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Chloroquine is still used as a first-line treatment for uncomplicated Plasmodium vivax malaria in India and resistance to this therapy can act as a major hurdle for malaria elimination. It is difficult to monitor drug-efficacy and drug resistance through in vivo and in vitro studies in case of Plasmodium vivax so analysis of molecular markers serves as an important tool to track resistance. Molecular methods that are currently in use for detecting single nucleotide polymorphisms in resistant genes including Polymerase chain reaction (PCR), Realtime-Polymerase chain reaction require highly sophisticated labs and are time consuming. So, with this background the study has been designed to optimize Loop Mediated Isothermal Amplification Assay to detect single nucleotide polymorphisms in chloroquine resistance gene of Plasmodium vivax in field settings. Eighty-eight Plasmodium vivax positive samples were collected. Pvmdr1 gene was amplified for all the samples and sequenced. Obtained sequences were analyzed for the presence of single nucleotide polymorphisms in the target gene. Further Loop Mediated Isothermal Amplification Assay primer sets were designed for the target mutants and the assay was optimized. Clinical as well as analytical sensitivity and specificity for the assay was calculated. Double mutants with variations at T958M and F1076L were detected in 100% of the Plasmodium vivax clinical isolates with haplotype M958 Y976 Y1028 L1076. Designed primers for Loop Mediated Isothermal Amplification Assay successfully detected both the mutants (T958M and F1076L) in 100% of the isolates and do not show cross-reactivity with other strains. So, the assay was 100% sensitive and specific for detecting single nucleotide polymorphisms in the target Pvmdr1 gene. Limit of detection was found to be 0.9 copies/µl and lowest DNA template concentration detected by designed assay was 1.5 ng/µL. Observed prevalence of single nucleotide polymorphisms in Pvmdr 1 gene is indicating a beginning of trend towards chloroquine resistance in Plasmodium vivax. The present study optimized LAMP for detecting single nucleotide polymorphisms in Plasmodium vivax cases in field settings, thus would help in finding significant hubs of emerging chloroquine drug resistance and ultimately helping in the management of suitable antimalarial drug policy. [ABSTRACT FROM AUTHOR]

Neuropsychiatric disorders, ranging from mild cognitive impairment to frank psychosis, have been associated with certain parasitic infections. The parasite may cause damage to the central nervous system in several ways: as a space‑occupying lesion (neuro‑cysticercosis), alteration of neurotransmitters (toxoplasmosis), generation of the inflammatory response (trypanosomiasis, schistosomiasis), hypovolemic neuronal injury (cerebral malaria), or a combination of these. Certain drugs like quinacrine (mepacrine), mefloquine, quinolone, and interferon alpha which are used to treat these parasitic infections can further cause neuropsychiatric adverse effects. This review summarizes the major parasitic infections that are associated with neuropsychiatric disorders and the pathogenesis involved in their processes. A high index of suspicion for parasitic diseases, especially in endemic areas, should be kept in patients presenting with neuropsychiatric symptoms. A multidimensional approach to identification of the offending parasite using serological, radiological, and molecular tests is required not only to ensure proper and prompt treatment of the primary parasitic infection but also to improve the prognosis of patients by complete resolution of neuropsychiatric symptoms. [ABSTRACT FROM AUTHOR]

Taenia solium is a widespread zoonotic tapeworm that predominantly affects regions of Latin America, South and South-East Asia, and Sub-Saharan Africa. Neurocysticercosis (NCC), the presence of T. solium cysts in the brain is associated with diverse clinical manifestations, such as epilepsy, seizures, and neurological deficits. It is a significant cause of preventable epilepsy globally, accounting for approximately 30% of cases in endemic regions. The diagnosis of neurocysticercosis relies on neuroimaging techniques, but these resources are often limited in low-income countries, resulting in an underestimation of the disease burden. The present study enrolled 141 patients who were clinically suspected and radiologically confirmed for NCC at the Neurology OPD of PGIMER, Chandigarh. Additionally, 98 control subjects attending the PGIMER OPD for investigation were also included. Plasma and urine samples were collected from all participants for further analysis. Cell-free DNA extraction was performed using specific kits, and the quality of the extracted DNA was assessed. The RT-LAMP assay targeted the cox1 gene. Real-time RT-LAMP results were evaluated using a fluorescence graph obtained with the Genei III fluorimeter. Among a group of patients diagnosed with NCC, the gene was identified in 74.4% of plasma samples and 67.3% of urine samples. In comparison, the T. solium cox1 gene was found in 6.1% of control subjects in plasma and urine samples using the LAMP assay. In conclusion, the study emphasises the need for improved diagnostic methods for NCC and presents promising alternatives, such as RT-LAMP and urine-based cell-free DNA analysis. These approaches offer advantages in terms of cost-effectiveness, simplicity, and diagnostic accuracy. • Real-time LAMP, targeting infection-induced cell-free DNA in plasma and urine samples. • RT-LAMP assay examines the presence of the cox1 gene from Taenia solium. • Six sets of primers, including loop primers, were employed to enhance efficiency. • LAMP assay demonstrated a sensitivity of 74.4% for plasma and 67.3% for urine samples. • LAMP assay exhibited a specificity of 93.8% for both plasma and urine samples. [ABSTRACT FROM AUTHOR]

