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Introduction: The ongoing visceral leishmaniasis (VL) elimination programme in India is targeting the elimination of the disease VL but not the pathogen. The persistence of hidden parasite pool may initiate a resurgence in suitable conditions. This study dealt with a novel approach to unearth such pathogen pool and their proper management to prevent the resurgence of VL., Materials and Methods: We deployed a new approach for detection of pathogen pool by following up the VL and post kala-azar dermal leishmaniasis patients treated during the last 10 years along with mass sero-surveillance within a radius of 500 m of recently treated individuals., Results: We followed up 72.6% (3026/4168) previously treated VL and post kala-azar dermal leishmaniasis patients and diagnosed 42 (1.4%) new and 38 (1.3%) recurrent post kala-azar dermal leishmaniasis. We detected 93 asymptomatic leishmanial infection, 8 VL and 1 post kala-azar dermal leishmaniasis by mass sero-surveillance., Conclusion: Our three-step process including mapping and follow-up of previously treated cases, mass surveillance within 500 m of radius of known cases, and 6 monthly follow-on clinical and serological screening of asymptomatic cases, enabled detection of previously undetected cases of post kala-azar dermal leishmaniasis and VL. Recurrent post kala-azar dermal leishmaniasis deserves special attention regarding their treatment guideline. Early diagnosis and effective treatment of all leishmaniasis cases will hasten pathogen elimination and prevent resurgence of VL. This may help the policymakers to develop appropriate strategy for elimination of pathogen to prevent resurgence of VL., (© 2024. The Author(s).)

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., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Guha et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

A comprehensive entomological survey was undertaken in Alipurduar District, West Bengal, from 2018 to 2020 and in 2022. This study was prompted by reported malaria cases and conducted across nine villages, seven Sub-Centres, and three Primary Health Centres (PHCs). Mosquitoes were hand-collected with aspirators and flashlights from human dwellings and cattle sheds during the daytime. Both morphological and molecular techniques were used for species identification. Additionally, mosquitoes were tested for Plasmodium parasites and human blood presence. Mosquito species such as An. barbirostris s.l. , An. hyrcanus s.l. , An. splendidus , and An. vagus were morphologically identified. For species like An. annularis s.l. , An. minimus s.s. , An. culicifacies s.l. , and An. maculatus s.s ., a combination of morphological and molecular techniques was essential. The mitochondrial cytochrome c oxidase gene subunit 1 (CO1) was sequenced for An. annularis s.l. , An. maculatus s.s. , An. culicifacies s.l. , An. vagus , and some damaged samples, revealing the presence of An. pseudowillmori and An. fluviatilis . The major Anopheles species were An. annularis s.l ., An. culicifacies s.l. , and An. maculatus s.s ., especially in Kumargram and Turturi PHCs. Plasmodium positivity was notably high in An. annularis s.l. and An. maculatus s.s. with significant human blood meal positivity across most species. Morphological, molecular, and phylogenetic analyses are crucial, especially for archived samples, to accurately identify the mosquito fauna of a region. Notably, this study confirms the first occurrence of An. pseudowillmori and An. sawadwongporni in West Bengal and implicates An. maculatus s.s ., An. culicifacies s.l. , and An. annularis s.l. as significant vectors in the Alipurduar region.

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 ., Competing Interests: There are no conflicts of interest., (Copyright: © 2024 Tropical Parasitology.)

Background: Japanese encephalitis (JE) is an emerging zoonotic disease caused by JE virus (JEV) and transmitted to humans from pigs or aquatic birds by vector mosquitoes in southeast Asian countries. In this study, JEV infection rate among vector mosquitoes and domestic pigs was determined by detecting viral RNA and anti-JEV antibody (immunoglobulin G), respectively., Materials and Methods: A total of 146 pool mosquitoes of Culexvishnui subgroup and 278 pig blood samples were analyzed by reverse transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay methods, respectively. E and premembrane (PrM) gene of JEV detected among vectors were sequenced and a phylogenetic tree was constructed., Results: Five (5.81%) pools of Culextritaeniorhynchus were positive for JEV with pooled infection rate 1.70/1000 mosquitoes. A total of 108 (38.84%) blood samples were positive for anti-JEV antibody. Phylogenetic analysis revealed that our own E and PrM gene sequence of JEV belonging to Genotype III and showed 96.95% sequence similarities with the vaccine strain SA14-14-2., Conclusion: It was observed that domestic pigs of northern West Bengal were highly infected with JEV. Hence, the transmission should be blocked by pig vaccination. A pilot study may be undertaken for mass vaccination of the prevailing pig population to observe any reduced rate of JEV transmission from both pig to pig and pig to human., (Copyright © 2023 Copyright: © 2023 Indian Journal of Public Health.)

Background & Objectives: Community participation is one of the key factors for implementation and success of a public health programme which depends upon knowledge about that disease. Therefore, understanding the community knowledge about malaria is important for designing sustainable control programmes. This study was conducted to assess the knowledge about malaria, to evaluate long lasting insecticidal nets (LLINs) distribution and their use by LQAS method in endemic areas of Bankura district, West Bengal state, India Methods: It was a community based cross-sectional survey conducted in Bankura during December 2019-March 2020. Structured questionnaire under four categories: socio-demographic variables, knowledge of malaria, owner ship of LLINs and its use were used for the interview. Ownership of LLINs and its use were analysed by LQAS method. Data were analysed by binary logistic regression model and chi-squared test., Results: Out of 456 respondents, 88.59% had good knowledge, 97.37% had good ownership of LLIN and 78.95% used LLINs properly. The knowledge about malaria was significantly associated with education level (p-value<0.0001). Out of 24 lots studied, 3, 2, 4 lots were underperforming with respect to knowledge, ownership of LLIN and its use, respectively., Interpretation & Conclusion: The study population had a good knowledge about malaria. In spite of good coverage of LLIN distribution, the use of LLINs was not up to the mark. LQAS analysis showed underperformance in few lots about knowledge, ownership of LLIN and its use. The IEC and BCC activities about LLIN should be done to achieve the impact of this intervention at the community level.

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., Competing Interests: There are no conflicts of interest., (Copyright: © 2023 Tropical Parasitology.)

Background: One major challenge in binning Metagenomics data is the limited availability of reference datasets, as only 1% of the total microbial population is yet cultured. This has given rise to the efficacy of unsupervised methods for binning in the absence of any reference datasets., Objective: To develop a deep clustering-based binning approach for Metagenomics data and to evaluate results with suitable measures., Methods: In this study, a deep learning-based approach has been taken for binning the Metagenomics data. The results are validated on different datasets by considering features such as Tetra-nucleotide frequency (TNF), Hexa-nucleotide frequency (HNF) and GC-Content. Convolutional Autoencoder is used for feature extraction and for binning; the K-means clustering method is used., Results: In most cases, it has been found that evaluation parameters such as the Silhouette index and Rand index are more than 0.5 and 0.8, respectively, which indicates that the proposed approach is giving satisfactory results. The performance of the developed approach is compared with current methods and tools using benchmarked low complexity simulated and real metagenomic datasets. It is found better for unsupervised and at par with semi-supervised methods., Conclusion: An unsupervised advanced learning-based approach for binning has been proposed, and the developed method shows promising results for various datasets. This is a novel approach for solving the lack of reference data problem of binning in metagenomics., (© 2022 Bentham Science Publishers.)

