Efficacy of quercetin in ameliorating hypoxia-induced hematological and histopathological alterations in rohu Labeo rohita.

Hypoxia, a major issue in aquatic ecosystems, in special reference to climate change, and exacerbated by anthropogenic activities. It is causing slow growth, disease outbreaks, and mortality in finfish and shellfish. Therefore, adaptation to lowering oxygen levels through supplementation of herbs or their extracts in diets is imperative. In this study, hypoxia was simulated in controlled conditions with quercetin-enriched diets. Quercetin is a plant pigment (flavonol) possessing anti-oxidant property and is present in vegetables, leaves, seeds, pulses, and fruits. The experiment was conducted on rohu Labeo rohita, which is most widely cultured in India. There were four treatments including T1 (Normoxia: > 5 ppm dissolved oxygen; DO2), T2 (hypoxia: 3-4 ppm DO2), T3 (hypoxia + 50 mg quercetin/kg diet), and T4 (hypoxia + 100 mg quercetin/kg diet). The study was conducted for 30 days, and water quality was measured regularly. The results revealed that the hematological parameters were negatively affected. The tissue micro-architecture illustrated the impairment through degeneration of neurons in the brain, increased pigmentation as melanosis in the kidney, increased thickness of primary lamellae in the gills, and dilatations of sinusoids in the liver in hypoxia groups, while quercetin-enriched diets improved the hematological and histomorphological parameters. The results confirm the utility of hematological and histopathological tools as biomarkers and reflect the possible threats of hypoxia on fish. In conclusion, quercetin in diets appeared to show resistance towards chronic hypoxia by restoring the structure and functions of the vital organs towards normalcy and could be recommended as a potential ameliorative agent.

Biology and biotechnological aspect of sweet potato (Ipomoea batatas L.): a commercially important tuber crop.

MAIN CONCLUSION: This review highlights the economic importance of sweet potato and discusses new varieties, agronomic and cultivation practices, pest and disease control efforts, plant tissue culture protocols, and unexplored research areas involving this plant. Abstract Sweet potato is widely consumed in many countries around the world, including India, South Africa and China. Due to its valuable nutritional composition and highly beneficial bioactive compounds, sweet potato is considered a major tuber crop in India. Based on the volume of production, this plant ranks seventh in the world among all food crops. Sweet potato is considered a "Superfood" by the 'Centre for Science in the Public Interest' (CSPI), USA. This plant is mostly propagated through vegetative propagation using vine cuttings or tubers. However, this process is costly, labour-intensive, and comparatively slow. Conventional propagation methods are not able to supply sufficient disease-free planting materials to farmers to sustain steady tuber production. Therefore, there is an urgent need to use various biotechnological approaches, such as cell, tissue, and organ culture, for the large-scale production of healthy and disease-free planting material for commercial purposes throughout the year. In the last five decades, a number of tissue culture protocols have been developed for the production of in vitro plants through meristem culture, direct adventitious organogenesis, callus culture and somatic embryogenesis. Moreover, little research has been done on synthetic seed technology for the in vitro conservation and propagation of sweet potato. The current review comprehensively describes the biology, i.e., plant phenotypic description, vegetative growth, agronomy and cultivation, pests and diseases, varieties, and conventional methods of propagation, as well as biotechnological implementation, of this tuber crop. Furthermore, the explored and unexplored areas of research in sweet potato using biotechnological approaches have been reviewed.

Assessment of prevalence and characteristics of asthma-COPD overlap among patients with chronic airflow obstruction.

