Meta-analysis on Plasmodium falciparum sulfadoxine-pyrimethamine resistance-conferring mutations in India identifies hot spots for genetic surveillance.

BACKGROUND: India is on track to eliminate malaria by 2030 but emerging resistance to first-line antimalarials is a recognised threat. Two instances of rapid development, spread, and natural selection of drug-resistant mutant parasites in India (chloroquine across the country and artesunate + sulfadoxine-pyrimethamine [AS+SP] in the northeastern states) translated into drug policy changes for Plasmodium falciparum malaria in 2010 and 2013, respectively. Considering these rapid changes in the SP drug resistance-conferring mutation profile of P. falciparum, there is a need to systematically monitor the validated mutations in Pfdhfr and Pfdhps genes across India alongside AS+SP therapeutic efficacy studies. There has been no robust, systematic countrywide surveillance reported for these parameters in India, hence the current study was undertaken. METHODS: Studies that reported data on WHO-validated SP resistance markers in P. falciparum across India from 2008 to January 2023 were included. Five major databases, PubMedⓇ, Web of ScienceTM, ScopusⓇ, EmbaseⓇ, and Google Scholar, were exhaustively searched. Individual and pooled prevalence estimates of mutations were obtained through random- and fixed-effect models. Data were depicted using forest plots created with a 95% confidence interval. The study is registered with PROSPERO (CRD42021236012). RESULTS: A total of 37 publications, and 533 Pfdhfr and 134 Pfdhps National Centre of Biotechnology Information (NCBI) DNA sequences were included from >4000 samples. The study included information from 80 districts, 21 states and 3 union territories (UTs) from India. The two PfDHFR mutations, C59R (62%) and S108N (74%), were the most prevalent mutations (pooled estimates 61% and 71%, respectively) and appeared to be stabilised/fixed. Although rarest overall, the prevalence of I164L was observed to be as high as 32%. The PfDHFR double mutants were the most prevalent overall (51%; pooled 42%). The prevalence of triple and quadruple mutations was 6% and 5%, respectively, and is an immediate concern for some states. The most prevalent PfDHPS mutation was A437G (39%), followed by K540E (25%) and A581G (12%). There was a low overall prevalence of PfDHFR/PfDHPS quintuple and sextuple mutations but surveillance for these mutations is critical for some areas. CONCLUSION: The analyses span the two critical policy changes, highlight the areas of concern, and guide policymakers in strategising and refining the anti-malaria drug policy for malaria elimination. The results of the analyses also highlight the SP-resistance hot spots, critical gaps and challenges, and indicate that focal and local malaria genetic surveillance (including drug-resistance markers) is needed until malaria is successfully eliminated.

Plasmodium vivax Duffy Binding Protein-Based Vaccine: a Distant Dream.

The neglected but highly prevalent Plasmodium vivax in South-east Asia and South America poses a great challenge, with regards to long-term in-vitro culturing and heavily limited functional assays. Such visible challenges as well as narrowed progress in development of experimental research tools hinders development of new drugs and vaccines. The leading vaccine candidate antigen Plasmodium vivax Duffy Binding Protein (PvDBP), is essential for reticulocyte invasion by binding to its cognate receptor, the Duffy Antigen Receptor for Chemokines (DARC), on the host's reticulocyte surface. Despite its highly polymorphic nature, the amino-terminal cysteine-rich region II of PvDBP (PvDBPII) has been considered as an attractive target for vaccine-mediated immunity and has successfully completed the clinical trial Phase 1. Although this molecule is an attractive vaccine candidate against vivax malaria, there is still a question on its viability due to recent findings, suggesting that there are still some aspects which needs to be looked into further. The highly polymorphic nature of PvDBPII and strain-specific immunity due to PvDBPII allelic variation in Bc epitopes may complicate vaccine efficacy. Emergence of various blood-stage antigens, such as PvRBP, PvEBP and supposedly many more might stand in the way of attaining full protection from PvDBPII. As a result, there is an urgent need to assess and re-assess various caveats connected to PvDBP, which might help in designing a long-term promising vaccine for P. vivax malaria. This review mainly deals with a bunch of rising concerns for validation of DBPII as a vaccine candidate antigen for P. vivax malaria.

Multiplexing for Plasmodium spp.? Think Again! Comment on Bhowmick et al. Dry Post Wintertime Mass Surveillance Unearths a Huge Burden of P. vivax, and Mixed Infection with P. vivax P. falciparum, a Threat to Malaria Elimination, in Dhalai, Tripura, India. Pathogens 2021, 10, 1259

The study by Bhowmick et al. [...].

Global distribution of single amino acid polymorphisms in Plasmodium vivax Duffy-binding-like domain and implications for vaccine development efforts.

Plasmodium vivax (Pv) malaria continues to be geographically widespread with approximately 15 million worldwide cases annually. Along with other proteins, Duffy-binding proteins (DBPs) are used by plasmodium for RBC invasion and the parasite-encoded receptor binding regions lie in their Duffy-binding-like (DBL) domains-thus making it a prime vaccine candidate. This study explores the sequence diversity in PvDBL globally, with an emphasis on India as it remains a major contributor to the global Pv malaria burden. Based on 1358 PvDBL protein sequences available in NCBI, we identified 140 polymorphic sites within 315 residues of PvDBL. Alarmingly, country-wise mapping of SAAPs from field isolates revealed varied and distinct polymorphic profiles for different nations. We report here 31 polymorphic residue positions in the global SAAP profile, most of which map to the PvDBL subdomain 2 (α1-α6). A distinct clustering of SAAPs distal to the DARC-binding sites is indicative of immune evasive strategies by the parasite. Analyses of PvDBL-neutralizing antibody complexes revealed that between 24% and 54% of interface residues are polymorphic. This work provides a framework to recce and expand the polymorphic space coverage in PvDBLs as this has direct implications for vaccine development studies. It also emphasizes the significance of surveying global SAAP distributions before or alongside the identification of vaccine candidates.

Malaria diagnosis by PCR revealed differential distribution of mono and mixed species infections by Plasmodium falciparum and P. vivax in India.

Malaria is a vector-borne infectious disease, caused by five different species of the genus Plasmodium, and is endemic to many tropical and sub-tropical countries of the globe. At present, malaria diagnosis at the primary health care level in India is conducted by either microscopy or rapid diagnostic test (RDT). In recent years, molecular diagnosis (by PCR assay), has emerged as the most sensitive method for malaria diagnosis. India is highly endemic to malaria and shoulders the burden of two major malaria parasites, Plasmodium falciparum and P. vivax. Previous studies using PCR diagnostic assay had unraveled several interesting facts on distribution of malaria parasites in India. However, these studies had several limitations from small sample size to limited geographical areas of sampling. In order to mitigate these limitations, we have collected finger-prick blood samples from 2,333 malaria symptomatic individuals in nine states from 11 geographic locations, covering almost the entire malaria endemic regions of India and performed all the three diagnostic tests (microscopy, RDT and PCR assay) and also have conducted comparative assessment on the performance of the three diagnostic tests. Since PCR assay turned out to be highly sensitive (827 malaria positive cases) among the three types of tests, we have utilized data from PCR diagnostic assay for analyses and inferences. The results indicate varied distributional prevalence of P. vivax and P. falciparum according to locations in India, and also the mixed species infection due to these two species. The proportion of P. falciparum to P. vivax was found to be 49:51, and percentage of mixed species infections due to these two parasites was found to be 13% of total infections. Considering India is set for malaria elimination by 2030, the present malaria epidemiological information is of high importance.