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Background: Typhoid fever is an infectious disease of serious public health concern. The anti-typhoid efficacy and safety of Ximenia americana stem bark extracts on the nephrocytes, hepatocytes, and haematological parameters of Salmonella typhi infected Wistar albino rats was evaluated. Methodology: Experimental rats in their respective groups were infected with S. typhi via oral administration of infective dose (0.5 ml of 1.5 × 104 cfu/ml) of cultured S. typhi, while group 1 were uninfected. Group 2 were untreated, group 3 were treated with the standard drug (ciprofloxacin), while groups 4, 5, 6 were treated with (100, 200, 300 mg/kg) body weight of the methanol extract of X. americana stem bark, and groups 7, 8 and 9 were treated with (100, 200, 300 mg/kg) body weight of the ethyl-acetate fraction of X. americana stem bark administered orally for 7 days. Acute toxicity test was carried out using standard methods and signs accompanying toxicity such as drowsiness, respiratory distress, diarrhoea, and possible death of animals were monitored for 24 h in an interval of 2 h. Physical parameters were also monitored to check symptom amelioration as the treatment progresses. Results: Acute toxicity test showed that experimental rats administered the methanol extract and ethyl-acetate fraction of X. americana had no clinical signs of toxicity such as drowsiness, respiratory distress, diarrhoea, and death. The Physical examination of rats in all the treatment groups showed improvement in symptoms following Salmonella infection as treatment progressed when compared to the untreated group. Salmonella infection led to a significant (p < 0.05) increase in serum urea, sodium, chloride levels, bilirubin concentration, total white cell count and activities of the liver enzymes as compared to normal and apparently healthy rats. A significant (p < 0.05) decrease in PCV, haemoglobin concentration, RBC count, serum potassium, total protein and creatinine levels of the infected groups was observed as compared to the normal uninfected group. The methanol extract at the dose of 300 mg/kg body weight had 15%, 37%, 52% reduction in sodium, urea, and alkaline phosphatase respectively and 57% increase in potassium while the ethyl-acetate fraction (300 mg/kg body weight) had 46% and 47% increase in PCV and haemoglobin and 36% reduction in ALT. Conclusion: The administration of the methanol extract and ethyl-acetate fraction of X. americana was accompanied by a significant (p

Due to low numbers of circulating tumor cells (CTCs) in liquid biopsies, there is much interest in enrichment of alternative circulating-like mesenchymal cancer cell subpopulations from in vitro tumor cultures for utilization within molecular profiling and drug screening. Viable cancer cells that are released into the media of drug-treated adherent cancer cell cultures exhibit anoikis resistance or anchorage-independent survival away from their extracellular matrix with nutrient sources and waste sinks, which serves as a pre-requisite for metastasis. The enrichment of these cell subpopulations from tumor cultures can potentially serve as an in vitro source of circulating-like cancer cells with greater potential for scale-up in comparison with CTCs. However, these live circulating-like cancer cell subpopulations exhibit size overlaps with necrotic and apoptotic cells in the culture media, which makes it challenging to selectively enrich them, while maintaining them in their suspended state. We present optimization of a flowthrough high frequency (1 MHz) positive dielectrophoresis (pDEP) device with sequential 3D field non-uniformities that enables enrichment of the live chemo-resistant circulating cancer cell subpopulation from an in vitro culture of metastatic patient-derived pancreatic tumor cells. Central to this strategy is the utilization of single-cell impedance cytometry with gates set by supervised machine learning, to optimize the frequency for pDEP, so that live circulating cells are selected based on multiple biophysical metrics, including membrane physiology, cytoplasmic conductivity and cell size, which is not possible using deterministic lateral displacement that is solely based on cell size. Using typical drug-treated samples with low levels of live circulating cells (<3%), we present pDEP enrichment of the target subpopulation to ∼44% levels within 20 minutes, while rejecting >90% of dead cells. This strategy of utilizing single-cell impedance cytometry to guide the optimization of dielectrophoresis has implications for other complex biological samples. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the use of Ceiba pentandra (C. pentandra) as a glucose lowering agent and the attendant physiological changes in albino rats. Methods: Acute oral toxicity study of the extract was carried out by the administration of 10, 100, 1000, 1600, 2900, and 5000 mg/kg body weight of C. pentandra to rats in their respective groups. Twenty healthy albino rats weighing between 140 and 150 g were randomly allotted to five groups of four rats each. 100 mg/kg body weight of alloxan monohydrate was i.p. administered to rats and rats with blood glucose ≥ 200 mg/dL were considered diabetic. 5 mg/kg body weight of standard drug, 200 and 400 mg/kg body weight of ethanol extract of C. pentandra were orally administered to diabetic rats in their respective groups once daily for 12 days while the control groups received 0.1 mL of normal saline for the same period. The blood glucose was checked after every 4 days and the experiment was terminated on the 17th day. Results: The safe dose (LD50) of the extract was greater than 5000 mg/kg body weight. The extract treated groups exhibited a remarkable reduction in blood glucose [(87.72 ± 7.67) mg/dL for 200 mg/kg body weight dose and (86.33 ± 4.54) mg/dL for 400 mg/kg body weight dose] competitively with the normoglycemic group [(88.71 ± 4.56) mg/dL]. The body weight of the extract and standard drug treated groups appreciated significantly (P

