the area at risk for An. culicifacies s.l., An. dthali and An. superpictus s.l. in the 2050s compared to the 2030s, but the reverse will be induced by RCP2.6 and RCP4.5 scenarios. For An. fluviatilis s.l., RCP2.6 will reduce the risk areas in the 2050s, whereas an increase is expected under the two other scenarios. Moreover, all scenarios would decrease the high-risk areas of An. maculipennis s.l. in the 2050s. For An. sacharovi, RCP2.6 would increase its high-risk areas, whereas RCP4.5 and RCP8.5 would decrease its exposure. The high-risk area of An. stephensi is expected to increase under RCP8.5 in the 2030s and RCP4.5 in 2050s, but it will be almost unchanged or reduced under other scenarios.