Given the paucity of research on asthma-chronic obstructive pulmonary disease (COPD) overlap (ACO) and the high prevalence of co-morbidities and healthcare utilization associated with it, the current study looked at the prevalence of ACO and its clinico-radiological phenotype in patients with chronic airflow obstruction. The study was conducted at a tertiary care hospital in North India. Patients over 40 with COPD or asthma were screened for inclusion in the ACO, asthma, and COPD groups. The ACO and COPD groups were further investigated. The clinical characteristics, lung functions, health-related quality of life, and radiological features of both groups were investigated and compared. ACO was discovered in 16.3% of patients with chronic airflow obstruction (asthma and COPD). The most commonly observed symptoms at presentation in the evaluated ACO patients (n=77) were shortness of breath, wheezing, cough, and expectoration (mean age at presentation: 57.9; mean duration of illness: 8.62 years). Exacerbation rates in ACO patients were significantly higher than in COPD patients (p<0.001). The ACO group had a significantly greater mean change in FEV1 post-bronchodilator in millilitres (ml) and percentage (379.61 ml and 37.72%) than the COPD group (p<0.001). The proportion of patients with emphysema was lower in the ACO group than in the COPD group (p<0.001). The ACO and COPD groups did not differ significantly in major airway wall thickness (p=0.3), but the COPD group had a significantly higher proportion of patients with vascular attenuation and distortion (p<0.001). Patients with COPD had a higher degree of hyperinflation, according to high resolution computed tomography (HRCT) indices. This study found that patients with ACO have a distinct phenotype in terms of clinical presentation and HRCT features. More research on the radiological features of ACO is required to identify the anatomical abnormalities involved in the disease's pathogenesis and to validate the radiological features of ACO.

Assessment of ASHA for knowledge, diagnosis and treatment on malaria in Mandla district of Madhya Pradesh as part of the malaria elimination demonstration project.

BACKGROUND: The role of Accredited Social Health Activist (ASHA) in the health care delivery services at the periphery level is crucial for achieving disease prevention, control and elimination goals. The objective of the study was to assess the knowledge, attitude, practices, priorities and capability of ASHA related to malaria diagnosis and treatment as part of the Malaria Elimination Demonstration Project in 1233 villages of district Mandla, Madhya Pradesh. METHODS: A cross sectional study was conducted using a fully structured, pre-tested interview schedule during June and July 2017 (before the field operations of MEDP were started). Two hundred twenty (17%) of the total ASHAs were selected for the interview from the 9 developmental blocks of Mandla district. RESULTS: Knowledge, Attitude and Practices (KAP) study revealed that most ASHAs knew that mosquitoes are the main agent for spread of malaria (97.7%). They mostly used Rapid Diagnostic Test (RDT) for diagnosis (91.8%). The majority (87.3%) correctly identified negative RDT result while only 15% and 10.5%, respectively, identified Plasmodium vivax and Plasmodium falciparum positive cases correctly. Further analysis showed that 85% ASHAs used chloroquine, 44.5% used artemisinin-based combination therapy (ACT), and 55.5% used primaquine for treatment of malaria. It was also found that only 38.2% ASHA gave PQ for 14 days in cases of P. vivax. At the time of the interview, 19.1% ASHAs did not have any RDTs for diagnosis and 47.7% reported not having ACT for treatment of P. falciparum malaria. CONCLUSIONS: This study has revealed that ASHAs in the test district were not adequately trained or stocked for malaria parasite species identification and treatment, which are the major components of malaria elimination programme. This study has, therefore, revealed a need for training ASHAs on testing by RDT and proper treatment regimen for P. vivax and P. falciparum.

New insights into the catalytic inactivity of mammary gland protein-40, a chitinase-like protein expressed during mammary gland involution.

MGP-40 is a mammary gland-specific glycoprotein which is expressed during involution and is an important marker for mammary gland apoptosis. It is an inactive chitinase-like protein belonging to Glycosyl Hydrolase family 18. The present study reports sequence characterization, tissue-specific expression analysis, production of recombinant MGP-40 and its mutant (A117D and L119E) in both E. coli and COS1 cells for their chitin-binding and chitinase activity analysis. The cDNA of buffalo MGP-40 was cloned and sequenced which corresponded to 1803 bp with an open reading frame of 1152 bp (361 aa), signal sequence of 63 bp (21 aa), 5' and 3' UTR of 144 bp and 507 bp, respectively. The 3' UTR analysis revealed potential sites for high level expression and stability during involution. The half-life of buffalo MGP-40 was found to be 11.7 h. MGP-40 was highly expressed in mammary gland followed by small intestine, spleen and mammary epithelial cells. The purified recombinant MGP-40 and its mutant expressed in E.coli were observed to bind chitin efficiently, however, no chitinase activity was observed. Further, chitinase activity was also not observed by expressing mutant recombinant MGP-40 in COS1 cells ruling out the possible role of post-translational modifications. Structure-based in-silico mutagenesis by FoldX algorithm showed a drastic decrease in overall fold stability which might be a possible reason for inability to recover its activity. Therefore, chitinase activity could not be restored in MGP-40 even after reverting back two critical residues in active site which may be due to detrimental effect of mutations on structural stability.