Objective: To evaluate the acute and sub-acute toxicity of aqueous and methanol extracts of Nelsonia campestris (N. campestris) in rats. Methods: Acute oral toxicity study of aqueous and methanol extracts was carried out by administration of 10, 100, 1000, 1600, 2900 and 5000 mg/kg body weight of N. campestris extracts to rats in the respective groups. Sub-acute toxicity study was conducted by oral administration of the extracts at daily doses of 100, 300 and 600 mg/kg body weight to another group of rats for 28 days, while rats in the control group received 0.5 mL of normal saline. Results: The LD50 of the N. campestris extracts in rats was determined to be greater than 5000 mg/kg body weight. There was no significant difference (P > 0.05) between the test groups administered with aqueous and methanol extracts in relation to the control group for serum electrolytes (Na+, K+, Cl−, HCO3−), serum albumin, total and conjugated bilirubin. Similarly, mean organ-to-body weight ratio and all haematological parameters (white blood cell, red blood cell, mean cell volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, packed cell volume) evaluated were not significantly different (P > 0.05) from the control. There was a significant increase (P

Urinary excretion of two orally-administered non-metabolizable sugars, lactulose and mannitol, is a valuable marker for evaluating intestinal permeability. Usually this test involves a time consuming procedure of about 5 hour’s urine collection, which makes the test incompatible to some extent. As the results are expressed as the ratio of lactulose and mannitol recovered in urine within certain time, it may be possible to get similar result despite the reduced urine collection time of 2 hours. Moreover, different laboratories do the test by different methods, which make the results incomparable between laboratories. Here, we are also trying to find the correlation between results from most commonly used methods: HPAE-PAD and LC-MSMS. The lactulose: mannitol (LM) test was performed in a cohort of Bangladeshi infants considered at-risk for environmental enteropathy. 208 urine specimens from 104 (52 male and 52 female) infants were collected at 2 and 5 hours after LM solution administration and were tested for lactulose and mannitol by two different methods, one HPAE-PAD platform and another LC-MSMS platform. Median age of the children was 15.0 months (range 6.9 to 25.8 months) and their mean weight-for-age z-score was -0.92. A higher percentage of lactulose and mannitol recovery was found in 5 hours urine collection than in the corresponding 2 hours by both HPAE-PAD and LC-MSMS method, but when results were expressed as lactulose to mannitol ratio (LMR) there was no significant difference between 2 and 5 hours urine collection in both HPAE-PAD (P = 0.138) and LC-MSMS (P = 0.099) method. LMR based on 2 hours urine collection correlated well with LMR based on traditional 5 hours urine collection (Spearman’s correlation coefficient 0.578 and 0.604 respectively for HPAE-PAD and LC-MSMS). In future, LM test to assess intestinal permeability in children can be simplified by shortening the urine collection time from 5 hours to 2 hours. [ABSTRACT FROM AUTHOR]