Background: Neurocysticercosis is a common cerebral parasitic infestation, caused due to pork tapeworm infection the infestations risks parallels the socio-economic status, personal hygiene and education. The effect of NCC was assessed in neurocognition. Objective: To study demographic characteristics and neurocognitive domains of patients with Neurocysticercosis Methods: Neurocysticercosis diagnosed patients by CT, MRI and LAMP tests. MMSE score was measured for assessment. Results: MMSE score were reduced in majority of the patients. In attention was the most common deficit found. Repeat MMSE assessment done in 6 patients showed an improvement of scores post therapy Conclusion: Cognitive involvement is common in NCC and is a major cause of morbidity. [ABSTRACT FROM AUTHOR]

Objective: The aim of study was to characterize Giardia isolates genetically among patients in Chandigarh region, India. For this, nested PCR targeting fragment of the glutamate dehydrogenase (GLUD1 earlier named as GDH) gene was used. Phylogenetic analysis was done by constructing neighbor-joining tree made out of the nucleotide sequences of G. intestinalis isolates obtained in this study and with the known sequences published in GenBank. Results: Out of 40 samples, GLUD1 gene was amplified in 33 samples (82.5%). The product of GLUD1 gene was successfully sequenced only in 32 samples. In these samples, assemblage B was found in 27 (84.37%) samples whereas 5 (15.6%) samples had assemblage A. Among assemblage B most of them were of BIII. Therefore, genotyping of Giardia would be helpful in conducting epidemiological studies. [ABSTRACT FROM AUTHOR]

Among the parasitic diseases, amoebic liver abscess (ALA) ranks second to malaria in terms of mortality. Due to the poor sensitivity of conventional diagnostic methods, there is a need for the development of effective and rapid diagnostic methods for ALA. Thus, the purpose of this work was to develop a real‐time loop‐mediated isothermal amplification (RT‐LAMP) assay specific to Entamoeba histolytica. Further, we compared the performance of real‐time LAMP with conventional and real‐time PCR (RT‐PCR) targeting 18S small subunit ribosomal RNA (18S SSU rRNA) gene of E. histolytica in patients with ALA. A total of 126 liver samples were obtained for the study. Of these, 96 aspirated pus samples were obtained from patients suffering from an ALA (serology confirmed, anti‐amoebic immunoglobulin IgG positive), 19 aspirated pus samples from patients with pyogenic liver abscess (PLA, 16S RNA gene positive) and 11 autopsy liver tissues. The results showed that the DNA of E. histolytica was detected in 81 samples by conventional PCR, 93 by RT‐PCR and 95 by RT‐LAMP. The analytical sensitivity of the RT‐LAMP assay was much higher than the other two techniques. RT‐LAMP assay was able to amplify up to one copy of the targeted gene of E. histolytica while conventional PCR and RT‐PCR could amplify up to 103 and 102 copies of the targeted gene of E. histolytica, respectively. In conclusion, RT‐LAMP proved to be a sensitive, specific and rapid test which can be utilised as an effective tool for the diagnosis of ALA. [ABSTRACT FROM AUTHOR]

Abstract: Entamoeba histolytica infection is associated with considerable morbidity and mortality in the form of intestinal and extraintestinal amoebiasis. No vaccine is yet available for amoebiasis. Heparan Sulphate Binding Proteins (HSBPs) from E. histolytica were evaluated for immunogenicity and protective efficacy in a Guinea pig model. Animals were immunized subcutaneously with 30μg of HSBP by three weekly inoculations. The immunogenicity of HSBP was determined by antibody response (IgG, IgM and IgA), splenocyte proliferation assay and in vitro direct amoebicidal assay with splenic lymphocytes and monocytes from vaccinated and control animals. The efficacy of the vaccine was evaluated by challenge infection to vaccinated and control animals by intra-caecal inoculation of E. histolytica trophozoites and comparing gross and histopathological findings in caeca of these animals. HSBP was found to induce specific anti-amoebic response as seen by specific antibody production and direct amoebicidal activity of splenocytes. The vaccine also showed partial protection against challenge infection in vaccinated animals as shown by mild/absent lesions and histopathological findings. [Copyright &y& Elsevier]

