A randomised controlled trial to compare the efficacy, safety, and tolerability of low dose, short course primaquine in adults with uncomplicated P. vivax malaria in two hospitals in India.

BACKGROUND: Plasmodium vivax remains a major challenge for malaria control and elimination due to its ability to cause relapsing illness. To prevent relapses the Indian National Center for Vector Borne Diseases Control (NCVBDC) recommends treatment with primaquine at a dose of 0.25 mg/kg/day provided over 14 days. Shorter treatment courses may improve adherence and treatment effectiveness. METHODS: This is a hospital-based, randomised, controlled, open-label trial in two centres in India. Patients above the age of 16 years, with uncomplicated vivax malaria, G6PD activity of ≥ 30% of the adjusted male median (AMM) and haemoglobin levels ≥ 8 g/dL will be recruited into the study and randomised in a 1:1 ratio to receive standard schizonticidal treatment plus 7-day primaquine at 0.50 mg/kg/day or standard care with schizonticidal treatment plus 14-day primaquine at 0.25 mg/kg/day. Patients will be followed up for 6 months. The primary endpoint is the incidence risk of any P. vivax parasitaemia at 6 months. Safety outcomes include the incidence risk of severe anaemia (haemoglobin < 8 g/dL), the risk of blood transfusion, a > 25% fall in haemoglobin and an acute drop in haemoglobin of > 5 g/dL during primaquine treatment. DISCUSSION: This study will evaluate the efficacy and safety of a 7-day primaquine regimen compared to the standard 14-day regimen in India. Results from this trial are likely to directly inform national treatment guidelines. TRIAL REGISTRATION: Trial is registered on CTRI portal, Registration No: CTRI/2022/12/048283.

Vaccination coverage and breakthrough infections of COVID-19 during the second wave among staff of selected medical institutions in India.

India experienced the second wave of SARS-CoV-2 infection from April 3 to June 10, 2021. During the second wave, Delta variant B.1617.2 emerged as the predominant strain, spiking cases from 12.5 million to 29.3 million (cumulative) by the end of the surge in India. Vaccines against COVID-19 are a potent tool to control and end the pandemic in addition to other control measures. India rolled out its vaccination programme on January 16, 2021, initially with two vaccines that were given emergency authorization-Covaxin (BBV152) and Covishield (ChAdOx1 nCoV- 19). Vaccination was initially started for the elderly (60+) and front-line workers and then gradually opened to different age groups. The second wave hit when vaccination was picking up pace in India. There were instances of vaccinated people (fully and partially) getting infected, and reinfections were also reported. We undertook a survey of staff (front line health care workers and supporting) of 15 medical colleges and research institutes across India to assess the vaccination coverage, incidence of breakthrough infections, and reinfections among them from June 2 to July 10, 2021. A total of 1876 staff participated, and 1484 forms were selected for analysis after removing duplicates and erroneous entries (n = 392). We found that among the respondents at the time of response, 17.6% were unvaccinated, 19.8% were partially vaccinated (received the first dose), and 62.5% were fully vaccinated (received both doses). Incidence of breakthrough infections was 8.7% among the 801 individuals (70/801) tested at least 14 days after the 2nd dose of vaccine. Eight participants reported reinfection in the overall infected group and reinfection incidence rate was 5.1%. Out of (N = 349) infected individuals 243 (69.6%) were unvaccinated and 106 (30.3%) were vaccinated. Our findings reveal the protective effect of vaccination and its role as an essential tool in the struggle against this pandemic.

Economic burden estimation associated with dengue and chikungunya in Gujarat, India.