Functional characterization of Mammary Gland Protein-40, a chitinase-like glycoprotein expressed during mammary gland apoptosis.

MGP-40 is a chitinase-like protein which is over expressed during mammary gland involution. However, its physiological function in the mammary gland is poorly understood. In the present investigation, we have reported the functional significance of buffalo specific MGP-40 in the mammary gland by using an in vitro model of the buffalo mammary epithelial cell (BuMEC) line. MGP-40 was highly up regulated in BuMECs in serum starved condition as well as after treatment with prolactin suggesting its role in the stress response. Subsequently, to study the effect of MGP-40 on BuMECs, the cells were transfected with a mammalian expression construct of pCI neo harboring MGP-40 gene. It was observed that over expression of MGP-40 enhanced proliferation of BuMECs and protected the cells from apoptosis under serum free condition. In contrast, MGP-40 attenuated the mitogenic effect of insulin in BuMECs. Besides, over expression of the MGP-40 reduced dome formation, acinar polarization and casein synthesis in BuMECs in the presence of lactogenic hormones, it also induced Stat3 phosphorylation and epithelial to mesenchymal transition (EMT) -like features. Together, our data suggest that MGP-40 is involved in protection of BuMECs under stress conditions, inhibits cellular differentiation and induces EMT-like features. A schematic diagram depicting possible association of MGP-40 in various molecular pathways has been presented.

Establishing communication mechanism for malaria prevention in Baiga tribal villages in Baiga Chak area of Dindori district, Madhya Pradesh.

BACKGROUND & OBJECTIVES: Malaria is a serious public health concern in several parts of India, particularly in tribal areas of Madhya Pradesh (MP). Dindori district inhabitated by Baiga tribe, contributes about 15 per cent to the total malaria burden in MP. The tribal and other local inhabitants believe in magico-religious treatment of malaria and use modern health facilities only as second line of treatment. The present study was planned in the villages of one of the particularly vulnerable tribal group of MP, the Baigas. The objective of the study was to generate awareness and utilization of health services for malaria by establishing a communication strategy using local students and unemployed youths as agents of change. METHODS: The study was undertaken in 47 villages and the need based IEC (information, education and communication) intervention was evaluated within four months of initiation by adopting before and after with control design. For both baseline and resurvey the households covered each time were 2350. RESULTS: The baseline data generated revealed that around 53 per cent of the people in the study villages were aware of malaria. Among the non Baigas, 59 per cent were aware of malaria, while among the Baigas it was 49 per cent. IEC intervention could raise the level of awareness to malaria significantly with a net intervention effect of 23 per cent. The IEC intervention also improved the utilization of modern health services significantly. INTERPRETATION & CONCLUSIONS: The IEC strategy designed by using local children and youths was effective as the malaria was on decline in the study area. The same strategy with necessary modifications may be replicated in other areas pandemic for malaria.

Similar and divergent responses to salinity stress of jamun (Syzygium cumini L. Skeels) genotypes.

