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The conventional paper-based system for malaria surveillance is time-consuming, difficult to track and resource-intensive. Few digital platforms are in use but wide-scale deployment and acceptability remain to be seen. To address this issue, we created a malaria surveillance mobile app that offers real-time data to stakeholders and establishes a centralised data repository. The MoSQuIT app was designed to collect data from the field and was integrated with a web-based platform for data integration and analysis. The MoSQuIT app was deployed on mobile phones of accredited social health activists (ASHA) working in international border villages in the northeast (NE) Indian states of Assam, Tripura and Arunachal Pradesh for 20 months in a phased manner. This paper shares the challenges and opportunities associated with the use of MoSQuIT for malaria surveillance. MoSQuIT employs the same data entry formats as the NVBDCP's malaria surveillance programme. Using this app, a total of 8221 fever cases were recorded, which included 1192 (14.5%) cases of P. falciparum malaria, 280 (3.4%) cases of P. vivax malaria and 52 (0.6%) mixed infection cases. Depending on network availability, GPS coordinates of the fever cases were acquired by the app. The present study demonstrated that mobile-phone-based malaria surveillance facilitates the quick transmission of data from the field to decision makers. Geospatial tagging of cases helped with easy visualisation of the case distribution for the identification of malaria-prone areas and potential outbreaks, especially in hilly and remote regions of Northeast India. However, to achieve the full operational potential of the system, operational challenges have to be overcome.

Background: Recent studies in experimental mice have shown that mild deficiency of methylenetetrahydrofolate reductase (MTHFR) enzyme confers protection against malaria, thus providing an important basis for the hypothesis that MTHFR polymorphism, i.e. C677T, might have been subjected to selection pressure against malaria. The present study was undertaken in a malaria endemic region in North East India to assess whether a similar selection advantage exists for other genes in folate metabolism pathway., Methods: A total of 401 subjects including 131 symptomatic malaria, 97 asymptomatic malaria and 173 normal healthy controls were analysed for nine polymorphisms (single-nucleotide polymorphisms [SNPs] in eight genes and insertion/deletion in one gene): MTHFR C677T, methionine synthase reductase (MTRR) A66G, glutamate carboxypeptidase II (GCPII) C1561T, cystathionine beta-synthase (CBS) 844ins68, reduced folate carrier-1 (RFC-1) G80A, serine hydroxymethyltransferase (SHMT) C1420T, methionine synthase (MTR) A2756G, MTHFR G1793A (D 919G), glycine N-methyltransferase (GNMT) 1289 by PCR-RFLP technique. Differences in frequencies of genotype distribution of each polymorphic marker between these groups were evaluated., Results: MTRR A2756G, SHMT C1420T, GCPII C1561T, MTRR A2756G and GNMT C1289T and RFC1 G80A polymorphisms showed significantly different prevalence between different groups analyzed. No significant differences were seen in the distribution of other polymorphisms., Conclusions: The study gives a clue for the possible selection of specific polymorphisms in the genes involved in the folate metabolism pathway by malaria parasite., (© The Author 2016. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The mechanism of chloroquine (CQ) resistance in Plasmodium falciparum is not clearly understood. However, CQ resistance has been shown to be associated with point mutations in Pfcrt and Pfmdr1. These genes encode for digestive vacuole transmembrane proteins Pfcrt and Pgh1, respectively. The present study was carried out to analyze the association of Pfcrt-K76T and Pfmdr1-N86Y mutations with CQ resistance in Northeast Indian P. falciparum isolates. 115 P. falciparum isolates were subjected to in vitro CQ sensitivity testing and PCR-RFLP analysis for the Pfmdr1-N86Y and Pfcrt-K76T mutations. 100 isolates of P. falciparum were found to be resistant to CQ by the in vitro susceptibility test (geometric mean EC50 2.21 µM/L blood) while 15 were found to be CQ sensitive (geometric mean EC50 0.32 µM/L blood). All the CQ resistant isolates showed the presence of Pfmdr1 and Pfcrt mutations. CQ sensitive isolates were negative for these mutations. Strong linkage disequilibrium was observed between the alleles at these two loci (Pfmdr1-N86Y and Pfcrt-K76T). The results indicate that Pfmdr1-N86Y and Pfcrt-K76T mutations can be used as molecular markers to identify CQ resistance in P. falciparum. The result necessitates the evaluation of CQ in vivo therapeutic efficacy in endemic areas for more effective malaria control strategies.