Adherence of pathogenic Entamoeba histolytica trophozoites mediated by Gal/GalNAc lectin is a prerequisite for killing naı¨ve T cells and monocytes but the activated T cells and monocyte derived macrophages (MDMs) not only resist the attack but can kill the parasite. In the present study, we have analysed the adherence and cytotoxicity of the immunecompetent cells from patients of amoebic liver abscess at the time of their diagnosis and after 3 months to elucidate the development of cell mediated cytotoxicity, a major mechanism of resistance to amoebic infection. The results show that CD3+ cells from amoebic liver abscess cases, when stimulated, in vitro, bound E. histolytica trophozoites with increased intensity and their viability was also increased. The activated lymphocytes (taken at 3 months post treatment) were also able to kill amoebae. MDMs bound amoebae with greater intensity than lymphocytes, until 3 months post infection. These MDMs were effective in killing approximately 40% amoebae which was significantly less than at the time of diagnosis but was very significant as compared to the controls. The data suggest that cell mediated cytotoxic responses are maximum until 1 month post treatment and are significantly reduced thereafter. [Copyright &y& Elsevier]

Background & objectives The emergence of drug resistance in leishmaniasis has remained a concern. Even new drugs have been found to be less effective within a few years of their use. Coupled with their related side effects and cost-effectiveness, this has prompted the search for alternative therapeutic options. In this study, the Computer Aided Drug Design (CADD) approach was used to repurpose already existing drugs against Leishmania major. The enzyme lanosterol 14-alpha demethylase (CYP51), in L. major , was chosen as the drug target since it is a key enzyme involved in synthesizing ergosterol, a crucial component of the cell membrane. Methods A library of 1615 FDA-approved drugs was virtually screened and docked with modeled CYP51 at its predicted binding site. The drugs with high scores and high affinity were subjected to Molecular Dynamics (MD) simulations for 100 ns. Finally, the compounds were tested in vitro using an MTT [3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide] assay against the promastigotes of L. major. Results Computational screening of FDA-approved drugs identified posaconazole and isavuconazole as promising candidates, as both drugs target the CYP51 enzyme in fungi. Molecular dynamics (MD) simulations demonstrated that both drugs form stable complexes with the target enzyme. In vitro studies of posaconazole and isavuconazole against promastigotes of L. major demonstrated significant efficacy, with IC50 values of 2.062±0.89 µg/ml and 1.202±0.47 µg/ml, respectively. Interpretation & conclusions The study showed that the existing FDA-approved drugs posaconazole and isavuconazole can successfully be repurposed for treating L. major by targeting the CYP51 enzyme, demonstrating significant efficacy against promastigotes. [ABSTRACT FROM AUTHOR]

Leishmaniasis is a vector-borne parasitic infection caused by the infective bite of female Phlebotomine sandflies. Treatment of leishmaniasis by conventional synthetic compounds is met by challenges pertaining to adverse effects which call for the discovery of newer anti-leishmanial molecules. This study was performed to evaluate the effect and modes of action of a sesquiterpene alcoholic molecule Farnesol on Leishmania major, the causative agent of Zoonotic CL. The cytotoxic effect of Farnesol against L.major promastigotes, amastigotes and macrophages was assessed by MTT test and counting. The IC50 on promastigotes by Farnesol on L.major was also evaluated by flow cytometry. In the findings, promastigotes were reduced at 167μM. The mean numbers of L.major amastigotes in macrophages were significantly decreased on exposure to Farnesol at 172μM. In addition, Farnesol induced significant apoptosis dose-dependent on L.major promastigotes. In silico protein-ligand_binding analyses indicated the effect of Farnesol in perturbation of the ergosterol synthesis pathway of Leishmania with attributes suggesting inhibition of Lanosterol-α-demethylase, the terminal enzyme of ergosterol synthesis machinery. Findings from flow cytometry reveal the role of Farnesol in apoptosis-induced killing in promastigotes. Farnesol was effective at very lower concentrations when compared to Paromomycin. Further studies are crucial to evaluate the therapeutic potential of Farnesol alone or in combination with other conventional drugs in animal models. [ABSTRACT FROM AUTHOR]