Background: Genetic variation for salt tolerance remains elusive in jamun (Syzygium cumini). Methods: Effects of gradually increased salinity (2.0-12.0 dS/m) were examined in 20 monoembryonic and 28 polyembryonic genotypes of jamun. Six genotypes were additionally assessed for understanding salt-induced changes in gas exchange attributes and antioxidant enzymes. Results: Salt-induced reductions in leaf, stem, root and plant dry mass (PDM) were relatively greater in mono- than in poly-embryonic types. Reductions in PDM relative to control implied more adverse impacts of salinity on genotypes CSJ-28, CSJ-31, CSJ-43 and CSJ-47 (mono) and CSJ-1, CSJ-24, CSJ-26 and CSJ-27 (poly). Comparably, some mono- (CSJ-5, CSJ-18) and poly-embryonic (CSJ-7, CSJ-8, CSJ-14, CSJ-19) genotypes exhibited least reductions in PDM following salt treatment. Most polyembryonic genotypes showed lower reductions in root than in shoot mass, indicating that they may be more adept at absorbing water and nutrients when exposed to salt. The majority of genotypes did not exhibit leaf tip burn and marginal scorch despite significant increases in Na+ and Cl-, suggesting that tissue tolerance existed for storing excess Na+ and Cl- in vacuoles. Jamun genotypes were likely more efficient in Cl- exclusion because leaf, stem and root Cl- levels were consistently lower than those of Na+ under salt treatment. Leaf K+ was particularly little affected in genotypes with high leaf Na+. Lack of discernible differences in leaf, stem and root Ca2+ and Mg2+ contents between control and salt treatments was likely due to their preferential uptake. Correlation analysis suggested that Na+ probably had a greater inhibitory effect on biomass in both mono- and poly-embryonic types. Discriminant analysis revealed that while stem and root Cl- probably accounted for shared responses, root Na+, leaf K+ and leaf Cl- explained divergent responses to salt stress of mono- and poly-embryonic types. Genotypes CSJ-18 and CSJ-19 seemed efficient in fending off oxidative damage caused by salt because of their stronger antioxidant defences. Conclusions: Polyembryonic genotypes CSJ-7, CSJ-8, CSJ-14 and CSJ-19, which showed least reductions in biomass even after prolonged exposure to salinity stress, may be used as salt-tolerant rootstocks. The biochemical and molecular underpinnings of tissue tolerance to excess Na+ and Cl- as well as preferential uptake of K+, Ca2+, and Mg2+ need to be elucidated.

Efficacy of Individualized Homeopathic Medicines in the Treatment of Hemorrhoids: Double-Blind, Randomized, Placebo-Controlled Trial.

Objectives: To investigate the efficacy and safety of individualized homeopathic medicines (IHMs) in treating hemorrhoids compared with placebo. Design: This is a double-blind, randomized (1:1), two parallel arms, placebo-controlled trial. Setting: The trial was conducted at the surgery outpatient department of the State National Homoeopathic Medical College and Hospital, Lucknow, Uttar Pradesh, India. Subjects: Patients were 140 women and men, aged between 18 and 65 years, with a diagnosis of primary hemorrhoids grades I-III for at least 3 months. Excluded were the patients with grade IV hemorrhoids, anal fissure, and fistula, hypertrophic anal papillae, inflammatory bowel disease, coagulation disorders, rectal malignancies, obstructed portal circulation, patients requiring immediate surgical intervention, and vulnerable samples. Interventions: Patients were randomized to Group 1 (n = 70; IHMs plus concomitant care; verum) and Group 2 (n = 70; placebos plus concomitant care; control). Outcome measures: Primary-the anorectal symptom severity and quality-of-life (ARSSQoL) questionnaire, and secondary-the EuroQol 5-dimensions 5-levels (EQ-5D-5L) questionnaire and EQ visual analogue scale (VAS); all of them were measured at baseline, and every month, up to 3 months. Results: Out of the 140 randomized patients, 122 were protocol compliant. Intention-to-treat sample (n = 140) was analyzed. The level of significance was set at p < 0.05 two tailed. Statistically significant between-group differences were elicited in the ARSSQoL total (Mann-Whitney U [MWU]: 1227.0, p < 0.001) and EQ-5D-5L VAS (MWU: 1228.0, p = 0.001) favoring homeopathy against placebos. Sulfur was the most frequently prescribed medicine. No harm or serious adverse events were reported from either of the groups. Conclusions: IHMs demonstrated superior results over placebo in the short-term treatment of hemorrhoids of grades I-III. The findings are promising, but need to be substantiated by further phase 3 trials. Clinical Trial Registration Number: CTRI/2020/03/024342.

Identification, validation and quantification of thymoquinone in conjunction with assessment of bioactive possessions and GC-MS profiling of pharmaceutically valuable crop Nigella (Nigella sativa L.) varieties.

