Prevalence of polymorphisms in marker genes associated with antimalarial drug resistance in Plasmodium falciparum following 10 years of artemisinin-based combination therapy implementation in urban Kolkata.

Context: Resistance to antimalarial drugs is one of the major challenges for malaria elimination. In India, artemisinin combination therapy (artesunate-sulfadoxin pyrimethamine) was introduced in place of chloroquine (CQ) for the treatment of uncomplicated falciparum malaria in 2010. Periodical monitoring of polymorphisms in antimalarial drug resistance marker genes will be useful for assessing drug pressure, mapping and monitoring of drug resistance status; and will be helpful for searching alternative treatments. Objectives: This study was conducted to study the polymorphisms in antimalarial drug resistance marker genes among clinical Plasmodium falciparum isolates collected from Kolkata after 10 years of artemisinin-based combination therapie (ACT) implementation. Materials and Methods: Blood samples were collected from P. falciparum mono-infected patients and polymorphisms in P. falciparum CQ resistance transporter (pfcrt), P. falciparum multidrug resistance (pfmdr-1), P. falciparum dihydrofolate reductase (pfdhfr), P. falciparum dihydropteroate synthetase (pfdhps), pfATPase6 and pfK-13 propeller genes were analysed by polymerase chain reaction and DNA sequencing. Results: In pfcrt gene, C72S, and K76T mutation was recorded in 100% isolates and no mutations was detected in any of the targeted codons of pfmdr-1 gene. A double mutant pfcrt haplotype SVMNT and wildtype haplotype NYD in pfmdr-1 were prevalent in 100% of study isolates. Triple mutant pfdhfr-pfdhps haplotype ANRNI-SGKAA was recorded. No polymorphism in pfK13 gene was documented in any of the isolates. Conclusions: Observed wild codon N86 along with Y184 and D1246 of pfmdr-1 gene might be an indication of the reappearance of CQ sensitivity. The absence of quadruple and quintuple haplotypes in pfdhfr-pfdhps gene along with the wild haplotype of pfK13 is evidence of ACT effectivity. Hence, similar studies with large sample size are highly suggested for monitoring the drug resistance status of P. falciparum.

Challenges for maintaining post elimination phase of visceral leishmaniasis control programme in India: A field-based study.

BACKGROUND: India is going through the maintenance phase of VL elimination programme which may be threatened by the persistence of hidden parasite pools among asymptomatic leishmanial infection (ALI) and PKDL. The present work was designed to determine the burden of VL, PKDL, and ALI and to assess the role of treatment of ALI in maintaining post-elimination phase. METHODS AND FINDING: The study was undertaken in Malda district, West Bengal, India during October 2016 to September 2021. Study areas were divided into 'Study' and 'Control' arms. VL and PKDL cases of both the arms were diagnosed by three active mass surveys with an interval of one year and treated as per National guideline. ALI of 'Study' arm was treated like VL. ALI of 'Control' arm was followed up to determine their fate. Fed sand-fly pools were analysed for parasitic DNA. No significant difference was noted between the incidence of VL and PKDL in both the arms. Incidence of ALI declined sharply in 'Study' arm but an increasing trend was observed in 'Control' arm. Significantly higher rate of sero-conversion was noted in 'Control' arm and was found to be associated with untreated ALI burden. Parasitic DNA was detected in 22.8% ALI cases and 2.2% sand-fly pools. CONCLUSION: Persistence of a significant number of PKDL and ALI and ongoing transmission, as evidenced by new infection and detection of leishmanial DNA in vector sand-flies, may threaten the maintenance of post-elimination phase. Emphasis should be given for elimination of pathogen to prevent resurgence of VL epidemics.

Assessment of Knowledge, Attitudes, and Practices about Visceral Leishmaniasis in Endemic Areas of Malda District, West Bengal, India.

