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Molecular evidence of speciation between island and continental populations of Anopheles (Cellia) sundaicus (Diptera: Culicidae), a principal malaria vector taxon in Southeast Asia

Dusfour I, Linton YM, Cohuet A, Harbach RE, Baimai V, Trung HD, et al.

J Med Entomol, 2004 May;41(3):287-95.
PMID: 15185927

Anopheles sundaicus s.l. is a principal malaria vector taxon on islands and along the coastal areas of Southeast Asia. It has a wide geographical distribution and exhibits a high level of ecological and behavioral variability. Study of this taxon is crucial for understanding its biology and implementing effectise vector control measures. We compared populations of An. sundaicus from Vietnam, Thailand, and Malaysian Borneo by using two mitochondrial DNA markers: cytochrome oxidase I and cytochrome b. Genetic divergence, geographic separation, and cladistic analysis of relationships revealed the presence of two cryptic species: Anopheles sundaicus s.s. on Malaysian Borneo and An. sundaicus species A in coastal areas of Thailand and Vietnam. A polymerase chain reaction (PCR) assay was developed to easily identify these two species throughout their geographic distributions. The assay was based on sequence characterized amplified region derived from random amplified polymorphic DNA. This PCR identification method needs to be validated and adapted for the recognition of other possible species in the Sundaicus Complex.

Matched MeSH terms: Anopheles/genetics*
The effect of Plasmodium vivax infection on SOCS gene expression in Anopheles dirus (Diptera: Culicidae)

Kittiwattanawong K, Ponlawat A, Boonrotpong S, Nanakorn N, Kongchouy N, Moonmake S, et al.

Trop Biomed, 2020 Jun 01;37(2):397-408.
PMID: 33612809

The Anopheles dirus mosquito is a primary malaria vector that transmits many species of Plasmodium parasites in Thailand and is widely spread across its geographic area. In the current study, the levels of expression of the suppressor of cytokine signaling (SOCS) gene in An. dirus mosquitoes infected with P. vivax were examined. The level of the gene's expression determined by mRNA extraction in An. dirus females (n=2,400) was studied at different times (0, 12, 24, 36, and 48 h after feeding), with different types of blood feeding (non-feeding, parasite-negative blood feeding, parasite-positive blood feeding) and in different parts of the body of mosquito samples (thorax and abdomen). The datasets were analyzed based on their relative expression ratio by the 2-ΔΔCT method and were tested for significant differences with ANOVA. The results showed that the An. dirus SOCS gene was stimulated in the abdomen 12 h and 24 h after blood feeding about three times more highly than in unfed females, with the difference being significant. At 24 h after P. vivax-infected blood feeding, the SOCS gene in the abdomen was expressed more highly than 24 h after parasite-negative blood feeding and expression was almost 36 times higher than in the control group who were not fed blood. However, in the thorax at all times after feeding and non-feeding, there was no expression of the SOCS gene. Therefore, the SOCS gene in An. dirus was most highly expressed 24 h post-feeding with a P. vivax-infected bloodmeal, which indicates that the SOCS gene in the major malaria vector in Thailand plays an important role in its immune system and its response to P. vivax infection.

Matched MeSH terms: Anopheles/genetics*
SCAR markers and multiplex PCR-based identification of isomorphic species in the Anopheles dirus complex in Southeast Asia

Manguin S, Kengne P, Sonnier L, Harbach RE, Baimai V, Trung HD, et al.

Med Vet Entomol, 2002 Mar;16(1):46-54.
PMID: 11963981

The Anopheles dirus Peyton & Harrison complex of mosquitoes (Diptera: Culicidae) comprises seven known species, including important malaria vectors in Southeast Asia. Specific identification of each species of the complex, which cannot be distinguished using morphological characters, is crucial for understanding vector ecology and implementing effective control measures. Derived from individual random amplified polymorphic DNA (RAPD) markers, sequence characterized amplified regions (SCAR) were analysed for the design of specific paired-primers. Combination of six SCAR primers resulted in the development of a simple, robust, single multiplex PCR able to identify three important malaria vectors among the four most common species (A, B, C, D) of the complex: species A from several Southeast Asian countries, species B from Perlis, Malaysia, and species C and D from Thailand.