Background: Plants have been pivotal in traditional and modern medicine globally, with historical evidence supporting their therapeutic applications. Nigella (Nigella sativa L.) is an annual herbaceous plant of the Ranunculaceae family and is cultivated in the Middle East, Eastern Europe, and Western and Central Asia. The medicinal use of plants dates back thousands of years, documented in ancient writings from various civilizations. Alkaloids, phenolics, saponins, flavonoids, terpenoids, anthraquinones, and tannins found in plants exhibit antioxidant, immunomodulatory, anti-inflammatory, anticancer, antibacterial, and antidiabetic activities. Methodology: This study specifically examines the pharmacological potential of Nigella sativa L., emphasizing thymoquinone-a compound with diverse nutraceutical benefits. The extraction, characterization, and quantification of thymoquinone, alongside other physicochemical parameters, were carried out using ethanol through Soxhlet extraction procedures on five nigella varieties. HPLC analysis was performed to determine the maximum accumulation of thymoquinone in the released variety of the plant and the chemical composition of the seed oil isolated from Nigella sativa L., varieties utilized in the study was determined through GC-MS analysis. Results: The research revealed that the Ajmer nigella-20 variety stands out, exhibiting elevated levels of thymoquinone (0.20 ± 0.07%), antioxidants (76.18 ± 1.78%), and substantial quantities of total phenols (31.85 ± 0.97 mg GAEg-1 seed) and flavonoids (8.150 ± 0.360 mg QE 100 g-1 seed) compared to other varieties. The GC-MS profiling showed the presence of 11 major compounds in the studied varieties, with p-cymene, longifolene, and myristic acid identified as the major chemical compounds present in the oil. Conclusion: The observed variations among Nigella varieties indicate the Ajmer nigella-20 variety as particularly promising for thymoquinone and bioactive compound extraction. This study underscores Nigella's potential as a source of pharmacologically active compounds, highlighting the need for further exploration in therapeutic applications.

Optimization of Oral Posaconazole Step Down Therapy in Management of Rhino-Orbital-Cerebral Mucormycosis (ROCM): Outcome of an Institutional Protocol.

All cases of ROCM received antifungal for a prolong period in step down phase. There is no definite guidelines regarding duration. We have formulated and followed an institutional protocol with good results. To determine the outcome of an institutional protocol and required optimum duration of oral posaconazole therapy to cure ROCM in step down phase, 30 ROCM patients were followed up in step down phase. Oral posaconazole tablet was given for minimum 3, 4 & 1/2 and 6 months to stage-II, III & IV cases respectively, with a provision of extension for another 6 weeks in suspected active diseases. Nasal endoscopy, MRI, histopathology /culture of suspected tissue and hematologic investigations were done routinely to identify the residual active disease or recurrences at earliest. Outcome of this protocol proved excellent as all cases of ROCM were cured. Oral posaconazole step down therapy for a duration according to the stages of disease cured 18 cases (60%) of ROCM. Fourteen patients required extended duration of therapy as per provision of our treatment protocol. Only one patient needed re-debridement. The cases with deformities needed extended period of posaconazole therapy. No one needed posaconazole therapy for more than 7and1/2 months.

Optimization of biogas potential using kinetic models, response surface methodology, and instrumental evidence for biodegradation of tannery fleshings during anaerobic digestion.

The optimization of the batch size experiment was run for a hydraulic retention time of 45 days using proteolytic enzyme pretreatment. The highest amounts of biogas were produced in comparison to conventional BDS (25:75), which is not processed with enzymes, and there was an increase in the biogas generation of 13.9 and 18.57%. The kinetic models show the goodness of fit between 0.993 and 0.998 and the correlation coefficient's value domain was [-1, 1] from a statistical perspective. The Box-Behnken design was carried out using the response surface methodology at different levels of independent parameters to optimize the process. Different instruments were evaluated to determine the chemical structure change and the contamination of the different treatments and the raw sample of tannery fleshings was determined. Thermogravimetric analysis was conducted to determine the loss of weight on thermal degradation. The Fourier transform infrared spectrometry was carried out to determine the different functional groups, such as -OH, -CH, -NH, and C-O, present in the samples of tannery fleshings. Scanning electron microscopy and energy dispersive X-ray analysis were carried out to determine the morphological alterations in the substrate, digestate, enzyme-pretreated fleshings, and the chemical composition of samples.

Development of B cell epitopes-based enzyme linked immune sorbent assay for detection of bovine anti-Mullerian hormone.

