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.

Case Report: A Series of Three Meningoencephalitis Cases Caused by Acanthamoeba spp. from Eastern India.

Acanthamoeba spp. are rare etiological agents of meningoencephalitis with high mortality. We present three cases of Acanthamoeba meningoencephalitis in immunocompetent individuals from Eastern India. The first patient presented with fever and headache; the second with headache, visual disturbance, and squint; and the third presented in a drowsy state. The cases presented on March 3, 18, and 21, 2023 respectively. The first two patients had concomitant tubercular meningitis for which they received antitubercular therapy and steroid. Their cerebrospinal fluid showed slight lymphocytic pleocytosis and increased protein. The diagnosis was done by microscopy, culture, and polymerase chain reaction. They received a combination therapy comprising rifampicin, fluconazole, and trimethoprim-sulfamethoxazole. The first patient additionally received miltefosine. She responded well to therapy and survived, but the other two patients died despite intensive care. Detection of three cases within a period of 1 month from Eastern India is unusual. It is imperative to sensitize healthcare providers about Acanthamoeba meningoencephalitis to facilitate timely diagnosis and treatment of the disease.

Prevalence of histidine-rich protein 2 deletion among the Plasmodium falciparum isolates from Kolkata.

Context: Histidine-rich protein 2 (HRP2) detecting rapid diagnostic tests (RDTs) have played an important role in enabling prompt malaria diagnosis in remote locations. HRP2 has advantages over other biomarkers because of its abundance in the bloodstream, repetitive binding epitopes, and falciparum-specificity. Most HRP2-based RDTs also exhibit some cross-reactivity to a closely related protein (HRP3). Plasmodium falciparum parasites lacking HRP2 (pfhrp2) and 3 (pfhrp3) genes escape detection by these RDTs. Objectives: The objective of the study was to study the sensitivity and specificity of hrp2-based RDT for diagnosis of falciparum, to compare the RDT results with microscopy and polymerase chain reaction (PCR), and to determine the prevalence of HRP2 gene deletion among the RDT-negative, microscopy-positive falciparum strains. Materials and Methods: Blood samples were collected and diagnosis was done by microscopic examination, RDTs, and PCR. Results: Out of 1000 patients examined, 138 were positive for P. falciparum. Fever was the most common symptom followed by chills with rigor and headache were recorded among more than >95% of the study patients. Three microscopy-confirmed P. falciparum cases were negative by HRP2-based RDT and were found to have deletion of HRP2 and HRP3 exon 2. Conclusions: Rapid and accurate diagnosis and prompt deployment of effective antimalarial medication are essential components of appropriate case management. P. falciparum strains that evade diagnosis by RDTs represent a major threat to malaria control and elimination efforts.

Survival of Persons Living With HIV/AIDS: A Multicentric Study From India.

BACKGROUND: It has been more than 17 years since the introduction of free ART in India. At this point, it would be prudent to look at the factors associated with the survival of persons living with human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) (PLHA) who are already enrolled in the ART program. METHODS: PLHAs enrolled from antiretroviral therapy (ART) centers located in three different cities in India - Delhi, Pune and Kolkata, and were followed up at six monthly intervals monitoring the WHO stage, CD4 counts, complete blood counts, and liver and kidney function tests, for a duration of three years. RESULTS AND DISCUSSION: The incidence of mortality among HIV/AIDS patients on ART was 5.0 per 1000 patient-years (21/1410, 1.4%). Age at initiation of ART, being above 35 years, was the only significant predictor of mortality (log-rank p = 0.018). Multivariable analysis showed a significant association of an unfavourable outcome (defined as mortality or development of opportunistic infection during follow-up) with male gender (adjusted odds ratio (AOR) = 5.26, p = <0.01) and being unmarried at ART initiation (AOR = 1.39, p = 0.005). CONCLUSION: The survival of PLHA with good adherence to ART is independent of the WHO stage or CD4 counts at the initiation of ART. Initiation of ART after 35 years of age was a significant predictor of mortality.

Efficacy of Bacillus Calmette-Guérin (BCG) Vaccination in Reducing the Incidence and Severity of COVID-19 in High-Risk Population (BRIC): a Phase III, Multi-centre, Quadruple-Blind Randomised Control Trial.