Background: Febrile illness is the most common symptom experienced by patients suspected of having malaria, especially in the malaria endemic areas of northeast India. Hospital-based data from two districts of upper Assam, namely Golaghat and Tinsukia, were analysed to assess the prevalence of malaria among inpatients who reported to health centres with febrile illness., Methods: A total of 16 hospitals were selected for the study. These included six government hospitals (three in rural and three in urban areas) and 10 private hospitals, which included four tea-garden hospitals. Selection was dependant on the availability of patients and inpatient treatment facilities. During the study, a total of 350 inpatients reported to the selected hospitals with fever; 324 were suspected of having malaria and were interviewed and included in the study., Results: The average age of patients was 24.2±15.2 years and 63.3% (205/324) of them were male. The majority of patients (77.5%; 251/324) belonged to the lower household income category (<5000 Indian rupees) and travelled to the nearest town or city for treatment. Overall, the prevalence of malaria (as diagnosed by clinical examination and blood tests) across the different categories such as location, types of hospitals and household income was found to be 29.9% (97/324). Multivariate analysis revealed that the adjusted odds ratios (AOR) of malaria cases was higher based on location (2.13), type of health centre (1.75) and the distance travelled to a health centre (2.09). The mean duration of hospital treatment was 4.2±3.6 days and the delay in reporting to hospital was 3.9±2.6 days., Conclusion: The study emphasises the need to strengthen and improve the treatment facilities for malaria in government hospitals, and to create more awareness among people regarding early treatment, especially in the rural periphery villages of the endemic areas., (© The Author 2014. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Malaria control is compromised worldwide by continuously evolving drug-resistant strains of the parasite demanding exploration of natural resources for developing newer antimalarials. The northeastern region of India is endemic for malaria characterized by high prevalence of drug-resistant Plasmodium falciparum strains. Many plants are used by the indigenous communities living in the northeast India in their traditional system of medicine for the treatment of malarial fever. Folklore claim of antimalarial property of one such plant Brucea mollis was evaluated in vitro and in vivo for antiplasmodial activity. Crude extracts from dried B. mollis root powder were prepared through soxhlet extraction using petroleum ether, methanol, and water sequentially. Methanol extract was further partitioned between chloroform and water. These extracts were tested in vitro against laboratory-adapted chloroquine-sensitive and chloroquine-resistant strains of P. falciparum. In in vitro evaluation, extracts were found more active on the chloroquine-sensitive strain. Methanolic-chloroform (IC(50) 5.1 μg ml(-1)) and methanolic-aqueous (IC(50) 13.9 μg ml(-1)) extracts recorded significant in vitro antiplasmodial activity which was also supported by their promising in vivo activity (ED(50) 72 and 30 mg kg(-1) bw day(-1), respectively) against chloroquine-resistant Plasmodium yoelli N-67 strain in Swiss albino mice. Methanolic-aqueous extract-treated mice survived on average for 14 days that was comparable to the reference drug chloroquine. This is the first report of antiplasmodial activity of B. mollis validating the traditional use of this plant as antimalarial in the northeast India and calls for further detailed investigations.

A study was conducted to evaluate the preventive efficacy of insecticide-treated mosquito nets (ITMNs) and mosquito repellent (MR) in a malaria-endemic foothill area of Assam, India, with forest ecosystem. During the first year, a survey was conducted in four demarcated sectors (A-D) to observe the malaria endemicity and vector prevalence patterns before implementing intervention measures. All four sectors were endemic for malaria. The prevalence of established malaria vectors such as Anopheles dirus, A. minimus and A. philippinensis was observed. During the second year, intervention measures were implemented in the four sectors as follows: A, ITMN + MR; B, ITMN; C, MR; D, no intervention. The most effective intervention was in sector A, followed by sectors B and C. Sectors A and B exhibited significantly higher (P < 0.001) malaria protective efficacy during both the first and second years of intervention compared with sector D. The total vector population in the three intervention sectors decreased significantly compared with that of the non-intervention one. Information-education-communication activities motivated the residents to participate actively in the intervention programme. The finding could be an effective model for containment of high malaria morbidity in inaccessible forest fringe areas of the northeastern region of India., (Copyright © 2011 Royal Society of Tropical Medicine and Hygiene. Published by Elsevier Ltd. All rights reserved.)