Matched MeSH terms: Anopheles/genetics*
Molecular identification of Anopheles (Cellia) sundaicus from the Andaman and Nicobar islands of India

Alam MT, Das MK, Ansari MA, Sharma YD

Acta Trop, 2006 Jan;97(1):10-8.
PMID: 16125659

Anopheles (Cellia) sundaicus (Rodenwaldt) is an important malaria vector in the Andaman and Nicobar islands of India where it breeds in freshwater as well as in brackish water. To establish the molecular identity of An. sundaicus on these islands we analyzed samples from four geographically isolated areas-Teressa, Nancowry, Car Nicobar and Katchal islands. PCR-amplification and nucleotide sequence analysis were performed for internal transcribed spacer 2 (ITS2) and domain-3 (D3) of 28S rRNA. The ITS2 region of An. sundaicus from all four islands was identical but different from An. sundaicus A of Vietnam and An. sundaicus s.s of Malaysia. Furthermore, freshwater and brackish water forms of An. sundaicus did not reveal any sequence variation. Similarly, the D3 sequences were identical among all An. sundaicus samples from the four islands. D3 sequences for a species of the Sundaicus Complex are reported here for the first time and thus could not be compared with other regional isolates of this species. In conclusion, probably only one member of the Sundaicus Complex exists on the Andaman and Nicobar islands, which breeds in freshwater as well as in brackish water and is different from the An. sundaicus A and Malaysian An. sundaicus s.s. The identification of a new sibling species of the Sundaicus Complex in these islands is significant from the viewpoint of vector control strategies.

Matched MeSH terms: Anopheles/genetics
Fulltext Temporal and spatial trends in insecticide resistance in Anopheles arabiensis in Sudan: outcomes from an evaluation of implications of insecticide resistance for malaria vector control

Ismail BA, Kafy HT, Sulieman JE, Subramaniam K, Thomas B, Mnzava A, et al.

Parasit Vectors, 2018 03 02;11(1):122.
PMID: 29499751 DOI: 10.1186/s13071-018-2732-9

BACKGROUND: Long-lasting insecticidal nets (LLINs) (with pyrethroids) and indoor residual spraying (IRS) are the cornerstones of the Sudanese malaria control program. Insecticide resistance to the principal insecticides in LLINs and IRS is a major concern. This study was designed to monitor insecticide resistance in Anopheles arabiensis from 140 clusters in four malaria-endemic areas of Sudan from 2011 to 2014. All clusters received LLINs, while half (n = 70), distributed across the four regions, had additional IRS campaigns.
METHODS: Anopheles gambiae (s.l.) mosquitoes were identified to species level using PCR techniques. Standard WHO insecticide susceptibility bioassays were carried out to detect resistance to deltamethrin (0.05%), DDT (4%) and bendiocarb (0.1%). TaqMan assays were performed on random samples of deltamethrin-resistant phenotyped and pyrethrum spray collected individuals to determine Vgsc-1014 knockdown resistance mutations.
RESULTS: Anopheles arabiensis accounted for 99.9% of any anopheline species collected across all sites. Bioassay screening indicated that mosquitoes remained susceptible to bendiocarb but were resistance to deltamethrin and DDT in all areas. There were significant increases in deltamethrin resistance over the four years, with overall mean percent mortality to deltamethrin declining from 81.0% (95% CI: 77.6-84.3%) in 2011 to 47.7% (95% CI: 43.5-51.8%) in 2014. The rate of increase in phenotypic deltamethrin-resistance was significantly slower in the LLIN + IRS arm than in the LLIN-only arm (Odds ratio 1.34; 95% CI: 1.02-1.77). The frequency of Vgsc-1014F mutation varied spatiotemporally with highest frequencies in Galabat (range 0.375-0.616) and New Halfa (range 0.241-0.447). Deltamethrin phenotypic-resistance correlated with Vgsc-1014F frequency.
CONCLUSION: Combining LLIN and IRS, with different classes of insecticide, may delay pyrethroid resistance development, but the speed at which resistance develops may be area-specific. Continued monitoring is vital to ensure optimal management and control.

Matched MeSH terms: Anopheles/genetics
Fulltext Microsatellite and mitochondrial markers reveal strong gene flow barriers for Anopheles farauti in the Solomon Archipelago: implications for malaria vector control

Ambrose L, Cooper RD, Russell TL, Burkot TR, Lobo NF, Collins FH, et al.

Int J Parasitol, 2014 Mar;44(3-4):225-33.
PMID: 24440418 DOI: 10.1016/j.ijpara.2013.12.001

Anopheles farauti is the primary malaria vector throughout the coastal regions of the Southwest Pacific. A shift in peak biting time from late to early in the night occurred following widespread indoor residue spraying of dichlorodiphenyltrichloro-ethane (DDT) and has persisted in some island populations despite the intervention ending decades ago. We used mitochondrial cytochrome oxidase I (COI) sequence data and 12 newly developed microsatellite markers to assess the population genetic structure of this malaria vector in the Solomon Archipelago. With geographically distinct differences in peak A. farauti night biting time observed in the Solomon Archipelago, we tested the hypothesis that strong barriers to gene flow exist in this region. Significant and often large fixation index (FST) values were found between different island populations for the mitochondrial and nuclear markers, suggesting highly restricted gene flow between islands. Some discordance in the location and strength of genetic breaks was observed between the mitochondrial and microsatellite markers. Since early night biting A. farauti individuals occur naturally in all populations, the strong gene flow barriers that we have identified in the Solomon Archipelago lend weight to the hypothesis that the shifts in peak biting time from late to early night have appeared independently in these disconnected island populations. For this reason, we suggest that insecticide impregnated bed nets and indoor residue spraying are unlikely to be effective as control tools against A. farauti occurring elsewhere, and if used, will probably result in peak biting time behavioural shifts similar to that observed in the Solomon Islands.