INTRODUCTION: Universal coverage of vaccines alone cannot be relied upon to protect at-risk populations in lower- and middle-income countries against the impact of the coronavirus disease 2019 (COVID-19) pandemic and newer variants. Live vaccines, including Bacillus Calmette-Guérin (BCG), are being studied for their effectiveness in reducing the incidence and severity of COVID-19 infection. METHODS: In this multi-centre quadruple-blind, parallel assignment randomised control trial, 495 high-risk group adults (aged 18-60 years) were randomised into BCG and placebo arms and followed up for 9 months from the date of vaccination. The primary outcome was the difference in the incidence of COVID-19 infection at the end of 9 months. Secondary outcomes included the difference in the incidence of severe COVID-19 infections, hospitalisation rates, intensive care unit stay, oxygen requirement and mortality at the end of 9 months. The primary analysis was done on an intention-to-treat basis, while safety analysis was done per protocol. RESULTS: There was no significant difference in the incidence rates of cartridge-based nucleic acid amplification test (CB-NAAT) positive COVID-19 infection [odds ratio (OR) 1.08, 95% confidence interval (CI) 0.54-2.14] in the two groups, but the BCG arm showed a statistically significant decrease in clinically diagnosed (symptomatic) probable COVID-19 infections (OR 0.38, 95% CI 0.20-0.72). Compared with the BCG arm, significantly more patients developed severe COVID-19 pneumonia (CB-NAAT positive) and required hospitalisation and oxygen in the placebo arm (six versus none; p = 0.03). One patient belonging to the placebo arm required intensive care unit (ICU) stay and died. BCG had a protective efficacy of 62% (95% CI 28-80%) for likely symptomatic COVID-19 infection. CONCLUSIONS: BCG is protective in reducing the incidence of acute respiratory illness (probable symptomatic COVID-19 infection) and severity of the disease, including hospitalisation, in patients belonging to the high-risk group of COVID-19 infection, and the antibody response persists for quite a long time. A multi-centre study with a larger sample size will help to confirm the findings in this study. CLINICAL TRIALS REGISTRY: Clinical Trials Registry India (CTRI/2020/07/026668).

The Bacillus Calmette­Guérin (BCG) vaccine has been studied previously in several settings, including reducing childhood mortalities due to viral infections and induction of trained immunity and reducing upper respiratory tract infections and pneumonia in older adults. This multi-centre trial has tried to evaluate the efficacy of BCG revaccination in reducing the incidence and severity of COVID-19 infections in adults between 18 and 60 years of age belonging to the high-risk group owing to the presence of comorbidities including diabetes, chronic kidney disease, chronic liver disease and chronic lung diseases. A single dose of BCG vaccine produced significantly high titres of BCG antibodies lasting for six months. While there was no significant reduction in the incidence of COVID-19 infection, there was an 8.4% reduction in the incidence of symptomatic COVID-19 disease at the end of 9 months of follow-up. In addition, there were significantly fewer severe COVID-19 infections requiring hospital stay and oxygen support. However, the overall numbers of severe COVID-19 infections were low. Thus, the study shows that BCG can protect against symptomatic and severe COVID-19 disease. However, it might not reduce the incidence of new infections. The study results are significant for low- and middle-income countries without adequate coverage of primary doses of COVID-19 vaccination, let alone the booster doses. Future studies should evaluate the BCG vaccine's efficacy as a booster compared with routine COVID-19 vaccine boosters.

Outcome of Adult Malarial Co-infections in Eastern India.