Entomological survey was conducted to know the breeding habitat preference of the forest breeder malaria vector Anopheles baimaii, known earlier as An. dirus species D in the northeastern region of India. Breeding potential of the vector in forest areas was found to be high in water stored in jungle pool (69.84%) followed by elephant footprints with clear water (39.13%) and with turbid water (26.19%), whereas in forest fringe areas, the vector breeding was more prominent in elephant footprints: 65.11% in clear water and 62.5% in turbid water. Although other habitats had shown only low breeding of the vector, all types of habitats were positively correlated with malaria occurrence. Cattle hoof marks (r = 0.998) and elephant footprint (turbid; r = 0.999) explained nearly the same amount of variance. It was observed that deforestation as well as elephant habitat-type destruction had engendered man-elephant conflicts intensively in fringe areas. Seasonal abundance pattern of this vector was found to vary in forest and forest fringe areas in relation to different habitats. Seasonal abundance of An. baimaii was significantly different in different habitats. The Tukey post hoc comparisons indicated that the abundance of An. baimaii in different habitats was significantly higher (P < 0.05) in monsoon season than that of premonsoon and postmonsoon seasons. No significant difference was observed between premonsoon and postmonsoon seasons. The findings therefore will eventually help to predict transmission of malaria in targeted area and in formulating an improved malaria control program in the northeastern region of India.

Background: This paper studies the determinants of utilization of health care services, especially for treatment of febrile illness in the malaria endemic area of north-east India., Methods: An area served by two districts of Upper Assam representing people living in malaria endemic area was selected for household survey. A sample of 1,989 households, in which at least one member of household suffered from febrile illness during last three months and received treatment from health service providers, were selected randomly and interviewed by using the structured questionnaire. The individual characteristics of patients including social indicators, area of residence and distance of health service centers has been used to discriminate or group the patients with respect to their initial and final choice of service providers., Results: Of 1,989 surveyed households, initial choice of treatment-seeking for febrile illness was self-medication (17.8%), traditional healer (Vaidya)(39.2%), government (29.3%) and private (13.7%) health services. Multinomial logistic regression (MLR) analysis exhibits the influence of occupation, area of residence and ethnicity on choice of health service providers. The traditional system of medicine was commonly used by the people living in remote areas compared with towns. As all the febrile cases finally received treatment either from government or private health service providers, the odds (Multivariate Rate Ratio) was almost three-times higher in favour of government services for lower households income people compared to private., Conclusion: The study indicates the popular use of self-medication and traditional system especially in remote areas, which may be the main cause of delay in diagnosis of malaria. The malaria training given to the paramedical staff to assist the health care delivery needs to be intensified and expanded in north-east India. The people who are economically poor and living in remote areas mainly visit the government health service providers for seeking treatment. So, the improvement of quality health services in government health sector and provision of health education to people would increase the utilization of government health services and thereby improve the health quality of the people.

Background & Objective: The National Vector Borne Disease Control Programme (NVBDCP), India has been promoting and scaling up the use of insecticide-treated nets (ITNs) in the tribal dominated malarious areas of north-east India. But, information on sleeping habits, bed net ownership and use practices, and feedback of communities in ITN--targeted areas is needed for formulating a strategic framework for upscaling the coverage of ITNs. We carried out a community-based cross-sectional survey in select areas of Nagaland and Mizoram (where ITNs were introduced) along with Assam (where ITNs were not introduced) to know the response of community., Methods: Following large scale introduction of ITNs during 2001-2002 in the north-eastern States of Nagaland and Mizoram by NVBDCP, India, a cross-sectional community-based survey was undertaken in April-May, 2003 covering 435 households of Nagaland and 464 households in Mizoram, using a structured questionnaire, to assess the demographic variables relevant to bed net use, bed net washing practices and acceptability of ITNs etc., for upscaling the coverage of ITNs in the surveyed communities. A total of 448 households in 8 villages in a non-ITN PHC area of Assam were served as the control area., Results: The average bet net usage per family (2.01 to 2.65). Bed net use was mostly seasonal in Nagaland where a majority (65.1-78.7%) used bed nets only during summers and monsoon as compared to the year round use in Mizoram (83.5%) and Assam (78.9%). Frequent washing of nets was most common in Assamese communities with 77 per cent households washing their nets at least once in a month. More than two third users favoured use of ITNs over the conventional indoor residual spray of DDT for malaria control., Interpretation & Conclusion: ITNs are widely acceptable in the user communities and a demand for ITNs is evident in the non-user communities of north-east India. Approach of treating community owned nets through an efficient service delivery mechanism will be a viable option for upscaling the ITN coverage.