Background: Dengue and chikungunya have been emerging as major vector-borne diseases. The global burden of the diseases is rising as a public health problem. The complexity of disease is governed by multiple constraints including only symptomatic treatment and inflicts heavy social and economic burden on society. The present study is designed to assess the economic burden of dengue and chikungunya infection by calculating cost per patient in Gujarat, India. Methods: A total of 210 patients were enrolled in the study from Ahmedabad and Kheda district of Gujarat from May 2018 to December 2019 of which 150 had dengue and 60 chikungunya infections, subject to the willingness of participation in the survey. Information on wage loss days, cost associated with medicines, diagnosis, special food and travel cost, etc., for the calculation of the direct and indirect costs associated with dengue and chikungunya were collected from these participants using a structured questionnaire. Informed consent was taken before including any participant in the study. Results: In the dengue sample, 86 were males (57.3%) and the rest were females, and in the chikungunya sample, 31 were males (51.7%) and the rest females. The median age of the participants with dengue and chikungunya was 18 (p25 to p75: 8 to 26) and 30 (p25 to p75: 21 to 45) years respectively. Median family income was recorded as Rs 15,000 (p25 to p75: 9000 to 25500) and Rs 12,000 (p25 to p75: 9000 to 18500) for the dengue and chikungunya cases, respectively. The average duration of the illness was observed to be higher in chikungunya (median days (P25 to p75): 15 (7-45)) than dengue (median days (P25 to p75): 10 (5-15)). The median indirect cost in the case of dengue was Rs 1,931 (p25 to p75: 300 to 4500) while Rs 2,550 (p25 to p75: 0 to 5250) was observed for chikungunya cases. Two types of direct cost, namely, direct cost related to medical expenses and direct cost related to other expenses were calculated. Direct cost related to medical expenses was observed to be higher in dengue (Md (P25 to p75): Rs 2,450 (400-5000)) than chikungunya (Md (P25 to p75): Rs 1,500 (150-5200)) while indirect cost related to other expenses were comparable between dengue (Md (P25 to p75): Rs 1,575 (1300-2600)) and chikungunya (Md (P25 to p75): Rs 1500 (850-2850)). The average total cost for one dengue episode was estimated to be Rs 6,860 (3700-12525) whereas it was Rs 7,000 (2550-14000) for one episode of Chikungunya. Conclusions: Overall, patients have to bear high costs while suffering from dengue and chikungunya infections. Furthermore, the duration of illness while suffering from viral diseases also contributes to the substantial economic burden. Improved knowledge about the impact of the cost and the economic burden associated with dengue and chikungunya will help policymakers allocate and appropriate resources accordingly.

Four decades of epidemiological data reveal trajectories towards malaria elimination in Kheda district (Gujarat), western part of India.

BACKGROUND: Malaria is a main public health problem in India and was so particularly in the state of Gujarat in the western part of the country. This study assesses the effects of various interventions on malaria cases using data from the last 33 years (1987-2019). METHODS: Here we have analysed 33 years of malaria epidemiological data from a malaria clinic in Kheda district in Gujarat. The data were digitised yearly and monthly, age-wise and gender-wise, and descriptive analysis was performed to assess the effects of several interventions on malaria burden. RESULTS: During 1987-2019, our clinic diagnosed 5466 Plasmodium vivax and 4732 P. falciparum malaria cases. Overall, there was a declining trend in malaria cases except for the years 1991, 1994, 2004 and 2005. The year 2004 especially witnessed an epidemic in Kheda as well as throughout Gujarat. Malaria infections were most common (40%) among the 21-40 years age group. Fever was the most common symptom in all age groups. INTERPRETATION: Introduction of revised drug policy and improved surveillance technique (rapid diagnosis kits) have strengthened the diagnosis and treatment of malaria in the district. Use of pyrethroid in indoor residual insecticide spray has also strengthened vector control. Among the various interventions used, long-lasting insecticide nets and introduction of artemisinin-based combination therapy have played significant roles in controlling malaria cases. A more drastic decline in P. falciparum cases versus P. vivax is evident, but the latter persists in high proportions and therefore new tools for malaria control will be needed for elimination.

Small-scale field evaluation of the entomological efficacy and the residual activity of Fludora® Fusion WP-SB indoor residual spraying against Anopheles culicifacies s.l. in Gujarat, India.