Introduction: Co-infection with different agents such as bacterial, viral, and Rickettsia is being increasingly recognized due to greater availability and utilization of the diagnostic tests among malaria patients. Methods: Consecutive admitted malarial cases were included and were subjected to test for general investigations, bacteria, typhoid, dengue, chikungunya, and rest for specific diagnosis. All patients were followed up till discharge or death and appropriate statistical tests were performed. Results: A total of 152 malaria patients were recruited and 27 (18.8%) had concurrent infections. It included 40.7% dengue only, 18.7% pneumonia, 11.1% urinary tract infection (UTI), 7.4% enteric fever, 3.7% leptospirosis, chikungunya, and tuberculous meningitis each, and 3.7% each of dengue with pneumonia and UTI. The organisms isolated were Streptococcus pneumoniae, Klebsiella pneumoniae, Escherichia coli, Salmonella typhi, and Mycobacterium tuberculosis. The mean duration of fever was 6.33 ± 3.63 days with a range of 3-20 days. Blood culture grew in 2 cases S. typhi and K. pneumonia,e. Dengue co-infections had significantly higher clinical and laboratory features of dengue and complications such as bleeding, jaundice, and cholecystitis, whereas rest concurrent infections had a significantly higher proportion of nausea and vomiting, convulsion, altered sensorium, productive cough, urinary symptoms, shock, acute kidney injury, anemia, and mean neutrophil count. There was significantly higher mortality among malaria-dengue concurrent infection group with 2 (15.4%) than malaria mono-infection group 3 (2.4%). Conclusion: Co-infections with malaria are not uncommon, especially dengue fever and other bacterial infections. The dominant clinical picture is of the superimposed infection. Decision should be clinically guided adjunct with specific diagnostic tests, and timely treatment has favorable outcome.

A report on incidence of COVID-19 among febrile patients attending a malaria clinic.

CONTEXT: Screening for malaria and coronavirus disease (COVID-19) in all patients with acute febrile illness is necessary in malaria-endemic areas to reduce malaria-related mortality and to prevent the transmission of COVID-19 by isolation. AIMS: A pilot study was undertaken to determine the incidence of SARS-CoV-2 infection among febrile patients attending a malaria clinic. SUBJECTS AND METHODS: All patients were tested for malaria parasite by examining thick and thin blood smears as well as by rapid malaria antigen tests. COVID-19 was detected by rapid antigen test and reverse transcriptase-polymerase chain reaction in patients agreeing to undergo the test. RESULTS: Out of 262 patients examined, 66 (25.19%) were positive for Plasmodium vivax, 45 (17.17%) for Plasmodium falciparum (Pf) with a slide positivity rate of 42.40%, and Pf% of 40.50%. Only 29 patients consented for COVID-19 testing along with malaria; of them, 3 (10.34%) were positive for COVID-19 alone and 2 (6.89%) were positive for both COVID-19 and P. vivax with an incidence of 17.24%. A maximum number of patients (196) did not examine for COVID-19 as they did not agree to do the test. CONCLUSION: Diagnosis of COVID-19 among three patients (10.34%) is significant both in terms of identification of cases and to isolate them for preventing transmission in the community. Detection of COVID-19 along with malaria is equally important for their proper management.

Entomophthoramycosis: An unusual cause of facial disfigurement.

Entomophthoramycosis is a rare fungal infection of nose, paranasal sinuses and subcutaneous tissues found in tropical and subtropical region. From India very few cases have been reported. Here we report a case of Entomophthoramycosis due to Conidiobolus coronatus from the eastern India who presented with slowly growing rhinofacial swelling and right sided nasal obstruction due to intranasal mass. The case was diagnosed by typical histopathological findings of broad aseptate hyphae with surrounding eosinophilic granular material (Splendore Hoeppli phenomenon) on microscopy of nasal biopsy material and confirmed by PCR assay of DNA and sequencing from biopsy tissue. Treatment with saturated solution of potassium iodide and itraconazole was successful and clinical cure was attained in 8 months.

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.

Pulmonary tuberculosis and pneumocystis jirovecii concurrent pneumonia in HIV infected patients at a resource limited setting in Eastern India: A case series.