Background & Objective: During a malaria epidemiological study in Arunachal Pradesh, Plasmodium malariae like human malaria parasites were seen in blood smears from fever cases. The study was undertaken to detect the presence of P. malariae and to confirm its identity through DNA based polymerase chain reaction approach., Methods: Fever survey was carried out in 22 villages in Indo-Myanmar bordering district of Lohit, Arunachal Pradesh in 2005. Morphologically suspected P. malariae cases were confirmed using nested PCR based on 18S small subunit ribosomal DNA gene sequence., Results: Screening of 1,995 fever cases resulted in 9 probable cases of P. malariae based on morphological identification in Chakma tribe people residing in 2 villages. Nested PCR confirmed the identity of all probable cases of P. malariae by producing diagnostic band of 144 bp. PCR method was able to detect mixed infection of P. malariae with P. vivax and with P. falciparum., Interpretation & Conclusion: P. malariae may have been present in Arunachal Pradesh but most probably is being misdiagnosed due to its close resemblance with P. vivax, especially in ring forms. Estimation of actual case load of P. malariae in north-east India is, therefore, important with accurate species identification using molecular methods.

The sibling species composition of the Anopheles minimus and Anopheles dirus complexes is poorly known in the highly malarious north-eastern region of India where these two vector taxa are accountable for most of the malaria transmission among 30.7 million inhabitants. Prevalent members of these two complexes in this part of India were identified using sequences for the second internal transcribed spacer (ITS2) of ribosomal DNA. Anopheles baimaii (species D) of the An. dirus complex and An. minimus s.s. (species A) of the An. minimus complex were detected in Arunachal Pradesh, Assam, Meghalaya and Nagaland states. No intraspecific variation was observed in the ITS2 sequence (479bp) of An. baimaii whereas a single substitution was detected in the ITS2 sequence (372bp) of An. minimus from Nagaland state.

The risk of acquiring malaria transmitted by Anopheles baimaii Sallum & Peyton, 2005, formerly known as An. dirus species D (Sallum et al., 2005) (Diptera: Culicidae), at different hours of the night in a forest-fringed village of Assam, North-east India was assessed through all-night mosquito landing catches during 1995-2000. An estimated overall mean biting rate of 36.1 bites/person/night (95% CI = 26.2-45.8), a sporozoite rate of 1.9% (95% CI = 1.1-2.9%) and a parous rate of 58.7% (95% CI = 55.3-62.0%) were recorded. Parous and sporozoite-positive females tended to be caught mainly before midnight. The effective entomological inoculation rate was the highest (0.249 positive bites/person/night) from 21.00 to 24.00 hours, suggesting that the second quartile of the night is the most risky period for malaria transmission by An. baimaii. Considering that approximately 21% of mean inoculations take place before 21.00 hours, it appears that there is a need for appropriate protective measures during the pre-bed time period to supplement the impact of insecticide-treated nets against An. baimaii in north-east India.

Anopheles philippinensis-nivipes complex mosquitoes, captured in outdoor human landing catches and light traps in human dwellings from four different sites in Assam state and adjoining areas, were examined (n=1670) for the presence of circumsporozoite antigen (CSA) through enzyme linked immunosorbent assays (ELISA), using species specific capture monoclonal antibodies, of Plasmodium falciparum and two Plasmodium vivax polymorphs (Pv 210 and VK 247). In ELISA, 28 pools were found positive for CSA that accounted for minimum sporozoite rate of 1.7% (95% CI 1.11-2.41). Twenty five percent (7/28) of the positive pools were reactive for P. falciparum and between the two polymorphs of P. vivax, VK 247 was predominant with 77% (20/26) of all P. vivax positive pools. Results were suggestive of most likely involvement of Anopheles philippinensis-nivipes complex mosquitoes in malaria transmission in north-east India.

To estimate the vectorial capacity of Anopheles dirus, the main vector of forest malaria in the northeastern region of India, in order to gain an understanding of entomological factors related to malaria transmission in forest-fringe areas of Assam, India, an isolated village in the tropical rain forest-fringed area in the district of Dibrugarh, Assam, under the influence of An. dirus alone was studied. Data on various entomological variables required for computation of the vectorial capacity were generated in each month from June 1999 to May 2000 in the field using standard techniques. Malaria prevalence was also studied during the same period in the study village and correlated with the estimated vectorial capacity of An. dirus. Vectorial capacity of An. dirus was highest, 0.779 for Plasmodium vivax (Pv) and 0.649 for Plasmodium falciparum (Pf), during the hot-monsoon season (June-September) and decreased to 0.08 (Pv) and 0.07(Pf) in the temperate postmonsoon season (October-November) before attaining zero values in the cool-dry season (December-February). With increasing temperature in the temperate premonsoon season (March-May), vectorial capacity recorded was 0.119 and 0.82 for Pv and Pf, respectively. Significant positive correlation was seen between the estimated vectorial capacity of An. dirus and the number of new Pf (r = 0.86, p < 0.001) and Pv (r = 0.69, p < 0.02) cases in the study village in different months. Thus, this study highlights the pattern of malaria transmission by An. dirus in a forest-fringe area of Assam that begins in March, peaks in July/August, subsides by November, and remains interrupted between December and February. Measures for controlling malaria in forest-fringe areas should be scheduled accordingly.

