Molecular analysis reveals a high diversity of Anopheles species in Karama, West Sulawesi, Indonesia.

BACKGROUND: Understanding local Anopheles species compositions and bionomic traits are vital for an effective malaria vector intervention strategy. Though eight malaria vectors, including species complexes, have been documented across the island of Sulawesi, Indonesia, a comprehensive survey linking morphological and molecular species identification has not been conducted in this global hotspot of biodiversity. RESULTS: Eighteen distinct species of Anopheles were molecularly identified in a 1 km2 area in Karama village, West Mamuju Province, Sulawesi. Known species included An. aconitus, An. karwari, An. peditaeniatus, An. vagus, An. barbirostris, An. tessellatus, An. nigerrimus, An. crawfordi, An. maculatus, An. flavirostris and An. kochi. Of the 18 distinct sequence groups identified through both ribosomal DNA internal transcribed spacer region 2, and mitochondrial DNA cytochrome c oxidase subunit 1 loci, 8 could not be identified to species through comparison to published sequences. The comparison of morphological and molecular identities determined that interpretations of local species compositions for primary and expected species in Karama (An. barbirostris and An. vagus) had the highest rate of accuracy (92.1% and 87.6%, respectively) when compared to molecular analysis. However, the remaining distinct sequences molecularly identified to species were identified correctly by morphological methods less frequently, from 0 to 83%. CONCLUSIONS: Karama, Indonesia has a high diversity of Anopheles spp. The unexpected high number of Anopheles species in a small area points to possible complex transmission dynamics and limitations with vector control based on possible varying behaviors and interactions with both humans and interventions. Morphological identification of Anopheles spp. in this study was more accurate for primary and expected species than secondary or unexpected species. Finally, the inability to identify seven sequence groups to species with consensus sequences implies that future studies employing sequencing are required to clarify species compositions in the Nigerrimus Subgroup, among others, as well as their distribution and vector status. Use of molecular methods in conjunction with morphological investigations for analysis of species composition, population dynamics and bionomic characteristics is directly implicated in understanding drivers of malaria transmission, intervention effectiveness, and the pursuit of malaria elimination.

Characterization of vector communities and biting behavior in South Sulawesi with host decoy traps and human landing catches.

BACKGROUND: Indonesia has high mosquito diversity, with circulating malaria and arboviruses. Human landing catches (HLC) are ethically questionable where arboviral transmission occurs. The host decoy trap (HDT) is an exposure-free alternative outdoor sampling device. To determine HDT efficacy for local culicids, and to characterize local mosquito fauna, the trapping efficacy of the HDT was compared to that of HLCs in one peri-urban (Lakkang) and one rural (Pucak) village in Sulawesi, Indonesia. RESULTS: In Lakkang the outdoor HLCs collected significantly more Anopheles per night (n = 22 ± 9) than the HDT (n = 3 ± 1), while the HDT collected a significantly greater nightly average of Culex mosquitoes (n = 110 ± 42), than the outdoor HLC (n = 15.1 ± 6.0). In Pucak, there was no significant difference in Anopheles collected between trap types; however, the HDT collected significantly more Culex mosquitoes than the outdoor HLC nightly average (n = 53 ± 11 vs 14 ± 3). Significantly higher proportions of blood-fed mosquitoes were found in outdoor HLC (n = 15 ± 2%) compared to HDT (n = 2 ± 0%). More blood-fed culicines were collected with outdoor HLC compared to the HDT, while Anopheles blood-fed proportions did not differ. For the HDT, 52.6%, 36.8% and 10.5% of identified blood meals were on cow, human, and dog, respectively. Identified blood meals for outdoor HLCs were 91.9% human, 6.3% cow, and 0.9% each dog and cat. Mosquitoes from Pucak were tested for arboviruses, with one Culex pool and one Armigeres pool positive for flavivirus, and one Anopheles pool positive for alphavirus. CONCLUSIONS: The HDT collected the highest abundance of culicine specimens. Outdoor HLCs collected the highest abundance of Anopheles specimens. Although the HDT can attract a range of different Asian mosquito genera and species, it remains to be optimized for Anopheles in Asia. The high proportion of human blood meals in mosquitoes collected by outdoor HLCs raises concerns on the potential exposure risk to collectors using this methodology and highlights the importance of continuing to optimize a host-mimic trap such as the HDT.