The application of computer vision in agriculture has already contributed immensely to restructuring the existing field practices starting from the sowing to the harvesting. Among the different plant parts, the economic part, the yield, has the highest importance and becomes the ultimate goal for the farming community. It depends on many genetic and environmental factors, so this curiosity about knowing the yield brought several precise pre-harvest prediction methods using different ways. Out of those techniques, non-invasive yield prediction techniques using computer vision have been proved to be the most efficient and trusted platform. This study developed a novel methodology, called SlypNet, using advanced deep learning networks, i.e., Mask R-CNN and U-Net, which can extract various plant morphological features like spike and spikelet from the visual image of the wheat plant and provide a high-throughput yield estimate with great precision. Mask R-CNN outperformed previous networks in spike detection by its precise detection performance with a mean average precision (mAP) of 97.57%, a F1 score of 0.67, and an MCC of 0.91 by overcoming several natural field constraints like overlapping and background interference, variable resolution, and high bushiness of plants. The spikelet detection module's accuracy and consistency were tested with about 99% validation accuracy of the model and the least error, i.e., a mean square error of 1.3 from a set of typical and complex views of wheat spikes. Spikelet yield cumulatively showed the probable production capability of each plant. Our method presents an integrated deep learning platform of spikelet-based yield prediction comprising spike and spikelet detection, leading to higher precision over the existing methods., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Maji, Marwaha, Kumar, Arora, Chinnusamy and Islam.)

Hippocampus is a part of the limbic system in human brain that plays an important role in forming memories and dealing with intellectual abilities. In most of the neurological disorders related to dementia, such as, Alzheimer's disease, hippocampus is one of the earliest affected regions. Because there are no effective dementia drugs, an ambient assisted living approach may help to prevent or slow the progression of dementia. By segmenting and analyzing the size/shape of hippocampus, it may be possible to classify the early dementia stages. Because of complex structure, traditional image segmentation techniques can't segment hippocampus accurately. Machine learning (ML) is a well known tool in medical image processing that can predict and deliver the outcomes accurately by learning from it's previous results. Convolutional Neural Networks (CNN) is one of the most popular ML algorithms. In this work, a U-Net Convolutional Network based approach is used for hippocampus segmentation from 2D brain images. It is observed that, the original U-Net architecture can segment hippocampus with an average performance rate of 93.6%, which outperforms all other discussed state-of-arts. By using a filter size of [Formula: see text], the original U-Net architecture performs a sequence of convolutional processes. We tweaked the architecture further to extract more relevant features by replacing all [Formula: see text] kernels with three alternative kernels of sizes [Formula: see text], [Formula: see text], and [Formula: see text]. It is observed that, the modified architecture achieved an average performance rate of 96.5%, which outperforms the original U-Net model convincingly., (© 2022. The Author(s) under exclusive licence to Society for Imaging Informatics in Medicine.)

Usage of effective classification techniques on Magnetic Resonance Imaging (MRI) helps in the proper diagnosis of brain tumors. Previous studies have focused on the classification of normal (nontumorous) or abnormal (tumorous) brain MRIs using methods such as Support Vector Machine (SVM) and AlexNet. In this paper, deep learning architectures are used to classify brain MRI images into normal or abnormal. Gender and age are added as higher attributes for more accurate and meaningful classification. A deep learning Convolutional Neural Network (CNN)-based technique and a Deep Neural Network (DNN) are also proposed for effective classification. Other deep learning architectures such as LeNet, AlexNet, ResNet, and traditional approaches such as SVM are also implemented to analyze and compare the results. Age and gender biases are found to be more useful and play a key role in classification, and they can be considered essential factors in brain tumor analysis. It is also worth noting that, in most circumstances, the proposed technique outperforms both existing SVM and AlexNet. The overall accuracy obtained is 88% (LeNet Inspired Model) and 80% (CNN-DNN) compared to SVM (82%) and AlexNet (64%), with best accuracy of 100%, 92%, 92%, and 81%, respectively.

Kala-azar or visceral leishmaniasis was at one time a scourge in the Bengal Presidency of British India comprising the present Indian states of Bengal, Bihar, Assam, and Odisha. The disease was rampant along the Ganga and Brahmaputra River adjoining areas. In the early 1900s, the treatment initiated was by the intravenous injection of tartar emetic, which had a narrow safety level and long-term use was marked with multiple side effects. In 1920, Upendranath Brahmachari discovered urea stibamine, which is the urea salt of para-amino phenyl stibnic acid and it revolutionized the treatment of Kala-azar with >90% cure rate and with minimal side effects. He is also credited with the description of post-kala-azar dermal leishmaniasis. He was conferred the knighthood of the British Empire as recognition of his important contribution. Although his name was twice nominated for Nobel Prize, unfortunately, he never received it., Competing Interests: There are no conflicts of interest., (Copyright: © 2021 Tropical Parasitology.)

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., Competing Interests: There are no conflicts of interest., (Copyright: © 2021 Tropical Parasitology.)

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.

Goldenseal (Hydrastis canadensis L.) is a medicinal plant widely used in various traditional systems of medicine and as a food supplement. It has been traditionally used by Native Americans as a coloring agent and as medicinal remedy for common diseases and conditions like wounds, digestive disorders, ulcers, skin and eye ailments, and cancer. Over the years, goldenseal has become a popular food supplement in the USA and other regions. The rhizome of this plant has been used for the treatment of a variety of diseases including, gastrointestinal disorders, ulcers, muscular debility, nervous prostration, constipation, skin and eye infections, cancer, among others. Berberine is one of the most bioactive alkaloid that has been identified in different parts of goldenseal. The goldenseal extract containing berberine showed numerous therapeutic effects such as antimicrobial, anti-inflammatory, hypolipidemic, hypoglycemic, antioxidant, neuroprotective (anti-Alzheimer's disease), cardioprotective, and gastrointestinal protective. Various research finding suggest the health promoting effects of goldenseal components and their extracts. However, few studies have also suggested the possible neurotoxic, hepatotoxic and phototoxic activities of goldenseal extract and its alkaloids. Thus, large randomized, double-blind clinical studies need to be conducted on goldenseal supplements and their main alkaloids to provide more evidence on the mechanisms responsible for the pharmaceutical activity, clinical efficacy and safety of these products. Thus, it is very important to review the scientific information about goldenseal to understand about the current scenario., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Context: Both malaria and lymphatic filariasis (LF) are mosquito-borne diseases caused by protozoal and nematode parasites, respectively, and are serious public health problem in India. Although the vectors of the diseases are different, they can coexist in favorable conditions. Fever is the common symptom for both the diseases, but the emphasis is given for diagnosis and treatment of malaria due to its life-threatening severity, LF remained neglected. Detection and management of microfilaria are equally important. During the diagnosis of malaria, a few microfilaria were detected, which prompted us to undertake this study with following objectives., Objectives: The objective of the study was to determine the incidence of microfilaremia among the febrile patients attending for malaria diagnosis., Subjects and Methods: Thick and thin peripheral blood smears from all patients attended were examined following Giemsa staining. Different malarial indexes were analyzed., Results: Out of 8681 patients examined, 1778 were positive for Plasmodium vivax and 328 for Plasmodium falciparum with a slide positivity rate 20.48%. Twenty-six patients were positive for microfilaria of Wuchereria bancroftii among which five were coinfected with P. vivax and one with P. falciparum . Most of the microfilaria-positive patients were adult and originally from northern districts of Bihar., Conclusions: High incidence of microfilaria among febrile patients attending for malaria is alarming for urban Kolkata. Although the patients were originally from Bihar, they are staying in Kolkata for a long time, might be a source for transmission. Epidemiological study by collecting night blood samples and entomological survey is highly suggestive to explore local transmission if any., Competing Interests: There are no conflicts of interest., (Copyright: © 2021 Tropical Parasitology.)