Community participation is an important aspect for the success of kala-azar (KA) elimination program implemented in five Southeast Asian countries by the WHO. The participation of community depends on the level of knowledge of, attitude toward, and practice around risk factors associated with KA transmission among the population. We assessed the knowledge, attitude, and practice toward KA elimination in endemic areas of Malda district, West Bengal, India. A total of 709 individuals from different villages of 12 sub-centers were interviewed during April-July 2019. Data were recorded in a structured questionnaire under four categories: sociodemographic parameters, knowledge, attitude, and practice. The association of dependent variables such as knowledge, attitude, and practice with independent variables such as the economy and sociodemographic parameters was analyzed by binary logistic regression model and chi-square test using SPSS software. Despite the endemicity of the disease for a long time, the adequacy of knowledge about the disease was found to be poor that can be attributed to low education level and socioeconomic status, but the attitude and practices were good. So, there is a scope of improvement in knowledge of the disease through proper health education. This will further improve the level of attitude and practices that will be helpful for the smooth implementation of different activities of the program by more active participation of the community.

Seasonal variation of sand fly populations in Kala-azar endemic areas of the Malda district, West Bengal, India.

Vector control is one of the main aspects to reach the target of eliminating visceral leishmaniasis from Indian sub-continent as set by the World Health Organisation. Data on different aspects of vector like ecology, behaviour, population dynamics and their association with environmental factors are very important for formulating an effective vector control strategy. The present work was designed to study the species abundance and impact of environmental factors on population dynamics of vector P. argentipes in a visceral leishmaniasis endemic area of Malda district, West Bengal. Adult sand flies were collected using light traps and mouth aspirators from twelve kala-azar affected villages of Habibpur block of Malda district, on a monthly basis from January to December, 2018. Morphological and molecular methods were used for species identification. Population dynamics were assessed by man hour density and per night per trap collection. Data were analysed using SPSS software to determine the impact of environmental factors on vector population P. argentipes was found to the predominant species and prevalent throughout the year. A significantly higher number of sand flies were collected from cattle sheds than human dwellings and peri-domestic vegetation. A portion of the P. argentipes population was exophilic and exophagic as evidenced by their collection from peri-domestic vegetation. The highest population density was recorded during April to September. Population dynamics were mostly influenced by average temperature along humidity and rain fall. Resting behaviour of sand flies was not restricted to the lower portion of the wall but equally distributed throughout the wall and ceiling. Programme officials should consider management of outdoor populations of the sand flies and timings of indoor residual spray for chemical control purpose.

Insecticide susceptibility status of Phlebotomus argentipes and polymorphisms in voltage-gated sodium channel (vgsc) gene in Kala-azar endemic areas of West Bengal, India.

Rational use of insecticides, as advocated by World Health Organisation, plays a crucial role for vector control in eliminating visceral leishmaniasis from endemic countries. Emergence and spread of resistance among vector sand flies is of increasing concern for achieving these goals. Information on insecticide susceptibility status of sand fly populations and potential association between the former and polymorphisms in the insecticide target genes is important for formulating proper vector control measures. The present study was designed to evaluate the susceptibility status of vector sand fly species (Phlebotomus argentipes) against deltamethrin (type II pyrethroid), DDT (organochlorine) and malathion (organophosphate) and to detect polymorphisms in voltage gated sodium channel (vgsc) gene and investigating their association with type II pyrethroid and DDT susceptibility in three Kala-azar endemic districts of West Bengal, India. Adult sand flies were collected from human dwelling and cattle sheds of the study areas and subjected to insecticide bioassay using insecticide impregnated papers as per WHO protocol. Polymorphisms in domain II segment 6 of vgsc gene of pyrethroid and DDT susceptible and tolerant P. argentipes were detected by DNA sequencing. P. argentipes population of the study area was found to be susceptible to deltamethrin and malathion with corrected mortality rate between 98.02% to 98.80% and 98.81% to 100% respectively, but resistant to DDT (corrected mortality rate = 65.62%-76.33%). Two non-synonymous mutations L1014S and L1014F were detected of which L1014F was found to be associated with deltamethrin/DDT resistance. The replacement of DDT by synthetic pyrethroid is aptly done by national vector borne disease control programme (NVBDCP). The prevalence of L1014F mutation in vgsc gene and its association with type II pyrethroid tolerability is an indication of emergence of resistance against it. Malathion may be used as an alternative in the study areas if needed in future. Similar studies at a regular interval are highly suggested for monitoring susceptibility of used insecticide and to detect early signs of emergence of resistance against them.