In an entomological study, covering three biotopes like village, panikheti (wet rice cultivated land) and urban agglomeration, a total of 35 species of mosquitoes in eight genera were recorded in Jairampur area of Arunachal Pradesh. Two mosquito species--Orthopodomyia anopheloides and Tripteroides indicus were recorded for the first time in Arunachal Pradesh. Anopheles dirus and An. minimus, the two well-known malaria vectors were encountered in the study along with several potential vector species like An. annularis, An. culicifacies, An. maculatus group and An. philippinensis/nivipes. The diversity of mosquito species as well as the man-vector contact was maximum in the panikheti and minimum in the village biotope.

An investigation was undertaken of a malaria outbreak in the Primary Health Centre Titabor, district Jorhat, Assam during May/June 1999. The fever rate in the community since March 1999, was 44.4 per cent with an average case load of 2.5 per family. The fever cases peaked in the third week of May. Slide positive and slide falciparum rates in mass blood survey, in the study village were 16.1 and 14.5 per cent respectively with 90 per cent infection of Plasmodium falciparum. Males (SPR 17.5%) suffered relatively more than females (SPR 14.7%). Malaria prevalence was significantly less in individuals above 15 yr of age (SPR 11.0%) as compared to those below 15 yr (SPR 22.9%). Prevalence of malaria as well as mosquito densities in different clusters of the village were inversely related to the distance from the forested Naga hills. Anopheles minimus and A. dirus were collected in good numbers with comparatively higher densities of the former. Several factors like unusual climatic conditions, inadequate surveillance, unsatisfactory laboratory services and inadequate indoor residual insecticide spray were instrumental for the outbreak.

In an entomological study in South Tripura district of Tripura state during June 1998, a total of 31 species of mosquitoes belonging to eight genera were recorded. Among the known malaria vectors in India, Anopheles annularis, An. jeyporiensis, An. minimus, An. philippinensis/nivipes and An. varuna were collected but none of them could be incriminated. During the survey, Aedes pallidostriatus and Malaya genurostris were recorded for the first time in the north-eastern region and Tripura state respectively. Presence of host seeking An. minimus in light-trap collections indoors and its absence during morning indoor resting collections indicated the probability of some degree of exophilic behaviour of An. minimus population in Tripura.

Seasonal abundance of Anopheles dirus (s.l.) and malaria prevalence in an isolated forest fringed village was monitored at monthly intervals during August 1995 to July 1996. An. dirus was the only vector species detected during the study period. Its density pattern showed distinct seasonality with the peak occurring in the month of July and very low number during cool dry months. Positive correlation (r = 0.721) was found between the density of An. dirus and the amount of rainfall occurring two weeks prior to the collections. Overall sporozoite rate of 1.6% and parous rate of 64.7% were found in the study. Malaria transmission closely followed the density pattern of An. dirus and was seasonal with slide positivity rate and P. falciparum percentage of 47 and 83% respectively. Mean malaria prevalence was higher (p < 0.05) in females.

The feeding behavior of Anopheles dirus, the forest breeding, major malaria vector in northeast India was studied. The analysis of blood meals collected from this mosquito revealed that the species was highly anthropophilic in nature, the anthropophilic index being 90.5. The results of bait collection on human and cattle bait also confirmed its biting preference for human hosts. The species was observed to land on human bait throughout the night, showing prominent biting time at 20.00-21.00, 23.00-24.00 and 02.00-03.00 hours during the study period.