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., Competing Interests: Declaration of Competing Interest We have no conflicts of interest concerning the work reported in this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Our previous study exhibited free radicals scavenging and antioxidant activities of ethanolic and aqueous extracts of Tamarindus indica L. leaves in chronic sodium fluoride poisoning in rats. Tamarindus indica L. seed extract was also reported to have anti-arthritic efficacy by inhibiting cartilage and bone degrading factors. Therefore, an attempt was made to evaluate the effects of ethanolic extract of Tamarindus indica L. leaves in septic arthritis., Methods: The safety study was performed by oral dosing of ethanolic extract of the plant leaves at 2 g kg - 1 for consecutive 28 days in rabbits. Septic arthritis was induced in rabbits by single intra-articular inoculation of 10 4 c.f.u. of Staphylococcus aureus to the left stifle joint and was monitored by bacterial colony count, some relevant biochemical parameters and histopathological interpretation of the affected joint. For efficacy evaluation in septic arthritis, linezolid at 75 mg kg - 1 twice daily for 10 days and the ethanolic extract of Tamarindus indica L. at 500 and 1000 mg kg - 1 for consecutive 14 days were administered orally to the rabbits after 48 h of induction of arthritis., Results: In sub-acute toxicity study of Tamarindus indica L. leaves ethanolic extract, no significant change between days was found for aspertate aminotransferase, alanine transaminase, alkaline phosphatase, blood urea nitrogen and creatinine compared to day 0 values of the same group. The bacterial colony count of synovial fluid following Staphylococcus aureus inoculation to left stifle joint was found to be 1.08 ± 0.47 and 1.19 ± 0.29 c.f.u. mL - 1 in ethanolic extract low dose and high dose groups respectively, on day 2 which was reduced to 0.057 ± 0.036 c.f.u. mL - 1 and nil on day 16. The test extract was also found to markedly reduce simultaneous glucose difference, total protein ratio of serum and synovial fluid, joint radius and joint narrowing., Conclusion: Ethanolic extract of Tamarindus indica L. leaves at 500 mg kg - 1 and 1000 mg kg - 1 produced anti-arthritic effects against S. aureus induced septic arthritis in rabbits. However, the ethanolic extract at 1000 mg kg - 1 orally for consecutive 14 days showed better effects in septic arthritis.

Background: Aedes albopictus and Aedes aegypti are the major vectors of arboviral diseases. As effective vaccines are not available for most of the arboviral diseases, vector control by using insecticides play the key role to reduce the disease transmission. The emergence and spread of resistance to different classes of insecticides by the vectors is a major obstacle to control the disease transmission. Information about vector susceptibility to different insecticides and their mechanisms are very important for formulating proper vector control measures. The present study was designed to assess the susceptibility of Ae. aegypti against three different classes of adulticides, one larvicidal agent available and polymorphisms in the voltage-gated sodium channel (VGSC) gene related to insecticide resistance., Methods: Immature stages of Ae. aegypti were collected from three dengue endemic municipal areas of West Bengal and reared in the laboratory. Larvae and adults (F1 progeny) were used for insecticide bioassay as per WHO protocols. Knock down resistance gene (kdr) mutations were assessed by direct sequencing of PCR products., Results: The Ae. aegypti population was found to be susceptible to type II pyrethroids and malathion but highly resistant to DDT. A high rate of polymorphisms in the VGSC gene was observed among the collected mosquitoes. A double mutant V1016G + F1534C was found to be associated with DDT resistance but neither V1016G nor F1534C alone showed the same association. Association between the kdr mutations and the susceptibility status of pyrethroids could not be established due to very small sample size. A low to moderate level of resistance was noticed against temephos among the larval population based on WHO criteria., Conclusion: The replacement of DDT by type II pyrethroids for the management of dengue vectors is an appropriate decision taken by the national program which is supported by the findings of a higher level of resistance to DDT. Persistence of polymorphisms in the VGSC gene might be an indication of emergence of resistance against pyrethroid insecticides that should be monitored at a regular interval. Attempts should be made to determine the effectiveness of other larvicides for replacement of temephos if needed in future. Along with the chemical insecticides different biological vector control methods as well as biopesticides should also be used in vector control programmes., Competing Interests: The authors have declared that no competing interests exist.

Mesoporous bioactive glass (MBG) has drawn much attention due to its superior surface texture, porosity and bioactive characteristics. Aim of the present study is to synthesize MBG using different surfactants, viz., hexadecyltrimethylamonium(CTAB) (M1), poly-ethylene glycol (PEG) (M2) and pluronic P123 (M3); bioactivity study; and to understand their bone regeneration efficacy in combination with insulin-like growth factors (IGF-1) in animal bone defect model. SBF study revealed the formation of calcium carbonate (CaCO 3 ) and hydroxyapatite (HAp) phase over 14 days. Formation of apatite layer was further confirmed by FTIR, FESEM and EDX analysis. M1 and M2 showed improved crystallinity, while M3 showed slightly decrease in crystalline peak of CaCO 3 and enhanced HAp phase. More Ca-P layer formed in M1 and M2 supported the in vivo experiments subsequently. Degree of new bone formation for all MBGs were high, i.e., M1 (80.7 ± 2.9%), M2 (74.4 ± 2.4%) and M3 (70.1 ± 1.9%) compared to BG (66.9 ± 1.8%). In vivo results indicated that the materials were non-toxic, biodegradable, biocompatible, and is suitable as bone replacement materials. Thus, we concluded that growth factor loaded MBG is a promising candidate for bone tissue engineering application., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

A new generation of nanostructured glasses called mesoporous bioactive glasses (MBGs) exhibit superior surface texture, porosity and bioactive characteristics. The present study is carried out to develop and detailed characterize of ternary SiO 2 -CaO-P 2 O 5 MBG structure, fabricated by three different variations using different surfactants, e.g., hexadecyltrimethylammonium bromide (CTAB), poly-ethylene glycol,(PEG) and Pluronic P123. After thorough physico-chemical characterization, MBG granules were investigated for in vivo bone regeneration in animal bone defect model (rabbit) where standard S53P4 bioactive glass was used as control. All the synthesized MBG powders showed nano-range median particle size of 80-120 nm (MBG-CTAB), 50-70 nm (MBG-PEG and MBG-P123) while their specific surface area as 473.2, 52.2 and 169.3 m 2 /g respectively. All MBGs showed mesoporous nature corroborating transmission electron microscopy (TEM) observation as well. Bone regeneration property was measured after 45 and 90 days post-implantation at distal epiphysis of rabbit femur by radiography, histology, fluorochrome labeling, micro computed tomography (micro-CT) and vital organ histology. Results from in vivo studies indicated that the MBG materials produce minimal toxicity to the body. Furthermore, the biocompatibility and biodegradability of the implant makes them more suitable for application in bone tissue engineering. Among various implants, MBG fabricated using suitable surfactant (CTAB) shown the best result compared to other implants. Nonetheless, all the materials are suitable for application in bone tissue engineering and have potential for bone regeneration and healing., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

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., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Resistance to almost all available antimalarial agents, particularly for Plasmodium falciparum , is an important issue throughout all malaria endemic countries. Recently, Plasmodium vivax also showed resistance to chloroquine in some part of the World. Monitoring efficacy of used antimalarial drugs plays an important role to determine the emergence of resistant power by the prevailing parasite population of a geographical area if any. There are four different methods for antimalarial drug efficacy study. In vivo therapeutic efficacy study remains the gold standard and provides primary data for formulating antimalarial drug policy by the National Malaria Control Programmes. Several in vitro assay methods for assessing antimalarial drug susceptibility have been developed during past few decades. These assay methods are simple, easier to handle and allow early detection of drug-resistant parasites and also for the screening of different newly developed agents for their antimalarial activity. Approaches of different assay methods for testing the susceptibility of antimalarial agents and their limitations are discussed in this review article., Competing Interests: There are no conflicts of interest.