OBJECTIVES: To evaluate the entomological efficacy and the residual activity of indoor residual spraying with Fludora® Fusion 562.5 WP-SB, a combination formulation containing clothianidin, a neonicotinoid and deltamethrin, a pyrethroid, against the main rural malaria vector, Anopheles culicifacies s.l., in India in a small-scale trial. METHODS: In three study villages, suitable households were randomly allocated to five treatments: Fludora® Fusion 562.5 WP-SB (target dose 225 mg active ingredient AI/m2 ); clothianidin 70 WG (target dose 200 mg AI/m2 ); K-Othrine 250 WG (deltamethrin, target dose 25 mg AI/m2 ); Ficam VC 80 WP-SB (bendiocarb, target dose 400 mg AI/m2 ) and unsprayed control. Insecticides were sprayed by hand compression sprayers with control flow valves and 8002E nozzles. Post-spray cone bioassays were done on insecticide-treated walls using a colonised, deltamethrin-resistant strain of An. culicifacies. Mosquitoes were collected from treated rooms by different methods. The insecticide content on filter papers collected from the sprayed walls was determined by chemical assay to assess the spray quality. RESULTS: The ratios of applied to target doses of insecticides were within 0.84 to 1.4, showing a good spray quality. The cone bioassays revealed residual action lasting 7 months for all insecticides without significant differences in mortality between different surfaces treated nor between the four treatment arms (P > 0.05). Considering all entomological parameters such as indoor resting density, excito-repellency, blood-feeding inhibition and delayed mortality, the overall efficacy of Fludora® Fusion WG-SB was equal or better compared with other insecticides. CONCLUSIONS: Fludora® Fusion showed overall equal or better efficacy than deltamethrin and bendiocarb alone against a pyrethroid-resistant malaria vector population and can be considered as an alternative product for management of pyrethroid resistance in malaria vectors.

Overlaying COVID-19 mitigation plans on malaria control infrastructures.

To counter the coronavirus disease 2019 (COVID-19) pandemic, each country must design sustainable control plans given the inherent disparities in wealth and healthcare systems. Most malaria-endemic countries run well-entrenched malaria control programs via their established frameworks for diagnosis, case management, treatment and overall surveillance. We propose that the malaria control infrastructures can be partially co-opted for launching sustainable COVID-19 mitigation plans.

In-silico Designing and Testing of Primers for Sanger Genome Sequencing of Dengue Virus Types of Asian Origin.

Rarity in reporting whole genome sequence of Dengue virus from dengue endemic countries leaves lacunae in understanding regional pattern of virus mutation and ultimately leading to non-understanding of transmission pattern and clinical outcomes emerging at regional levels. Due to inter-serotype genomic similarity and intra-serotype genomic diversity, appropriate designing of primer pairs appears as an exhaustive exercise. Present paper reports new Dengue virus type-specific primer which may help in characterizing virus specific to Asian origin. Genomes of dengue virus serotypes of Asian region were searched and using advanced bioinformatics tools, serotype specific primers were designed and tested for their targeted amplification efficiency. 19 primers sets for DENV-1, 18 primer sets for DENV-2, 17 for DENV-3 and 18 for DENV-4 were designed. In-silico and experimental testing of the designed primers were performed on virus isolated from both clinical isolates and passaged cultures. While all 17 and 18 primer sets of DENV-3 and DENV-2 respectively yielded good quality sequencing results; in case of DENV-4, 16 out of 18 primer sets and in DENV-1, 16 out of 19 primer sets yielded good results. Average sequencing read length was 382 bases and around 82% nucleotide bases were Phred quality QV20 bases (representing an accuracy of circa one miscall every 100 bases) or higher. Results also highlighted importance of use of primer development algorithm and identified genomic regions which are conservative, yet specific for developing primers to achieve efficiency and specificity during experiments.