Matched MeSH terms: Anopheles/genetics*
Fulltext New vectors in northern Sarawak, Malaysian Borneo, for the zoonotic malaria parasite, Plasmodium knowlesi

Ang JXD, Kadir KA, Mohamad DSA, Matusop A, Divis PCS, Yaman K, et al.

Parasit Vectors, 2020 Sep 15;13(1):472.
PMID: 32933567 DOI: 10.1186/s13071-020-04345-2

BACKGROUND: Plasmodium knowlesi is a significant cause of human malaria in Sarawak, Malaysian Borneo. Only one study has been previously undertaken in Sarawak to identify vectors of P. knowlesi, where Anopheles latens was incriminated as the vector in Kapit, central Sarawak. A study was therefore undertaken to identify malaria vectors in a different location in Sarawak.
METHODS: Mosquitoes found landing on humans and resting on leaves over a 5-day period at two sites in the Lawas District of northern Sarawak were collected and identified. DNA samples extracted from salivary glands of Anopheles mosquitoes were subjected to nested PCR malaria-detection assays. The small subunit ribosomal RNA (SSU rRNA) gene of Plasmodium was sequenced, and the internal transcribed spacer 2 (ITS2) and mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of the mosquitoes were sequenced from the Plasmodium-positive samples for phylogenetic analysis.
RESULTS: Totals of 65 anophelines and 127 culicines were collected. By PCR, 6 An. balabacensis and 5 An. donaldi were found to have single P. knowlesi infections while 3 other An. balabacensis had either single, double or triple infections with P. inui, P. fieldi, P. cynomolgi and P. knowlesi. Phylogenetic analysis of the Plasmodium SSU rRNA gene confirmed 3 An. donaldi and 3 An. balabacensis with single P. knowlesi infections, while 3 other An. balabacensis had two or more Plasmodium species of P. inui, P. knowlesi, P. cynomolgi and some species of Plasmodium that could not be conclusively identified. Phylogenies inferred from the ITS2 and/or cox1 sequences of An. balabacensis and An. donaldi indicate that they are genetically indistinguishable from An. balabacensis and An. donaldi, respectively, found in Sabah, Malaysian Borneo.
CONCLUSIONS: Previously An. latens was identified as the vector for P. knowlesi in Kapit, central Sarawak, Malaysian Borneo, and now An. balabacensis and An. donaldi have been incriminated as vectors for zoonotic malaria in Lawas, northern Sarawak.

Matched MeSH terms: Anopheles/genetics
Fulltext Genome-Wide Transcription and Functional Analyses Reveal Heterogeneous Molecular Mechanisms Driving Pyrethroids Resistance in the Major Malaria Vector Anopheles funestus Across Africa

Riveron JM, Ibrahim SS, Mulamba C, Djouaka R, Irving H, Wondji MJ, et al.

G3 (Bethesda), 2017 06 07;7(6):1819-1832.
PMID: 28428243 DOI: 10.1534/g3.117.040147

Pyrethroid resistance in malaria vector, An. funestus is increasingly reported across Africa, threatening the sustainability of pyrethroid-based control interventions, including long lasting insecticidal nets (LLINs). Managing this problem requires understanding of the molecular basis of the resistance from different regions of the continent, to establish whether it is being driven by a single or independent selective events. Here, using a genome-wide transcription profiling of pyrethroid resistant populations from southern (Malawi), East (Uganda), and West Africa (Benin), we investigated the molecular basis of resistance, revealing strong differences between the different African regions. The duplicated cytochrome P450 genes (CYP6P9a and CYP6P9b) which were highly overexpressed in southern Africa are not the most upregulated in other regions, where other genes are more overexpressed, including GSTe2 in West (Benin) and CYP9K1 in East (Uganda). The lack of directional selection on both CYP6P9a and CYP6P9b in Uganda in contrast to southern Africa further supports the limited role of these genes outside southern Africa. However, other genes such as the P450 CYP9J11 are commonly overexpressed in all countries across Africa. Here, CYP9J11 is functionally characterized and shown to confer resistance to pyrethroids and moderate cross-resistance to carbamates (bendiocarb). The consistent overexpression of GSTe2 in Benin is coupled with a role of allelic variation at this gene as GAL4-UAS transgenic expression in Drosophila flies showed that the resistant 119F allele is highly efficient in conferring both DDT and permethrin resistance than the L119. The heterogeneity in the molecular basis of resistance and cross-resistance to insecticides in An. funestus populations throughout sub-Saharan African should be taken into account in designing resistance management strategies.

Matched MeSH terms: Anopheles/genetics*