Emergence and spread of resistance among vectors toward different insecticides is a serious problem for the Japanese encephalitis (JE) control program. Regularly monitoring the status of susceptibility of vector species to insecticides is important for formulating proper vector control measures. In this study, we studied the susceptibility status of major JE vectors from northern West Bengal, toward 4% DDT, 0.05% deltamethrin, and 5% malathion. Two- to three-day-old unfed female mosquitoes were subjected to a susceptibility bioassay using a World Health Organization kit. Corrected mortality (CM) and knockdown times were estimated. Culex tritaeniorhynchus, Cx. vishnui, Cx. pseudovishnui, and Cx. gelidus were the major JE vectors present in the study areas. All 4 vector species were highly tolerant to DDT with CM ＜ 90%. Cx. tritaeniorhynchus, Cx. vishnui, Cx. pseudovishnui, and Cx. gelidus were tolerant to deltamethrin with CM ＜ 90%, except for Cx. gelidus of Darjeeling and Malbazar. At most of the study sites, malathion was effective against Cx. vishnui, Cx. pseudovishnui, and Cx. gelidus with CM ≥ 98%. In contrast, Cx. tritaeniorhynchus was tolerant to malathion in all study areas. Predominant JE vector populations were highly tolerant to all 3 analyzed insecticides, except deltamethrin for Cx. gelidus and malathion for Cx. vishnui, Cx. pseudovishnui, and Cx. gelidus. The results of this study may be useful for better planning and implementing a JE control strategy.

Background: The control and prevention of dengue largely depends on vector control measures, environmental management, and personal protection. Dengue control programmes are facing great challenges due to development of insecticide resistance among vector mosquitoes. Information on susceptibility status to different insecticides is important for national programmes to formulate vector control strategies., Methods: We have studied the larval susceptibility of Aedes albopictus to temephos and adult susceptibility to 4% DDT, 0.05% deltamethrin, and 5% malathion as per WHO protocols in the northern districts of West Bengal. Polymorphisms in the VGSC gene were studied by direct sequencing of PCR products., Results: The Ae. albopictus larval population showed sensitive [Resistance Ratio (RR99)<3] to moderate levels of resistance (510) to temephos at different study sites. Adult bioassay results revealed that Ae. albopictus was highly resistant to DDT [Corrected Mortality (CM) < 90%] in all the study sites and susceptible to deltamethrin and malathion (CM > 98%), except in Dhupguri where a low level of resistance to deltamethrin (CM = 96.25%) was recorded. None of the six important kdr mutations (S953P, I975M/V, L978, V980G, F1474C, D1703Y) were found in the VGSC of studied mosquitoes, but we identified 11 synonymous and 1 non-synonymous mutation in the VGSC gene., Conclusion: The higher susceptibility level to deltamethrin and malathion, along with the absence of important kdr mutations indicates that these two insecticides are still effective against Ae. albopictus in the study areas. The susceptibility status of temephos should be monitored closely as low to moderate levels of resistance were observed in few sites. A similar study is recommended for monitoring and early detection of insecticide resistance in other parts of the country.

Background: In India, sulphadoxine-pyrimethamine (SP) is now in use as a partner drug of ACT (AS+SP) to treat uncomplicated falciparum malaria since 2010. Declined trend of AS+SP efficacy has been reported from north-eastern states of the country. It is not possible to determine the efficacy of SP alone from any study with ACT. So, this work was designed to study the pattern of polymorphisms in pfdhfr and pfdhps genes to predict the SP resistance status among parasite population of urban Kolkata after five years of ACT implementation., Methods: A total of 125 P. falciparum positive patients were enrolled in the study during December 2014 to July 2016 and treated with AS+SP. Parasitic DNA was isolated and subjected to sequencing of pfdhfr and pfdhps genes directly from purified PCR products., Results: Genotyping of both the genes was successfully done in 113 isolates. In pfdhfr, 94.69% (107/113) isolates showed mutations at codon 59 and 108. A double mutant genotype ANRNI was mostly prevalent (107/113, 94.69%), while wild-type genotype ANCSI was found only in 5.3% (6/113) isolates. In pfdhps, mutations were recorded at codon 436 and 437 in 65.49% (74/113) and 23.01% (26/113) isolates, respectively. In combined pfdhfr-pfdhps genes, triple mutant ANRNI-FAKAA was most prevalent (45/113, 39.82%) followed by double mutant ANRNI-SAKAA (37/113, 32.74%) and quadruple mutant ANRNI-FGKAA (24/113, 21.24%)., Conclusion: SP resistance hallmark mutations i.e., quadruple (AIRNI-SAEAA) or quintuple (AIRNI-SGEAA) genotype in pfdhfr and pfdhps was absent which indicates that SP components of used ACT is still effective in the study area. It is also evident by the clinical response of AS+SP. Monitoring the efficacy of this combination (both by therapeutic and molecular marker study) at a regular interval is highly suggested to record any development of SP resistance in near future., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Asymptomatic leishmaniasis may drive the epidemic and an important challenge to reach the goal of joint Visceral Leishmaniasis (VL) elimination initiative taken by three Asian countries. The role of these asymptomatic carriers in disease transmission, prognosis at individual level and rate of transformation to symptomatic VL/Post Kala-azar Dermal Leishmaniasis (PKDL) needs to be evaluated. Asymptomatic cases were diagnosed by active mass survey in eight tribal villages by detecting antileishmanial antibody using rK39 based rapid diagnostic kits and followed up for three years to observe the pattern of sero-conversion and disease transformation. Out of 2890 total population, 2603 were screened. Antileishmanial antibody was detected in 185 individuals of them 96 had a history of VL/PKDL and 89 without such history. Seventy nine such individuals were classified as asymptomatic leishmaniasis and ten as active VL with a ratio of 7.9:1. Out of 79 asymptomatic cases 2 were lost to follow up as they moved to other places. Amongst asymptomatically infected persons, disease transformation in 8/77 (10.39%) and sero-conversion in 62/77 (80.52%) cases were noted. Seven (9.09%) remained sero-positive even after three years. Progression to clinical disease among asymptomatic individuals was taking place at any time up to three years after the baseline survey. If there are no VL /PKDL cases for two or more years, it does not mean that the area is free from leishmaniasis as symptomatic VL or PKDL may appear even after three years, if there are such asymptomatic cases. So, asymptomatic infected individuals need much attention for VL elimination programme that has been initiated by three adjoining endemic countries.