Molecular, biochemical characterization and assessment of immunogenic potential of cofactor-independent phosphoglycerate mutase against Leishmania donovani: a step towards exploring novel vaccine candidate.

Despite immense efforts, vaccine against visceral leishmaniasis has yet not been developed. Earlier our proteomic study revealed a novel protein, cofactor-independent phoshoglycerate mutase (LdiPGAM), an important enzyme in glucose metabolism, in T helper cells type 1 (Th1) stimulatory region of soluble Leishmania donovani antigen. In this study, LdiPGAM was biochemically and molecularly characterized and evaluated for its immunogenicity and prophylactic efficacy against L. donovani. Immunogenicity of recombinant LdiPGAM (rLdiPGAM) was initially assessed in naïve hamsters immunized with it by analysing mRNA expression of inducible nitric oxide (NO) synthase (iNOS) and other Th1/T helper cells type 2 cytokines, which revealed an upregulation of Th1 cytokines along with iNOS. Immunogenicity of rLdiPGAM was further evaluated in lymphocytes of treated Leishmania-infected hamsters and peripheral blood mononuclear cells of Leishmania patients in clinical remission by various parameters, viz. lymphoproliferation assay and NO production (hamsters and patients) and levels of various cytokines (patients). rLdiPGAM induced remarkable Lymphoproliferative response and NO production in treated Leishmania-infected hamsters as well as in patients and increase in interferon gamma (IFN-γ), interleukin-12 (IL-12p40) responses in Leishmania patients in clinical remission. Vaccination with rLdiPGAM exerted considerable prophylactic efficacy (73%) supported by increase in mRNA expression of iNOS, IFN-γ and IL-12p40 with decrease in transforming growth factor beta and interleukin-10. Above results indicate the importance of rLdiPGAM protein as a potential vaccine candidate against visceral leishmaniasis.

Characterization of the proliferating cell nuclear antigen of Leishmania donovani clinical isolates and its association with antimony resistance.

Previously, through a proteomic analysis, proliferating cell nuclear antigen (PCNA) was found to be overexpressed in the sodium antimony gluconate (SAG)-resistant clinical isolate compared to that in the SAG-sensitive clinical isolate of Leishmania donovani. The present study was designed to explore the potential role of the PCNA protein in SAG resistance in L. donovani. For this purpose, the protein was cloned, overexpressed, purified, and modeled. Western blot (WB) and real-time PCR (RT-PCR) analyses confirmed that PCNA was overexpressed by ≥ 3-fold in the log phase, stationary phase, and peanut agglutinin isolated procyclic and metacyclic stages of the promastigote form and by ~5-fold in the amastigote form of the SAG-resistant isolate compared to that in the SAG-sensitive isolate. L. donovani PCNA (LdPCNA) was overexpressed as a green fluorescent protein (GFP) fusion protein in a SAG-sensitive clinical isolate of L. donovani, and modulation of the sensitivities of the transfectants to pentavalent antimonial (Sb(V)) and trivalent antimonial (Sb(III)) drugs was assessed in vitro against promastigotes and intracellular (J774A.1 cell line) amastigotes, respectively. Overexpression of LdPCNA in the SAG-sensitive isolate resulted in an increase in the 50% inhibitory concentrations (IC50) of Sb(V) (from 41.2 ± 0.6 µg/ml to 66.5 ± 3.9 µg/ml) and Sb(III) (from 24.0 ± 0.3 µg/ml to 43.4 ± 1.8 µg/ml). Moreover, PCNA-overexpressing promastigote transfectants exhibited less DNA fragmentation compared to that of wild-type SAG-sensitive parasites upon Sb(III) treatment. In addition, SAG-induced nitric oxide (NO) production was found to be significantly inhibited in the macrophages infected with the transfectants compared with that in wild-type SAG-sensitive parasites. Consequently, we infer that LdPCNA has a significant role in SAG resistance in L. donovani clinical isolates, which warrants detailed investigations regarding its mechanism.