Pneumocytis jirovecii pneumonia (PJP) and Pulmonary TB (PTB) both are common opportunistic infections among HIV infected individuals. But concurrent infections pose a diagnostic challenge owing to similar clinical features. Data suggests a high prevalence of such concurrent infections in developing countries but limited diagnostic modalities especially in resource constraint setup limits accurate diagnosis. At our centre we came across 6 newly diagnosed PTB patients among HIV infected ones had persistent shortness of breath (SOB) and hypoxia despite starting anti-tuberculous treatment (ATT). We excluded concomitant bacterial pneumonia by imaging, sputum examination and blood culture. Serum lactate dehydrogenase (LDH) was estimated and hypoxia by arterial blood gas (ABG). We found all 6 patients had elevated serum LDH, hypoxia and imaging suggestive of PJP were offered sputum for Geisma stain and standard treatment for PJP in form of Bactrim-double strength and steroid. 1 patient had PJ cysts in sputum. 5 patient's classical radiologic findings in form of ground glass opacities in lower lobes along with bilateral infiltrates and 1 had honeycombing. Serum LDH was elevated all 6 subjects. 5 were newly diagnosed HIV and 4 had CD4 count below 50 cells/mm3 and 2 had below 200 cells/mm3.1 patient had developed bilateral pneumothorax as complication. 4 patients responded to treatment and 2 (33.3%) died of respiratory failure during treatment. We were able to diagnose only severe PJP cases as concurrent infection with PTB as there was no availability of broncho alveolar lavage (BAL) as well as direct fluorescent antigen (DFA) test for PJ detection. A high index of suspicion for PJP even in PTB patients with low CD4 count will guide to appropriate therapy for both infections and eventually reduces morbidity and mortality.

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.

Characterization of human immunodeficiency virus-infected patients of suspected first-line antiretroviral treatment failure within 5 years - Evidence from a tertiary hospital, Kolkata.

OBJECTIVES: Antiretroviral therapy (ART) has immense survival benefit on human immunodeficiency virus (HIV)-infected people. However, every year, a proportion of patients were failing to the first-line drugs. The aim of this study is to characterize the patients developing first-line failure within 5 years of ART. MATERIALS AND METHODS: A retrospective observational study was carried out at the Centre of Excellence in HIV care, School of Tropical Medicine, Kolkata. A total of 190 referred patients' data of suspected first-line treatment failure who failed first-line ART within 5 years of initiation were collected and analyzed using R software. RESULTS: Among 190 patients, 100 (52.4%) patients had virologic failure. Male patients 78 (41.05%) outnumbered females 22 (11.57%) and needed to switch to the second-line drugs. The median age was 37 years (range 8-65 years), and the median duration of first-line ART taken was 2.85 years. Among the first-line failed patients, zidovudine, lamivudine, and nevirapine (23.6%) was the most common antiretroviral regimen and 77 (40.5%) referred in the WHO stage I of illness. Seventy-three (38.42%) patients were referred for immunological failure, 26 (13.7%) for both immunological and clinical failure, and only 1 (0.52%) had only clinical failure at the time of referral. We found a significant association of suboptimal adherence (P < 0.05) and high viral load in this study. CONCLUSION: This study enables that poor adherence was the most important factor responsible for the first-line treatment failure. As adherence is a dynamic process, interventions in every visit following ART initiation should be optimized, and a multidisciplinary approach toward adherence is needed to get the highest treatment outcome benefit.

Lamivudine plus tenofovir versus lamivudine plus adefovir for the treatment of hepatitis B virus in HIV-coinfected patients, starting antiretroviral therapy.

Background: Combination of tenofovir disoproxil fumarate (TDF), lamivudine (3TC) and efavirenz (EFV) is preferred in the treatment of HIV/hepatitis B virus (HBV) coinfection. We postulated that a HBV active nucleoside reverse transcriptase (RT) inhibitor/nucleotide RT inhibitor backbone of adefovir dipivoxil (ADV) +3TC would be as effective as TDF +3TC for the Indian population. Objective: ADV + 3TC could be an alternative option for these HIV/HBV coinfected individuals, preserving the dually active TDF + 3TC as second-line nucleoside backbone following failure of the first-line ART. Materials and Methods: This randomised control trial (CTRI/2012/03/002471) was carried out at the ART Centre of Calcutta School of Tropical Medicine, India. Seventy-eight (39 on each arm) treatment-naïve HIV/HBV coinfected patients were randomised to receive either the combination of lamivudine + tenofovir + EFV or lamivudine + adefovir + zidovudine + EFV and followed up for 120 weeks. Results: Median age of the study participants was 36 years (21-62), majority were male (61/78; 78.2%) and heterosexually (39/78; 50%) infected. Baseline characteristics were identical in both arms. There was no statistically significant difference in median aspartate aminotransferase (37 vs. 29.5 U/L), alanine aminotransferase (ALT) (36 vs. 34.5 U/L), ALT normalisation rate (80 vs. 70%), AST to platelet ratio index (0.45 vs. 0.33), CD4 count (508 vs. 427 cells/mm3), HBV DNA suppression (81.8 vs. 70%), hepatitis B e antigen loss (9 vs. 5%), hepatitis B surface antigen seroclearance rate (6.06 vs. 18.75%) and death (3 vs. 3) at 120 weeks between TDF (n = 33) and ADV (n = 32), respectively. Conclusions: Adefovir plus lamivudine is an effective alternative of tenofovir plus lamivudine in long-term HBV treatment outcome in HIV/HBV coinfected patients.