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., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The study of genetic diversity of Plasmodium falciparum is necessary to understand the distribution and dynamics of parasite populations. The genetic diversity of P. falciparum merozoite surface protein-1 and 2 has been extensively studied from different parts of world. However, limited data are available from India. This study was aimed to determine the genetic diversity and multiplicity of infection (MOI) of P. falciparum population in Kolkata, West Bengal, India. A total of 80day-zero blood samples from Kolkata were collected during a therapeutic efficacy study in 2008-2009. DNA was extracted; allelic frequency and diversity were investigated by PCR-genotyping method for msp1 and msp2 gene and fragment sizing was done by Bio-Rad Gel-Doc system using Image Lab (version 4.1) software. P. falciparum msp1 and msp2 markers were highly polymorphic with low allele frequencies. In Kolkata, 27 msp1 different genotypes (including 11of K1, 6 of MAD20 and 10 of Ro33 allelic families) and 30 different msp2 genotypes (of which 17 and 13 belonged to the FC27 and 3D7 allelic families, respectively) were recorded. The majority of these genotypes occurred at a frequency below 10%. The mean MOI for msp1 and msp2 gene were 2.05 and 3.72, respectively. The P. falciparum population of Kolkata was genetically diverse. As the frequencies of most of the msp1 and msp2 alleles were low, the probability of new infection with genotype identical to that in pretreatment infection was very rare. This information will serve as baseline data for evaluation of malaria control interventions as well as for monitoring the parasite population structure., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Dietary spices and their active constituents provide various beneficial effects on the gastrointestinal system by variety of mechanisms such as influence of gastric emptying, stimulation of gastrointestinal defense and absorption, stimulation of salivary, intestinal, hepatic, and pancreatic secretions. Capsicum annuum (Solanaceae), commonly known as chilli, is a medicinal spice used in various Indian traditional systems of medicine and it has been acknowledged to treat various health ailments. Therapeutic potential of chilli and capsaicin were well documented; however, they act as double-edged sword in many physiological circumstances. In traditional medicine chilli has been used against various gastrointestinal complains such as dyspepsia, loss of appetite, gastroesophageal reflux disease, gastric ulcer, and so on. In chilli, more than 200 constituents have been identified and some of its active constituents play numerous beneficial roles in various gastrointestinal disorders such as stimulation of digestion and gastromucosal defense, reduction of gastroesophageal reflux disease (GERD) symptoms, inhibition of gastrointestinal pathogens, ulceration and cancers, regulation of gastrointestinal secretions and absorptions. However, further studies are warranted to determine the dose ceiling limit of chilli and its active constituents for their utilization as gastroprotective agents. This review summarizes the phytochemistry and various gastrointestinal benefits of chilli and its various active constituents.

Post Kala-azar Dermal Leishmaniasis (PKDL) is a chronic but not life-threatening disease; patients generally do not demand treatment, deserve much more attention because PKDL is highly relevant in the context of Visceral Leishmaniasis (VL) elimination. There is no standard guideline for diagnosis and treatment for PKDL. A species-specific PCR on slit skin smear demonstrated a sensitivity of 93.8%, but it has not been applied for routine diagnostic purpose. The study was conducted to determine the actual disease burden in an endemic area of Malda district, West Bengal, comparison of the three diagnostic tools for PKDL case detection and pattern of lesion regression after treatment. The prevalence of PKDL was determined by active surveillance and confirmed by PCR based diagnosis. Patients were treated with either sodium stibogluconate (SSG) or oral miltefosine and followed up for two years to observe lesion regression period. Twenty six PKDL cases were detected with a prevalence rate of 27.5% among the antileishmanial antibody positive cases. Among three diagnostic methods used, PCR is highly sensitive (88.46%) for case confirmation. In majority of the cases skin lesions persisted after treatment completion which gradually disappeared during 6-12 months post treatment period. Reappearance of lesions noted in two cases after 1.5 years of miltefosine treatment. A significant number of PKDL patients would remain undiagnosed without active mass surveys. Such surveys are required in other endemic areas to attain the ultimate goal of eliminating Kala-azar. PCR-based method is helpful in confirming diagnosis of PKDL, referral laboratory at district or state level can achieve it. So a well-designed study with higher number of samples is essential to establish when/whether PKDL patients are free from parasite after treatment and to determine which PKDL patients need treatment for longer period.

In India, artemisinin-based combination therapy (ACT; specifically artesunate + sulfadoxine-pyrimethamine) has been implemented for uncomplicated falciparum malaria since 2010. But for vivax malaria drug policy remained unchanged i.e., chloroqine and primaquine. We observed the impact of this intervention in urban Kolkata by analyzing data from the Malaria Clinic from 2001 to 2013. In Kolkata, we observed that Plasmodium vivax was perennial, whereas P. falciparum infection was seasonal. Before ACT implementation, the proportion of P. falciparum was as high as 50% and it steadily decreased during 4 successive years post intervention. No change was observed in the number of P. vivax cases. ACT may be an effective measure in reducing falciparum malaria cases. Artemisinin-derivative combination therapies should be explored in vivax malaria to reduce the overall burden of malaria.

Molecular markers associated with artemisinin resistance in Plasmodium falciparum are yet to be well defined. Recent studies showed that polymorphisms in K13 gene are associated with artemisinin resistance. The present study was designed to know the pattern of polymorphisms in propeller region of K13 gene among the clinical isolates collected from urban Kolkata after five years of ACT implementation. We collected 59 clinical isolates from urban Kolkata and sequenced propeller region of K13 gene in 51 isolates successfully. We did not find any mutation in any isolate. All patients responded to the ACT, a combination of artesunate + sulphadoxine-pyrimethamine. The drug regimen is still effective in the study area and there is no sign of emergence of resistance against artemisinin as evidenced by wild genotype of K13 gene in all isolates studied.

Objective: The objective was to study the effect of Bauhinia variegata L. stem bark powder as adjunct therapy in chronic Staphylococcus aureus mastitis in goat., Materials and Methods: Mastitis was induced by intracisternal inoculation of coagulase positive S. aureus (J638) at the concentration of 2000 colony forming units. Group I animals were treated with repeated dose of ceftriaxone at 20 mg/kg intravenously, and Group II animals were treated with once daily oral administration of B. variegata L. stem bark powder at 6 g/kg for 7 days followed by maintenance dose at 3 g/kg for next 7 days along with repeated dose of the antibiotic at 20 mg/kg intravenously at 4 days interval., Results: No significant improvement in the clinical condition of the udder was noticed in the group treated with repeated dose of ceftriaxone alone. However, in the group treated with B. variegata L. stem bark powder along with repeated dose of ceftriaxone, no S. aureus colony was seen at 96 h and onwards in milk samples with a marked decrease in somatic cell count and milk alkaline phosphatase activity and increased lactoperoxidase activity. Further, plasma and milk concentration of ceftriaxone/ceftizoxime was increased, which indicated antibacterial, bioenhancing and antiinflammatory properties of the bark powder. The Group II animals also exhibited marked reduction in polymorphonuclear cells and fibrous tissue indicating antifibrotic property of B. variegata L., Conclusion: B. variegata L. stem bark powder can be considered as an effective adjunct therapy to intravenous ceftriaxone in S. aureus chronic mastitis in goat.

Sulfadoxine-pyrimethamine has never been recommended for the treatment of Plasmodium vivax malaria as the parasite is intrinsically resistant to pyrimethamine. The combination was introduced as a promising agent to treat Plasmodium falciparum malaria in many countries but was withdrawn after a few years due to development and spread of resistant strains. Presently, sulfadoxine-pyrimethamine is used as a partner drug of artemisinin-based combination therapy to treat uncomplicated falciparum malaria, and a combination of artesunate-sulfadoxine-pyrimethamine is currently in use in India. In countries like India, where both P. vivax and P. falciparum are equally prevalent, some proportion of P. vivax bacteria is exposed to sulfadoxine-pyrimethamine due to misdiagnosis and mixed infections. As reports on the in vivo therapeutic efficacy of sulfadoxine-pyrimethamine in P. vivax are rare, the study of mutations in the marker genes P. vivax dhfr (pvdhfr) and pvdhps is important for predicting drug selection pressure and sulfadoxine-pyrimethamine resistance monitoring. We studied the prevalence of point mutations and haplotypes of both the genes in 80 P. vivax isolates collected from urban Kolkata, India, by the DNA sequencing method. Point mutation rates in both the genes were low. The double mutant pvdhfr A15N50R58N117I173 (mutations are in boldface) and the single mutant pvdhps genotype S382G383K512A553V585 were more prevalent, while 35% of the isolates harbored the wild-type genotype. The triple mutant ANRNI-SGKAV was found in 29.9% isolates. No quintuple mutant genotype was recorded. The P. vivax parasites in urban Kolkata may still be susceptible to sulfadoxine-pyrimethamine. Hence, a combination of antimalarial drugs like artesunate-sulfadoxine-pyrimethamine introduced for P. falciparum infection might be effective in P. vivax infection also. Study of the therapeutic efficacy of this combination in P. vivax is thus strongly suggested. (The study protocol was registered in the Clinical Trial Registry-India [CTRI] of the Indian Council of Medical Research under registration number CTRI/2011/09/002031.).