Entamoeba histolytica causes intestinal disease in endemic settings throughout the world. Diagnosis of E. histolytica infection would be improved by the identification of biomarkers that are expressed by cysts of E. histolytica, but not by cysts of closely related commensal species of Entamoeba. Herein, we describe two novel monoclonal antibodies (1A4 and 1D3) produced against a spacer region of the E. histolytica Jacob2 lectin, an outer cyst wall protein. These reagents demonstrated no cross-reaction to E. dispar recombinant antigen and low picomolar molecular detection limits when paired in ELISA sandwich assays. In an immunofluorescence microscopy assay, the α-Jacob2 murine antibodies labeled cysts of three xenically cultured E. histolytica isolates but did not label cysts of three E. bangladeshi isolates. Monoclonal antibody 1A4 did not cross-react with xenic cultures of three E. dispar isolates, demonstrating specificity to E. histolytica, while monoclonal antibody 1D3 cross-reacted with two out of three E. dispar isolates. Both antibodies labeled cysts in formalin-fixed slides, a potential logistical advantage in some settings. The monoclonal antibody 1A4 was also used in an immunofluorescence microscopy assay with formalin-fixed stool specimens. Seven out of ten ELISA-positive stool specimens exhibited 1A4-labeled cyst-like objects, compared to one out of seven ELISA-negative specimens. These results demonstrate that antibodies generated against the flexible spacer of E. histolytica Jacob2 lectin recognize and bind to Jacob2 protein in whole cysts and are capable of differentiating Entamoeba species in fixed specimens. Thus, Jacob2 is a promising biomarker for use in diagnosing E. histolytica infection. [ABSTRACT FROM AUTHOR]

Background The combination of deworming and improved sanitation or hygiene may result in greater reductions in soil-transmitted helminth (STH) infection than any single intervention on its own. We measured STH prevalence in rural Bangladesh and assessed potential interactions among deworming, hygienic latrines, and household finished floors. Methodology We conducted a cross-sectional survey (n = 1,630) in 100 villages in rural Bangladesh to measure three exposures: self-reported deworming consumption in the past 6 months, access to a hygienic latrine, and household flooring material. We collected stool samples from children 1-4 years, 5-12 years, and women 15-49 years. We performed mini-FLOTAC on preserved stool samples to detect Ascaris lumbricoides, Enterobius vermicularis, hookworm, and Trichuris trichiura ova. Approximately one-third (32%) of all individuals and 40%of school-aged children had an STH infection. Less than 2%of the sample had moderate/heavy intensity infections. Deworming was associated with lower Ascaris prevalence (adjusted prevalence ratio (PR) = 0.53; 95% CI 0.40, 0.71), but there was no significant association with hookworm (PR = 0.93, 95%CI 0.60, 1.44) or Trichuris (PR = 0.90, 95%CI 0.74, 1.08). PRs for hygienic latrine access were 0.91 (95% CI 0.67,1.24), 0.73 (95%CI 0.43,1.24), and 1.03 (95%CI 0.84,1.27) for Ascaris, hookworm, and Trichuris, respectively. Finished floors were associated with lower Ascaris prevalence (PR = 0.56, 95%CI 0.32, 0.97) but not associated with hookworm (PR = 0.48 95%CI 0.16,1.45) or Trichuris (PR = 0.98, 95%CI 0.72,1.33). Across helminths and combinations of exposures, adjusted prevalence ratios for joint exposures were consistently more protective than those for individual exposures. Conclusions We found moderate STH prevalence in rural Bangladesh among children and women of childbearing age. This study is one of the first to examine independent and combined associations with deworming, sanitation, and hygiene. Our results suggest that coupling deworming with sanitation and flooring interventions may yield more sustained reductions in STH prevalence. [ABSTRACT FROM AUTHOR]