Mark-release-recapture studies reveal preferred spatial and temporal behaviors of Anopheles barbirostris in West Sulawesi, Indonesia.

BACKGROUND: Population density, dispersion patterns, flight distances, and survival rate of vector mosquitoes are all contributors to vectorial capacity that may be estimated in a single experimental method: mark-release-recapture (MRR). In this study, these key parameters were measured for mosquito populations in Karama, West Sulawesi, Indonesia. METHODS: Two mark-release-recapture (MRR) experiments were carried out in Karama village to characterize seasonality differences, if any: wet season (December 2013, MRR1) and dry season (May 2014, MRR2). For both experiments, mosquitoes were marked according to release site/date and were released on four consecutive nights. Four sampling methodologies were utilized to enable recapture: human landing catches (HLCs), kelambu traps and barrier screens. RESULTS: 98.7% of all catches were molecularly confirmed as Anopheles barbirostris. During the wet season, An. barbirostris demonstrated no preference toward endophagy. In the dry season, An. barbirostris demonstrated an endophagic preference. The duration of the feeding cycle for An. barbirostris was determined to be 5 days during the wet season and 3.7 days during the dry season, though an anomaly likely caused the wet season feeding cycle to be overestimated. The largest percentages of recaptured mosquitoes were collected in a single site during both seasons. The only significant relationship with mosquito dispersal was site of release and recapture. Finally, dispersal rates of An. barbirostris frequently ranged up to 800 m (the maximum measurable distance in this study) within a single day of release. CONCLUSIONS: This study estimated key vector parameters for An. barbirostris an understudied species complex, in Karama, West Sulawesi, Indonesia. Despite the length of the feeding cycle, the high indoor biting rates demonstrated by An. barbirostris in Karama suggest that the use of IRSs and LLINs, especially during the dry season, would have a substantial impact on the panmictic An. barbirostris population.

Using barrier screens to characterize mosquito composition, flight activity, and abdominal status in South Lampung, Indonesia.

BACKGROUND: Mosquito sampling methods target different aspects of mosquito behavior and are subject to trap and location specific biases. The barrier screen sampling method was developed and tested to sample free-flying, blood-fed, and host-seeking mosquitoes. During a pilot study, this method was useful in obtaining an unbiased sample of mosquitoes flying between outdoor larval habitats, and sites where blood meals were obtained. However, a relatively small number of blood-fed Anopheles mosquitoes were collected in Indonesia during the pilot study. The sampling method was extended in South Lampung, Indonesia, to enable the collection of blood-fed mosquitoes. This study aimed to intercept mosquitoes flying between human habitations and larval habitats with a barrier screen and to characterize mosquito composition, flight characteristics (direction, height and time), abdominal status, and parity. RESULTS: Barrier screens intercepted 15 different mosquito species in South Lampung: eight Anopheles spp. and seven Culex spp. Species compositions varied among the villages in South Lampung. About 15% of Anopheles spp. caught were blood-fed, of which 28.2% of those tested had fed on humans. This is the first time human blood-fed anophelines have been collected in Indonesia using barrier screens. Blood meals identified included cow, dog, goat, and human, as well as mixed blood meals. Activity of unfed An. subpictus, the primary vector collected, flying towards human habitations peaked between 20:00-12:00 h, with a slow decline in activity until 18:00 h. Unfed and fed An. sundaicus, had a different activity profile compared to An. subpictus. Other species demonstrated varied peak activity times, with earlier activity occurring as a general trend. For the Anopheles mosquitoes collected, 55.5% were collected below 0.5 m and 83.9% were captured resting < 1 m from the ground. Parity dissections enabled age structure by species, which revealed species-specific traits such as nulliparous An. subpictus being more active early in the night relative to An. sundaicus. CONCLUSIONS: This study demonstrates that barrier screens are an effective mosquito sampling method that can be used to gain insights into local mosquito species composition, flight characteristics (direction, height and time), abdominal status, and parity.