Pueraria tuberosa (Willd.) DC is a perennial herb commonly known as 'vidarikanda', distributed throughout south east Asia. The plant's tuber is widely used in ethanomedicine as well as in traditional systems of medicine, particularly in ayurveda. It has been used in various ayurvedic formulations as restorative tonic, antiaging, spermatogenic and immune booster and has been recommended for the treatment of cardiovascular diseases, hepatosplenomegaly, fertility disorders, menopausal syndrome, sexual debility and spermatorrhoea. Numerous bioactive phytochemicals, mostly isoflavonoids such as puerarin, genistein, daidzein, tuberosin and so on have been identified in the tuber. In vivo and in vitro studies have provided the support against traditional demands of the tuber as spermatogenic, immune booster, aphrodisiac, anti-inflammatory, cardiotonic and brain tonic. However, further studies are required to define the active phytochemical compositions and to validate its clinical utilisation in the herbal formulations for human uses. This review provides an overview of traditional applications, current knowledge on the phytochemistry, pharmacology and toxicology of P. tuberosa. This review also provides plausible hypotheses about how various isoflavones particularly puerarin, genistein and daidzein, individually or collectively, may be responsible for the therapeutic potential against a wide range of ailments.

Development of a vaccine against Plasmodium falciparum infection is an urgent priority particularly because of widespread resistance to most traditionally used drugs. Multiple evidences point to apical membrane antigen-1(AMA-1) as a prime vaccine candidate directed against P. falciparum asexual blood-stages. To gain understanding of the genetic and demographic forces shaping the parasite sequence diversity in Kolkata, a part of Pfama-1 gene covering domain-I was sequenced from 100 blood samples of malaria patients. Statistical and phylogenetic analyses of the sequences were performed using DnaSP and MEGA. Very high haplotype diversity was detected both at nucleotide (0.998±0.002) and amino-acid (0.996±0.001) levels. An abundance of low frequency polymorphisms (Tajima's D=-1.190, Fu & Li's D(∗) and F(∗)=-3.068 and -2.722), unimodal mismatch distribution and a star-like median-joining network of ama-1 haplotypes indicated a recent population expansion among Kolkata parasites. The high minimum number of recombination events (Rm=26) and a significantly high dN/dS of 3.705 (P<0.0001) in Kolkata suggested recombination and positive selection as major forces in the generation and maintenance of ama-1 allelic diversity. To evaluate the impact of observed non-synonymous substitutions in the context of AMA-1 functionality, PatchDock and FireDock protein-protein interaction solutions were mapped between PfAMA-1-PfRON2 and PfAMA-1-host IgNAR. Alterations in the desolvation and global energies of PfAMA-1-PfRON2 interaction complexes at the hotspot contact residues were observed together with redistribution of surface electrostatic potentials at the variant alleles with respect to referent Pf3D7 sequence. Finally, a comparison of P. falciparum subpopulations in five Indian regional isolates retrieved from GenBank revealed a significant level of genetic differentiation (FST=0.084-0.129) with respect to Kolkata sequences. Collectively, our results indicated a very high allelic and haplotype diversity, a high recombination rate and a signature of natural selection favoring accumulation of non-synonymous substitutions that facilitated PfAMA-1-PfRON2 interaction and hence parasite growth in Kolkata clinical isolates., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Objective: To study the in vivo efficacy of these two ACTs in the treatment of Plasmodium falciparum (P. falciparum malaria) in Kolkata and to determine the prevalence of mutant S769N codon of the PfATPase6 gene among field isolates of P. falciparum collected from the study area., Methods: A total of 207P. falciparum positive cases were enrolled randomly in two study arms and followed up for 42 days as per WHO (2009) protocol. A portion of PfATPase6 gene spanning codon S769N was amplified and sequenced by direct sequencing method., Results: It was observed that the efficacy of both the ACT regimens were highly effective in the study area and no mutant S769N was detected from any isolate., Conclusions: The used, combination AS+SP is effective and the other combination AM+LF might be an alternative, if needed., (Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.)

Asymptomatic infection by Plasmodium falciparum is an important obstacle to eliminating malaria. Asymptomatic carriers do not seek treatment for infection, and therefore they become a reservoir for the parasite. For this reason, these carriers pose a real public health risk. The systematic identification and treatment of asymptomatic infections should reduce the parasite reservoir. A large reduction in this pool will lower the chance of transmission of the disease. In this study, we screened a tribal population of 1,040 individuals in the Purulia district of West Bengal by using a dual-antigen rapid diagnostic kit (RDK), microscopy, and species-specific PCR. All positive individuals were treated with artemisinin-based combination therapy (ACT) (artesunate plus sulfadoxine-pyrimethamine) and followed for 42 days. Polymorphisms in candidate genes were screened by DNA sequencing. A significant proportion (8.4%) of the study population was infected with P. falciparum but showed no clinical manifestations. The PCR method was more sensitive in detecting infection than the RDK or microscopy. The efficacy of the ACT was 97%. In the pfcrt gene, the mutation K76T (the mutated amino acid is indicated by bold type) was found in 100% of the cases. In the pfmdr1 gene, the mutations N86Y and Y184F were noted in 55.5% and 11% of the cases, respectively. Six different haplotypes were identified in the pfdhfr-pfdhps genes. Most importantly, the quintuple mutant A(16)I(51)R(59)N(108)I(164)-S(436)G(437)E(540)A(581)A(613) was found in 10% of the isolates, which is potentially important for the development of sulfadoxine-pyrimethamine resistance. A significant proportion of the study population harboring P. falciparum does not seek treatment and therefore serves as a reservoir for the parasite, maintaining the natural cycle. If the National Vector Borne Disease Control Programme (NVBDCP) of India is to eliminate malaria, then this hidden parasite burden needs to be addressed properly. Similar study in other parts of the country could help to determine the magnitude of the problem.

Plasmodium vivax malaria, though benign, has now become a matter of concern due to recent reports of life-threatening severity and development of parasite resistance to different antimalarial drugs. The magnitude of the problem is still undetermined. The present study was undertaken to determine the in vivo efficacy of chloroquine (CQ) and chloroquine plus primaquine in P. vivax malaria in Kolkata and polymorphisms in the pvmdr1 and pvcrt-o genes. A total of 250 patients with P. vivax monoinfection were recruited and randomized into two groups, A and B; treated with chloroquine and chloroquine plus primaquine, respectively; and followed up for 42 days according to the WHO protocol of 2009. Data were analyzed using per-protocol analyses. We assessed polymorphisms of the pvmdr1 and pvcrt-o genes by a DNA-sequencing method. Out of the 250 patients recruited, 204 completed a 42-day follow-up period, 101 in group A and 103 in group B. In group A, the non-PCR-corrected efficacy of CQ was 99% (95% confidence interval [CI], 0.944 to 1.00), and in group B, all cases were classified as adequate clinical and parasitological response (ACPR). Day 3 positivity was observed in 11 (5.3%) cases. No specific mutation pattern was recorded in the pvcrt-o gene. Eight nonsynonymous mutations were found in the pvmdr1 gene, three of which were new. The Y976F mutation was not detected in any isolate. Chloroquine, either alone or in combination with primaquine, is still effective against P. vivax malaria in the study area. (The study protocol was registered in CTRI [Clinical Trial Registry-India] of the Indian council of Medical Research under registration no. CTRI/2011/09/002031.).