The present study aimed to generate antibodies against predicted B cell epitopic peptides encoding bAMH for developing different ELISA models. Sandwich ELISA was determined to be an excellent technique for assessing bAMH in bovine plasma based on sensitivity tests. The assay's specificity, sensitivity, inter- and intra-assay CV, recovery %, Lower limit of quantification (LLOQ), and Upper limit of quantification (ULOQ) were determined. The test was selective since it did not bind to AMH-related growth and differentiation factors (LH and FSH) or non-related components (BSA, progesterone). The intra-assay CV was 5.67%, 3.12%, 4.94%, 3.61% and 4.27% for 72.44, 183.11, 368.24, 522.24 and 732.25 pg/ml AMH levels, respectively. At the same time, the inter-assay CV was 8.77%, 7.87%, 4.53%, 5.76% and 6.70% for 79.30, 161.27, 356.30, 569.33 and 798.19 pg/ml AMH levels, respectively. The average (Mean ± SEM) recovery percentages were 88-100%. LLOQ was 5 pg/ml and ULOQ at 50 µg/ml (CV < 20%). In conclusion, we developed a new highly sensitive ELISA against bAMH using epitope specific antibodies.

Managed pollination is a much better way of increasing productivity and essential oil content of dill seeds crop.

Dill seeds (Anethum graveolens L.) is the most valuable medicinal seed spice crop of Apiaceae. It bears small yellow flowers in the form of umbels. Being a cross-pollinated crop, floral visitors play vital role in pollination and seed sets. Hence, the present study was conducted at the ICAR-National Research Centre on Seed Spices, Ajmer (Rajasthan), India to discover the pollinator's community, foraging behaviour and abundance of most frequent pollinators and different modes of pollination on seed yield and quality of this seed spice crop. The insect visitors community of dill seeds was composed of 28 insect species belonging to 14 families of 6 orders. Most of floral visitors started their foraging activity at 8.00 h, reached peak activity between 12.00 and 14.00 h and their activity ceased at 18.00 h. Apis florea, A. dorsata, A. mellifera, solitary bee, Halictus sp. and two unidentified species of Hymenoptera; Episyrphus balteatus (DeGeer), Episyrphus sp., Eristalis sp and two other Musca species of Diptera were identified as potential and regular floral visitors of dill seeds. The highest seed yield of 1505.63 kg/ha was recorded in the treated plots provided with only 10% jaggery solution and was at par with the open pollination. A lower seed yield of 1432.5 kg/ha was recorded in plots pollinated only with A. mellifera inside insect cages. Open pollination with 10% jaggery solution spray increased the seed yield of dill seed crop by 57%, one-thousand seed test weight by 96% and the essential oil content by 27% over control plots. These results show that managed pollination is a much better way to enhance yields and quality of dill seed crop than other treatments including only honeybee-based pollination.

Diversity in Child Mortality and Life Expectancy at Birth Among Major Tribes in Selected States of India.

OBJECTIVE: To provide tribe- specific child mortality rates and health indicators from selected states in India. METHODS: We used Census 2011 data and Coale Demney methodology to estimate the infant mortality rate (IMR), under-five mortality rate (U5MR) and expectation of life at birth (LEB) for 123 tribes of selected states of India. RESULTS: The estimated IMR and U5MR were higher in scheduled tribe population compared to respective state's total population. The IMR varied from 124 in the Birhore tribe of Chhattisgarh and Jharkhand, and the Bharias of Madhya Pradesh to 48 per 1000 live births in the Gamit tribe of Maharashtra. Similarly, the U5MR varied from the highest (203) in the Birhore tribe of Chhattisgarh to the lowest (57/1000 live births) in the Gamit tribe. The LEB varied from 72 years in the Gamit tribe of Maharashtra to 51 years in the Birhore tribe of Chhattisgarh. The study reveals that tribes have gross variation in child mortality rates and there is pressing need to prioritize tribe-specific action plans to improve their health indicators.

Transcriptional Repression of MFG-E8 Causes Disturbance in the Homeostasis of Cell Cycle Through DOCK/ZP4/STAT Signaling in Buffalo Mammary Epithelial Cells.