Background: The category B agent of bioterrorism, Entamoeba histolytica has a two-stage life cycle: an infective cyst stage, and an invasive trophozoite stage. Due to our inability to effectively induce encystation in vitro, our knowledge about the cyst form remains limited. This also hampers our ability to develop cyst-specific diagnostic tools. Aims: Three main aims were (i) to identify E. histolytica proteins in cyst samples, (ii) to enrich our knowledge about the cyst stage, and (iii) to identify candidate proteins to develop cyst-specific diagnostic tools. Methods: Cysts were purified from the stool of infected individuals using Percoll (gradient) purification. A highly sensitive LC-MS/MS mass spectrometer (Orbitrap) was used to identify cyst proteins. Results: A total of 417 non-redundant E. histolytica proteins were identified including 195 proteins that were never detected in trophozoite-derived proteomes or expressed sequence tag (EST) datasets, consistent with cyst specificity. Cyst-wall specific glycoproteins Jacob, Jessie and chitinase were positively identified. Antibodies produced against Jacob identified cysts in fecal specimens and have potential utility as a diagnostic reagent. Several protein kinases, small GTPase signaling molecules, DNA repair proteins, epigenetic regulators, and surface associated proteins were also identified. Proteins we identified are likely to be among the most abundant in excreted cysts, and therefore show promise as diagnostic targets. Major Conclusions: The proteome data generated here are a first for naturally-occurring E. histolytica cysts, and they provide important insights into the infectious cyst form. Additionally, numerous unique candidate proteins were identified which will aid the development of new diagnostic tools for identification of E. histolytica cysts. Author Summary: We used tandem mass spectrometry to identify E. histolytica cyst proteins in 5 cyst positive stool samples. We report the identification of 417 non-redundant E. histolytica proteins including 195 proteins that were not identified in existing trophozoite derived proteome or EST datasets, consistent with cyst specificity. Because the cysts were derived directly from patient samples with incomplete purification, a limited number of proteins were identified (N = 417) that probably represent only a partial proteome. Nevertheless, the study succeeded in identifying proteins that are likely to be abundant in the cyst stage of the parasite. Several of these proteins may play roles in E. histolytica stage conversion or cyst function. Proteins identified in this study may be useful markers for diagnostic detection of E. histolytica cysts. Overall, the data generated in this study promises to aid the understanding of the cyst stage of the parasite which is vital for disease transmission and pathogenesis in E. histolytica. [ABSTRACT FROM AUTHOR]

Cellular biophysical metrics exhibit systematic alterations during processes, such as metastasis and immune cell activation, which can be used to identify and separate live cell subpopulations for targeting drug screening. Image-based biophysical cytometry under extensional flows can accurately quantify cell deformability based on cell shape alterations but needs extensive image reconstruction, which limits its inline utilization to activate cell sorting. Impedance cytometry can measure these cell shape alterations based on electric field screening, while its frequency response offers functional information on cell viability and interior structure, which are difficult to discern by imaging. Furthermore, 1-D temporal impedance signal trains exhibit characteristic shapes that can be rapidly templated in near real-time to extract single-cell biophysical metrics to activate sorting. We present a multilayer perceptron neural network signal templating approach that utilizes raw impedance signals from cells under extensional flow, alongside its training with image metrics from corresponding cells to derive net electrical anisotropy metrics that quantify cell deformability over wide anisotropy ranges and with minimal errors from cell size distributions. Deformability and electrical physiology metrics are applied in conjunction on the same cell for multiparametric classification of live pancreatic cancer cells versus cancer associated fibroblasts using the support vector machine model., (© 2024 The Author(s). Small published by Wiley‐VCH GmbH.)