Context: Visceral leishmaniasis (VL), also known as Kala-azar (KA) is a public health problem of tropical and subtropical countries, which infects about 12 million people annually, out of which about 1.5 million are new cases. India contributes a major share of the global burden of VL. For many years leishmaniasis has been treated with pentavalent antimonials. Antimony resistance is a problem in India and in other different geographic areas of the world. Amphotericin B deoxycholate and pentamidine isethionate are effective by parenteral administration and associated with toxicities. The quest for an effective, orally administered, non-toxic and less expensive alternative resulted in the identification of miltefosine (hexadecylphosphocholine). In India, therapeutic efficacy of miltefosine in VL was assessed by many groups of scientists, mainly from Bihar and Uttar Pradesh. No such data is available from West Bengal., Aims: The present study was designed to observe the efficacy of miltefosine in VL in rural West Bengal., Materials and Methods: A total of 71 parasitologically proven VL patients participated in the study who received miltefosine in accordance with the National Vector Born Disease Control Programme (NVBDCP) of India and were followed up for the following one year., Results: The overall efficacy of the drug was 93% and no significant adverse side effects were observed during the study period., Conclusions: The study concludes that miltefosine is effective, well tolerated, and easily administrable drug in the treatment of visceral leishmaniasis at the field levels.

Background: Pueraria tuberosa (Fabaceae) is a well-known medicinal herbs used in Indian traditional medicines. The puerarin is one of the most important bioactive constituent found in the tubers of this plant. Quantitative estimation of bioactive molecules is essential for the purpose of quality control and dose determination of herbal medicines. The study was designed to develop a validated reversed phase high-performance liquid chromatography (RP-HPLC) method for the quantification of puerarin in the tuber extract of P. tuberosa., Materials and Methods: The RP-HPLC system with Luna C18 (2) 100 Å, 250 × 4.6 mm column was used in this study. The analysis was performed using the mobile phase: 0.1% acetic acid in acetonitrile and 0.1% acetic acid in water (90:10, v/v) under column temperature 25°C. The detection wavelength was set at 254 nm with a flow rate of 1 ml/min. The method validation was performed according to the guidelines of International Conference on Harmonization., Results: The puerarin content of P. tuberosa extract was found to be 9.28 ±0.09%. The calibration curve showed good linearity relationship in the range of 200-1000μg/ml (r (2)>0.99). The LOD and LOQ were 57.12 and 181.26μg/ml, respectively and the average recovery of puerarin was 99.73% ±1.02%. The evaluation of system suitability, precision, robustness and ruggedness parameters were also found to produce satisfactory results., Conclusions: The developed method is very simple and rapid with excellent specificity, accuracy and precision which can be useful for the routine analysis and quantitative estimation of puerarin in plant extracts and formulations.

The major disadvantage of a Serological test like Direct Agglutination Test (DAT) for Visceral Leishmaniasis (also called Kala-azar) is its inability to distinguish between recent and past infection. The objective of our study was to look at rate of decline of antibodies in fully cured cases of Kala-azar and length of time it takes for DAT to become negative. Cohort Study involving completely treated Kala-azar cases from Government Hospital during one calendar year of study. Cases were selected on the basis of treatment cohorts 0, 3, 6, 9 & 12 mo after completion of treatment.. Phase I--The cases were traced and after obtaining the informed consent they were subjected to Direct Agglutination Test (DAT). Phase II--The five treatment cohorts, constituting 82 cured cases (average of 15 cured cases per each treatment cohort) were tested again with DAT three months after the first test. The titers of Phase-I and phase-II tests were analyzed for the dynamics of the antibodies for the period. Cutoff-Values of DAT below 1:800 are considered negative. Values of 1:800, 1:1200, 1:1600 and so on are considered positive. The mean titer [Geometric Mean Titer (GMT)] at the start of treatment was 1:1120, which showed steady decline up to six months, plummeting below the cutoff titer for the DAT (1:800) at the ninth month. Antibodies continue to linger for about one year in cured Kala-azar cases even after correct and complete treatment. Single DAT results may be misleading due to high false positivity up to one year after the cure. Paired test defined as two tests 3 mo apart on the same subject. Paired test is highly recommended for diagnosis and prognosis. DAT is still a very useful tool for diagnosis if used along with clinical correlation.

In India, chloroquine has been replaced by a combination of artesunate and sulfadoxine-pyrimethamine (AS-SP) for uncomplicated P. falciparum malaria. Other available combinations, artemether-lumefantrine (AM-LF) and artesunate-mefloquine (AS-MQ), not included in the national program, are widely used by private practitioners. Little is known about the therapeutic efficacy of these artemisinin combinations and the prevalence of molecular markers associated with antimalarial drug resistance. A total of 157 patients with P. falciparum monoinfection were recruited and randomized into three study groups (AS-SP, AM-LF, and AS-MQ). All patients were followed up for 42 days to study the clinical and parasitological responses according to the WHO protocol (2009). We assessed the polymorphism of the pfATPase6, pfcrt, pfdhfr, and pfdhps genes by the DNA-sequencing method. The PCR-corrected therapeutic efficacies of AS-SP, AM-LF, and AS-MQ were 90.6% (95% confidence interval [CI], 0.793 to 0.969), 95.9% (95% CI, 0.860 to 0.995), and 100% (95% CI, 0.927 to 1.00), respectively. No specific mutational pattern was observed in the pfATPase6 gene. All isolates had a K76T mutation in the pfcrt gene. In the pfdhfr-pfdhps genotype, quadruple mutation was frequent, and quintuple mutation was documented in 6.3% of P. falciparum isolates. The significant failure rate of AS-SP (9.5%), although within the limit (10%) for drug policy change, was due to SP failure because of prevailing mutations in pfdhfr, I(51)R(59)N(108), with pfdhps, G(437) and/or E(540). The efficacy of this ACT needs periodic monitoring. Artemether-lumefantrine and artesunate-mefloquine are effective alternatives to the artesunate-sulfadoxine-pyrimethamine combination.