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.

Occult hepatitis B virus infection in HIV positive patients at a tertiary healthcare unit in eastern India.

Occult HBV infection (OBI), defined by the presence of HBV DNA in absence of hepatitis B surface antigen (HBsAg), is a significant concern in the HIV-infected population. Of 441 HIV+/HBsAg- patients analyzed, the overall prevalence of OBI was 6.3% (28/441). OBI was identified in 21 anti-HBc positives (17.8%), as well as among those who lacked any HBV-specific serological markers (2.2%). Comparison with HIV/HBV co-infection revealed that the levels of CD4, ALT, and HBV DNA were significantly lower during occult infection. Discrete differences were also observed with respect to quasispecies divergence. Additionally, subgenotype D1 was most frequent in occult infection, while D2 was widespread during chronic infection. The majority (~90%) of occult D1 sequences had the sQ129R mutation in the surface gene. This study highlights several distinct features of OBI in India and underscores the need for additional HBV DNA screening in HIV-positive individuals.

Baseline characteristics of HIV & hepatitis B virus (HIV/HBV) co-infected patients from Kolkata, India.

BACKGROUND & OBJECTIVES: Hepatitis B virus (HBV) and HIV co-infection has variable prevalence worldwide. In comparison to HBV mono-infection, the course of chronic HBV infection is accelerated in HIV/HBV co-infected patients. the present study was carried out to analyse the baseline characteristics (clinical, biochemical, serological and virological) of treatment naïve HIV/HBV co-infected and HIV mono-infected patients. METHODS: Between July 2011 and January 2013, a total number of 1331 HIV-seropositive treatment naïve individuals, enrolled in the ART Centre of Calcutta School of Tropical Medicine, Kolkata, India, were screened for hepatitis B surface antigen (HBsAg). A total of 1253 HIV mono-infected and 78 HIV/HBV co-infected patients were characterized. The co-infected patients were evaluated for HBeAg and anti-HBe antibody by ELISA. HIV RNA was quantified for all co-infected patients. HBV DNA was detected and quantified by real time-PCR amplification followed by HBV genotype determination. RESULTS: HIV/HBV co-infected patients had proportionately more advanced HIV disease (WHO clinical stage 3 and 4) than HIV mono-infected individuals (37.1 vs. 19.9%). The co-infected patients had significantly higher serum bilirubin, alanine aminotransferase (ALT), alkaline phosphatase and ALT/platelet ratio index (APRI). CD4 count was non-significantly lower in co-infected patients. Majority (61.5%) were HBeAg positive with higher HIV RNA (P<0.05), HBV DNA (p<0.001) and APRI (p<0.05) compared to those who were HBeAg negative. HBV/D was the predominant genotype (73.2%) and D2 (43.7%) was the commonest subgenotype. INTERPRETATION & CONCLUSIONS: HIV/HBV co-infected patients had significantly higher serum bilirubin, ALT, alkaline phosphatase and lower platelet count. HBeAg positive co-infected patients had higher HIV RNA and HBV DNA compared to HBeAg negative co-infected patients. Prior to initiation of antiretroviral treatment (ART) all patients should be screened for HBsAg to initiate appropriate ART regimen.