Introduction: MicroRNAs (miRNAs) are small, non-coding RNAs that post-transcriptionally regulate gene expression. Changes in miRNA expression have been reported in a number of intestinal diseases, in both tissue samples and readily accessible specimens like stools. Pathogenic infections, diet, toxins, and other environmental factors are believed to influence miRNA expression. However, modulation of miRNAs in humans is yet to be thoroughly investigated. In this study, we examined the expression levels of two human miRNAs (miRNA-122 and miRNA-21) in stool samples of a group of Bangladeshi children who had an altered/increased intestinal permeability (IIP). Methods: Stool samples were collected from children with IIP (L:M > 0.09) and normal intestinal permeability (NIP) (L:M ≤ 0.09). Quantitative PCR was performed to quantify the levels of miRNA-122 and miR-21 in stools. Commercial ELISA kits were used to measure gut inflammatory markers Calprotectin and REG1B. Serum samples were tested using Human Bio-Plex Pro Assays to quantify IL-1β, IL-2, IL-5, IL-10, IL-13, IFN-γ, and TNF-α. Total nucleic acid extracted from stool specimens were used to determine gut pathogens using TaqMan Array Card (TAC) system real-time polymerase chain reaction. Results: The expression levels of miRNA-122 (fold change 11.6; p < 0.001, 95% CI: 6.14-11.01) and miR-21 (fold change 10; p < 0.001, 95% CI: 5.05-10.78) in stool were upregulated in children with IIP than in children with normal intestinal permeability (NIP). Significant correlations were observed between stool levels of miR-122 and miR-21 and the inflammatory cytokines IL-1β, IL-2, IFN-γ, and TNF-α ( p < 0.05). Children with IIP were frequently infected with rotavirus, Campylobacter jejuni , Bacteroides fragilis , adenovirus, norovirus, astrovirus, and various Escherichia coli strains (ETEC&#95;STh, ETEC&#95;STp, EAEC&#95;aaiC, EAEC&#95;aatA) ( p < 0.001). miR-122 significantly correlated with the fecal inflammatory biomarkers REG1B ( p = 0.015) and Calprotectin ( p = 0.030), however miR-21 did not show any correlation with these fecal biomarkers., 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 © 2021 Rashid, Siddiqua, Hossain, Siddique, Kabir, Noor, Alam, Ahmed and Haque.)

Rapid point-of-care detection of enteric protozoa in diarrheal stool is desirable in clinical and research settings to efficiently determine the etiology of diarrhea. We analyzed the ability of the third-generation E. histolytica Quik Chek assay developed by Techlab to detect amebic antigens in fecal samples collected from independent study populations in South Africa and Bangladesh. We compared the performance of this recently released rapid test to that of the commercially available ProSpecT Entamoeba histolytica microplate assay from Remel and the E. histolytica II enzyme-linked immunosorbent assay (ELISA) from Techlab, using real-time and nested-PCR for Entamoeba species to resolve any discrepant results. After discrepant resolution, The E. histolytica Quik Chek assay exhibited sensitivity and specificity compared to the E. histolytica II ELISA of 98.0% (95% confidence interval [CI], 92.9% to 99.8%) and 100% (95% CI, 99.0% to 100%), respectively. Compared to the ProSpecT microplate assay, the E. histolytica Quik Chek (Quik Chek) assay exhibited 97.0% sensitivity (95% CI, 91.5% to 99.4%) and 100% specificity (95% CI, 99.0% to 100%). Our results indicate that the Quik Chek is a robust assay for the specific detection of E. histolytica trophozoites in unfixed frozen clinical stool samples., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Entamoeba histolytica, Giardia intestinalis, and Cryptosporidium spp. are not only three of the most important and common diarrhea-causing parasitic protozoa, but they often have similar clinical presentations. Microscopic diagnosis of these parasites is neither sensitive nor specific. Recently, more specific and sensitive alternative molecular methods (polymerase chain reaction [PCR] and antigen detection tests) have been introduced for all three of these parasitic infections. The use of these molecular diagnostic tests in routine diagnostic laboratories is still limited. In this study, we developed a multiplex real-time PCR assay for the simultaneous detection of E. histolytica, G. intestinalis, and Cryptosporidium spp. in one reaction using species-specific probes. This assay was evaluated using clinical specimens and was found to be quite sensitive and specific. The reagents used in this multiplex PCR assay can also be used for detection of these parasites individually. The use of this real-time PCR multiplex assay in developing countries at present will have limited scope for routine diagnosis because the cost will be high for a single test, although in the developed world, the test could see immediate application.