Genetic variations in toll-like receptors and cytokine genes of the innate immune pathways have been implicated in controlling parasite growth and the pathogenesis of Plasmodium falciparum mediated malaria. We previously published genetic association of TLR4 non-synonymous and TNF-α promoter polymorphisms with P.falciparum blood infection level and here we extend the study considerably by (i) investigating genetic dependence of parasite-load on interleukin-12B polymorphisms, (ii) reconstructing gene-gene interactions among candidate TLRs and cytokine loci, (iii) exploring genetic and functional impact of epistatic models and (iv) providing mechanistic insights into functionality of disease-associated regulatory polymorphisms. Our data revealed that carriage of AA (P = 0.0001) and AC (P = 0.01) genotypes of IL12B 3'UTR polymorphism was associated with a significant increase of mean log-parasitemia relative to rare homozygous genotype CC. Presence of IL12B+1188 polymorphism in five of six multifactor models reinforced its strong genetic impact on malaria phenotype. Elevation of genetic risk in two-component models compared to the corresponding single locus and reduction of IL12B (2.2 fold) and lymphotoxin-α (1.7 fold) expressions in patients'peripheral-blood-mononuclear-cells under TLR4Thr399Ile risk genotype background substantiated the role of Multifactor Dimensionality Reduction derived models. Marked reduction of promoter activity of TNF-α risk haplotype (C-C-G-G) compared to wild-type haplotype (T-C-G-G) with (84%) and without (78%) LPS stimulation and the loss of binding of transcription factors detected in-silico supported a causal role of TNF-1031. Significantly lower expression of IL12B+1188 AA (5 fold) and AC (9 fold) genotypes compared to CC and under-representation (P = 0.0048) of allele A in transcripts of patients' PBMCs suggested an Allele-Expression-Imbalance. Allele (A+1188C) dependent differential stability (2 fold) of IL12B-transcripts upon actinomycin-D treatment and observed structural modulation (P = 0.013) of RNA-ensemble were the plausible explanations for AEI. In conclusion, our data provides functional support to the hypothesis that de-regulated receptor-cytokine axis of innate immune pathway influences blood infection level in P. falciparum malaria.

Background: The Plasmodium vivax that was once prevalent in temperate climatic zones typically had an interval between primary infection and first relapse of 7-10 months, whereas in tropical areas P.vivax infections relapse frequently at intervals of 3-6 weeks. Defining the epidemiology of these two phenotypes from temporal patterns of illness in endemic areas is difficult or impossible, particularly if they overlap., Methods: A prospective open label comparison of chloroquine (CQ) alone versus CQ plus unobserved primaquine for either 5 days or 14 days was conducted in patients presenting with acute vivax malaria in Kolkata. Patients were followed for 15 months and primary and recurrent infections were genotyped using three polymorphic antigen and up to 8 microsatellite markers., Results: 151 patients were enrolled of whom 47 (31%) had subsequent recurrent infections. Recurrence proportions were similar in the three treatment groups. Parasite genotyping revealed discrete temporal patterns of recurrence allowing differentiation of probable relapse from newly acquired infections. This suggested that 32 of the 47 recurrences were probable relapses of which 22 (69%) were genetically homologous. The majority (81%) of probable relapses occurred within three months (16 homologous, 10 heterologous) and six genetically homologous relapses (19%) were of the long latency (8-10 month interval) phenotype., Conclusions: With long follow-up to assess temporal patterns of vivax malaria recurrence, genotyping of P.vivax can be used to assess relapse rates. A 14 day unobserved course of primaquine did not prevent relapse. Genotyping indicates that long latency P.vivax is prevalent in West Bengal, and that the first relapses after long latent periods are genetically homologous., Trial Registration: Controlled-Trials.com ISRCTN14027467.

Objective: In India, till recently, Chloroquine was used as first-line therapy in areas with Chloroquine sensitive Plasmodium falciparum malaria cases. The National Vector Borne Disease Control Programme (NVBDCP) has introduced artemisinin combination therapy (ACT) as first-line option to treat all P. falciparum cases in the country. This study was carried out to ascertain the efficacy of Chloroquine and Sulphadoxine-Pyrimethamine, either alone or in combination, before the launch of ACT by NVBDCP., Methods: A total of 300 P. falciparum malaria cases were enrolled randomly in three study arms, Chloroquine (CQ), Sulphadoxine-Pyrimethamine (SP) and Chloroquine plus Sulphadoxine-Pyrimethamine (CQ + SP). All patients were followed up for 28 days as per WHO (Assessment and Monitoring of Antimalarial Drug Efficacy for the Treatment of Uncomplicated Falciparum Malaria, Geneva, 2003) Protocol. Paired blood samples of treatment failure cases were collected and subjected to MSP 1, MSP 2 and GLURP genotyping for differentiation between re-infection and recrudescence. The data were analysed by Kaplan-Meier survival curve according to WHO standard procedures., Results: The overall failure rate including both early treatment failure (ETF) and late treatment failure (LTF) of CQ, SP and CQ + SP were 61%, 14% and 8%, respectively, in the study area. Of 60 recurrent malaria cases, genotyping was successful in 49 cases, revealing that most of the (46/49; 94%) cases of recurrent malaria were due to recrudescence., Conclusion: In Jalpaiguri District the overall failure rate of CQ was 61% and of SP 14%, which was well above the WHO recommended cut-off threshold level (10%) for change of drug policy., (© 2011 Blackwell Publishing Ltd.)

Dysregulated innate immune responses due to inappropriate signaling by Toll-like receptors (TLRs) and aberrant production of pro-inflammatory cytokines are implicated in the immunopathology and disease outcome in Plasmodium falciparum malaria. This study investigates the relationship between polymorphic variability of candidate genes including TLR-2, -4, -9, tumor necrosis factor-alpha and lymphotoxin-alpha and blood infection level in Indian mild malaria patients. Genotyping was carried out by PCR-RFLP and sequencing. Association of parasite load with genotypes was examined using model based and model free approaches. Allele and haplotype based risk assessment for disease severity was performed by stratifying the patients into high and low parasitemic groups on the basis of a threshold value derived by employing a two-component mixture model and expectation-maximization algorithm. The mean parasitemia was significantly increased for variant homozygous genotype (C/C) at TNF-alpha promoter -1031 and major homozygous genotypes encoding Asp/Asp and Thr/Thr at codons 299 and 399, respectively, on TLR4 polypeptide. Individuals harboring combined genotype C/C-Asp/Asp-Thr/Thr on TNF-alpha and TLR4 presented the highest parasite load. The frequencies of variant allele C in TNF-1031 (OR=1.91 with 95% CI=1.24-2.94) and TNF-alpha promoter haplotypes C-C-G-G (OR=1.99 with 95% CI=1.21-3.27) and C-C-G-A (OR=2.96 with 95% CI=1.19-7.37) pertaining to loci TNF-1031/-857/-308/-238 were significantly elevated in the high parasitemic group. On the contrary, the frequencies of variant allele encoding Ile at 399 (OR=0.55 with 95% CI=0.32-0.94) and haplotype corresponding to Gly-Ile (299-399) (OR=0.51 with 95% CI=0.28-0.9) in TLR4 were higher in low parasitemic group. In silico analysis indicate differential binding of transcription factors to TNF-alpha promoter haplotypes and alteration in the surface charge distribution of the TLR4 variant proteins. Our results support a genetic role of TLR4 and TNF-alpha in controlling the blood infection level in mild malaria., (Copyright 2010 Elsevier B.V. All rights reserved.)

Objectives: Nelumbo nucifera Gaertn. (Nymphaeaceae), also known as sacred lotus, is a well known medicinal plant. This article reviews the traditional uses, phytochemistry and therapeutic reports on different parts of N. nucifera viz. the seeds, rhizomes, leaves and flowers. This review also describes various compounds isolated from different parts of this plant and the therapeutic benefits derived from those phytoconstituents., Key Findings: There are several therapeutic benefits of this plant for which different parts are used. The extracts of rhizomes, seeds, flowers and leaves have been reported to have varied therapeutic potential. Several bioactive compounds have been derived from these plant parts belonging to different chemical groups, including alkaloids, flavonoids, glycosides, triterpenoid, vitamins etc., which all have their own therapeutic impact. Thus, the pharmacological effects and various active ingredients of different parts of N. nucifera are well understood., Summary: In this review we explore the current pharmaceutical, phytochemical and pharmacological knowledge about this well known plant species as well as several emerging aspects for research on N. nucifera.