Background: The emergence of antimalarial drug resistance malaria parasite is widespread in North eastern region of India. During January 2012-December 2013, we conducted active surveillance for detection of antifolate resistance-associated genetic polymorphisms in Plasmodium falciparum malaria parasite from different malaria endemic areas of Assam. Materials and Methods: A total of 281 field samples were collected from suspected malaria patients of which 106 malaria P. falciparum positive cases were detected in microscopic slide examination. A nested PCR was done for amplification of a 648 bp portion of the dhfr gene and 710 bp portion of the dhps gene. Results: Mutation analysis revealed existence of three different haplotypes of the P. falciparum dhfr gene of which ANRNI was highly prevalent (90%). Triple mutant haplotypes AIRNI (N51I + C59R + S108N) of the dhfr gene associated with pyrimethamine resistance were prevalent in Chirang district of Assam. Whereas, dhps mutation study revealed that triple mutant haplotype AGEAA (S436A + A437G + K540E) associated with Sulphadoxine resistance was found among 26% of P. falciparum field isolates. However, P. falciparum dhfr-dhps two locus mutation analysis showed that there were a total of nine dhfr-dhps genotypes. Conclusion: It was noticed that 93.62% (88/94) isolates had mutations in the sequences of both enzymes, which is an indication of prevalence of high grade of Sulphadoxine -- pyrimethamine resistance in P. falciparum malaria parasites in Assam. [ABSTRACT FROM AUTHOR]

Background & objectives: Severe anaemia in Plasmodium falciparum (Pf) associated malaria is a leading cause of death despite low levels of parasitaemia. In an effort to understand the pathogenesis of anaemia we studied expression level of RBC complement regulatory proteins, CR1 (CD35), CD55 and CD59 with haemoglobin status in a group of malaria cases from Assam, Goa and Chennai, and in healthy controls. Methods: Flowcytometry was used to study expression of CRI, CD55 and CD59 in 50 Pf cases and 30 normal healthy volunteers. Giemsa stained thick and thin blood films were used for microscopic detection and identification of malarial parasites and parasite count. Results: No correlation was found between degree of expression of RBC surface receptors CR1, CD55 and CD59 with haemoglobin level. However, expression of CD55 was less in malaria cases than in healthy controls. Interpretation & conclusions: The present findings indicate that malaria infection changes the expression profile of complement regulatory protein CD55 irrespective of severity status of anaemia. Further studies are needed to explore the pathophysiology of anaemia in malaria cases in Assam where expression of RBC complement receptors appears to be low even in normal healthy population. [ABSTRACT FROM AUTHOR]

Abstract: Anopheles philippinensis-nivipes species of mosquitoes are considered secondary malaria vectors in the north-east India. However, difficulty in separating An. philippinensis and An. nivipes accurately on the basis of morphological identification, due to the overlapping female wing characters, has led to the paucity of information on their spatial distribution and vectorial importance, essential for understanding malaria disease epidemiology in areas under their influence. In view of this a study was undertaken to generate authentic information on distribution of An. philippinensis and An. nivipes in the north-east India using molecular tools. Between the two, An. nivipes was recorded as the dominant species in Assam, and Nagaland states whereas An. philippinensis was found predominant in the state of Mizoram and Arunachal Pradesh. Species differentiation based on female wing characters proved wrong in about 74% specimens of An. philippinensis but found accurate in case of An. nivipes. No intra-specific variation in second internal transcribed spacer (ITS2) region of ribosomal DNA(r-DNA) in either An. nivipes or An. philippinensis from different geographical areas of north-east India was noted. [Copyright &y& Elsevier]

Abstract: The identity and distribution of the prevalent member species of the Anopheles dirus complex mosquitoes in the north-eastern region of India was investigated in a cross-sectional study. We altogether collected 267 individuals of An. dirus s.l. from 27 forested/forest fringed locations spread across the seven north-eastern states, identified the species using a ribosomal DNA (r-DNA) second internal transcribed spacer (ITS2) based allele specific polymerase chain reaction (ASPCR) method and sequenced the ITS2 locus in a sub set of mosquitoes. An. baimaii was identified as the main, almost exclusive (266/267), species of the Dirus complex throughout the north-east India with no intraspecific variation in the 479 base pair long ITS2 sequences in 59 of the 60 individuals sequenced. Ribosomal DNA of one individual from Assam state did not amplify in the ASPCR, possessed 786 base pair long ITS2 sequence and showed 99.7% similarity with the sequence of An. dirus species D (An. baimaii) from Yunnan province of China, later referred to as species X of the Dirus complex. These observations suggest the presence of another, possibly the new, species of the Dirus complex, sympatric with An. baimaii, in Assam warranting investigations on its distribution, biology and role in human malaria transmission in north-east India. [Copyright &y& Elsevier]