Polymorphisms in Pfcrt and Pfmdr-1 genes after five years withdrawal of chloroquine for the treatment of Plasmodium falciparum malaria in West Bengal, India.

BACKGROUND: The emergence of resistant power against different antimalarial agents particularly by Plasmodium falciparum is a challenge to combat malaria. Regular monitoring is essential not only to determine the efficacy and development of resistance by the parasite but also to detect early sign of regaining sensitivity to any anti-malarial agent that has been withdrawn for a long period. Studies on molecular markers associated with antimalarial drug resistance of prevailing Plasmodium population play an important role in this aspect. The present protocol was designed to study the polymorphisms in pfcrt and pfmdr-1 gene to determine any sign of regaining sensitivity to chloroquine among P. falciparum after five years of artemisinin combination therapy (ACT) implementation. METHODS: Clinical isolates were collected from P. falciparum positive patients attending the malaria clinic of Calcutta School of Tropical Medicine during December 2014 to December 2015. Genomic parasitic DNA was extracted and subjected to sequencing of pfcrt and pfmdr-1 gene directly from purified PCR products. RESULTS: A total of 89 isolates were sequenced for pfcrt and 73 isolates for pfmdr-1 genes. In pfcrt gene mutant K76T was detected in all isolates and all were SVMNT haplotype. Out of three important polymorphisms in pfmdr-1 gene mutant Y184F was detected among all isolates. One synonymous G182G and one non-synonymous S232F/Y, mutation were detected in 99% isolates. CONCLUSION: All isolates carrying mutant K76T in pfcrt gene, considered as hall mark for CQ resistance, indicate that there is no sign of regaining CQ sensitivity among the prevailing P. falciparum population of the study area after five years of ACT implementation.

Comparative Study of Effectiveness and Resistance Profile of Chloroquine and Sulfadoxine-Pyrimethamine in Uncomplicated Plasmodium falciparum Malaria in Kolkata.

INTRODUCTION: Malaria is one of the major public health problems of the country. Factors responsible for reemergence of malaria in India was due to emergence and spread of chloroquine resistant Plasmodium falciparum strains across the country coupled with steady rise in insecticide resistance of the vector mosquitoes. Very little is known about the drug resistance status of P. falciparum in India. As per National Vector Borne Diseases Control Programme (NVBDCP), chloroquine is the drug of choice for uncomplicated P. falciparum cases and the combination of Artesunate and Sulfadoxine-Pyrimethamine (SP) is being used to treat the documented chloroquine-resistant uncomplicated cases. To evaluate the comparative effectiveness and resistance profile of Chloroquine vis-à-vis Sulfadoxine-Pyrimethamine (SP) in uncomplicated Plasmodium falciparum cases as the first-line therapy a study was undertaken at the Malaria Clinic of Calcutta School of Tropical Medicine, Kolkata during the period from July 2007 to December 2007 at Kolkata Municipal Corporation, Kolkata. MATERIAL & METHODS: Following WHO protocol 2003, a total of 100 parasitologically confirmed Plasmodium falciparum cases were recruited as per the recruitment criteria. Among them, 50 patients were given Chloroquine and another 50 patients were given SP. Eight patients were excluded or lost to follow-up during the follow-up period because of failure to follow the protocol. RESULTS: It was observed that in the Chloroquine group out of 50 patients, 30 (60%) showed adequate clinical and parasitological response (ACPR), 15 (30%) had late treatment failure (LTF) and remaining 5 (10%) were lost during the follow up period (LFU). On the other hand in the SP group out of 50 patients, 46 (92%) showed ACPR and only one (2%) had LTF and 3 patients were LFU. The difference of LTF in Chloroquine and Sulfadoxine-pyrimethamine groups was statistically significant (p value < 0.05). Also there was statistically significant difference of the mean parasite clearance time (PCT) of Chloroquine (82.7 hours) and SP group (61.3 hours). CONCLUSIONS: Chloroquine failure rate was high which was well above the WHO recommended cut off threshold for drug policy change (> 10%), Sulfadoxine- Pyrimethamine can be used in place of Chloroquine as the first line drug in uncomplicated P. falciparum cases.