The mammary gland is a unique apocrine gland made up of a branching network of ducts that end in alveoli. It is an ideal system to study the molecular mechanisms associated with cell proliferation, differentiation, and oncogenesis. MFG-E8, also known as Lactadherin, is a vital glycoprotein related to the milk fat globule membrane and initially identified to get secreted in bovine milk. Our previous report suggests that a high level of MFG-E8 is indicative of high milk yield in dairy animals. Here, we showed that MFG-E8 controls the cell growth and morphology of epithelial cells through a network of regulatory transcription factors. To understand the comprehensive action, we downregulated its expression in MECs by MFG-E8 specific shRNA. We generated a knockdown proteome profile of differentially expressed proteins through a quantitative iTRAQ experiment on a high-resolution mass spectrometer (Q-TOF). The downregulation of MFG-E8 resulted in reduced phagocytosis and cell migration ability, whereas it also leads to more lifespan to knockdown vis-a-vis healthy cells, which is confirmed through BrdU, MTT, and Caspase 3/7. The bioinformatics analysis revealed that MFG-E8 knockdown perturbs a large number of intracellular signaling, eventually leading to cessation in cell growth. Based on the directed network analysis, we found that MFG-E8 is activated by CX3CL1, TP63, and CSF2 and leads to the activation of SOCS3 and CCL2 for the regulation of cell proliferation. We further proved that the depletion of MFG-E8 resulted in activated cytoskeletal remodeling by MFG-E8 knockdown, which results in the activation of three independent pathways ZP4/JAK-STAT5, DOCK1/STAT3, and PIP3/AKT/mTOR. Overall, this study suggests that MFG-E8 expression in mammary epithelial cells is an indication of intracellular deterioration in cell health. To date, to the best of our knowledge, this is the first study that explores the downstream targets of MFG-E8 involved in the regulation of mammary epithelial cell health.

Direct detection of OXA-48-like carbapenemase variants with and without co-expression of an extended-spectrum ß-lactamase from bacterial cell lysates using mass spectrometry.

INTRODUCTION: Antibiotic-resistant Gram-negative bacteria are of a growing concern globally, especially those producing enzymes conferring resistance. OXA-48-like carbapenemases hydrolyze most ß-lactam antibiotics, with typically low-level hydrolysis of carbapenems, but have limited effect on broad-spectrum cephalosporins. These are frequently co-expressed with extended spectrum ß-lactamases, especially CTX-M-15, which typically shows high level resistance to broad-spectrum cephalosporins, yet is carbapenem susceptible. The combined resistance profile makes the need for successful detection of these specific resistance determinants imperative for effective antibiotic therapy. OBJECTIVES: The objective of this study is to detect and identify OXA-48-like and CTX-M-15 enzymes using mass spectrometry, and to subsequently develop a method for detection of both enzyme types in combination with liquid chromatography. METHODS: Cells grown in either broth or on agar were harvested, lysed, and, in some cases buffer-exchanged. Lysates produced from bacterial cells were separated and analyzed via liquid chromatography with mass spectrometry (LC-MS) and tandem mass spectrometry (LC-MS/MS). RESULTS: The intact proteins of OXA-48, OXA-181, and OXA-232 (collectively OXA-48-like herein) and CTX-M-15 were characterized and detected. Acceptance criteria based on sequence-informative fragments from each protein group were established as confirmatory markers for the presence of the protein(s). A total of 25 isolates were successfully tested for OXA-48 like (2), CTX-M-15 (3), or expression of both (7) enzymes. Thirteen isolates served as negative controls. CONCLUSIONS: Here we present a method for the direct and independent detection of both OXA-48-like carbapenemases and CTX-M-15 ß-lactamases using LC-MS/MS. The added sensitivity of MS/MS allows for simultaneous detection of at least two co-eluting, co-isolated and co-fragmented proteins from a single mass spectrum.

Rapid MRSA detection via tandem mass spectrometry of the intact 80 kDa PBP2a resistance protein.

Treatment of antibiotic-resistant infections is dependent on the detection of specific bacterial genes or proteins in clinical assays. Identification of methicillin-resistant Staphylococcus aureus (MRSA) is often accomplished through the detection of penicillin-binding protein 2a (PBP2a). With greater dependence on mass spectrometry (MS)-based bacterial identification, complementary efforts to detect resistance have been hindered by the complexity of those proteins responsible. Initial characterization of PBP2a indicates the presence of glycan modifications. To simplify detection, we demonstrate a proof-of-concept tandem MS approach involving the generation of N-terminal PBP2a peptide-like fragments and detection of unique product ions during top-down proteomic sample analyses. This approach was implemented for two PBP2a variants, PBP2amecA and PBP2amecC, and was accurate across a representative panel of MRSA strains with different genetic backgrounds. Additionally, PBP2amecA was successfully detected from clinical isolates using a five-minute liquid chromatographic separation and implementation of this MS detection strategy. Our results highlight the capability of direct MS-based resistance marker detection and potential advantages for implementing these approaches in clinical diagnostics.

Identification of critical areas and evaluation of best management practices using SWAT for sustainable watershed management.

Identification of critical erosion-prone areas and selection of best management practices (BMPs) for watersheds are necessary to control their degradation by reducing sediment yields. The current research assesses and proposes a combination of potential BMPs for the Baitarani catchment in India using the Soil and Water Assessment Tool (SWAT). After the successful calibration and validation of the SWAT model developed for this catchment, the model was applied to evaluate the efficacy of eight agricultural and structural management practices and their combinations (three scenarios) for controlling sediment yields at watershed and sub-watershed levels as well as to assess the impacts of combined BMPs on water balance components. A combination of BMPs was found more effective in reducing sediment yields than individual BMPs. Comparative evaluation revealed that structural BMPs (0.66-70%) are better than agricultural BMPs (2-7%) in minimizing sediment yields at watershed level. The costly measures like grade and streambank stabilization structures can reduce the sediment yield up to 70% at the watershed level. The modeling results of the impacts of different combinations of BMPs (three scenarios) indicated that if all the eight BMPs are implemented, the reduction of sediment yields is increased by 76% and 80% at sub-watershed and watershed levels, respectively compared to the Base Scenario. Based on funds availability, a suitable combination of BMPs can be adopted by the concerned decision-makers to effectively reduce sediment yields in the study area. Further, the simulation results of BMPs impacts on water balance components revealed that the annual average surface runoff reduces by 4-14% in the three scenarios, while aquifer recharge (6.8-8.7%), baseflow (8-10.5%), and percolation (1.2-3.9%) increase due to implementation of BMPs. Overall, the findings of this study are very useful for ensuring sustainable management of land and other resources at a catchment scale.

Implantation associated changes in expression profile of indoleamine-2, 3-dioxygenase 1, Th1-Th2 cytokines and interferon-stimulated genes on neutrophils and peripheral blood mononuclear cells of crossbred cows.

Effective bidirectional communication between the embryo and dam improves the reproductive efficiency of dairy cows. Possible role of immunosuppressive indolamine-2, 3-dioxygenase 1 (IDO1) enzyme in the regulation of maternal systemic cytokine balance/shift during early pregnancy establishment along with various interferon-stimulated genes (ISGs) expression in neutrophils and peripheral blood mononuclear cell (PBMCs) were investigated in crossbred cows. Blood was collected on days 0 i.e. day of Artificial Insemination (AI), 10, 18 and 36 post-AI followed by isolation of neutrophils and PBMCs for gene expression study of IDO1, anti-inflammatory cytokines (IL-4, IL-10 and TGFß1), pro-inflammatory cytokines (IFNγ and TNFα) and ISGs (ISG15, MX1, MX2, OAS1) in pregnant and non-pregnant cows. Cows were grouped as pregnant and non-pregnant after pregnancy confirmation by non-return to heat, ultrasonography, per rectal examination along with progesterone and IFNτ assay. Significantly (P < 0.05) higher relative mRNA expression of IDO1 and anti-inflammatory cytokines on days 10 and 18 post-AI were observed in both neutrophils and PBMCs of pregnant cows. Pregnant cows showed significantly (P < 0.05) higher mRNA transcripts of IFNγ and TNFα genes on days 18 post-AI in both neutrophils and PBMCs. Expression of ISGs was higher (P < 0.05) on day 10th and 18th post AI in both the neutrophils and PBMCs of pregnant cows. The study indicates that systemic immune regulation by IDO1 (through cytokine shift) and ISGs in peripheral immune cells are essential for the establishment of pregnancy and may be targeted in future as biomarkers for pregnancy diagnosis.