Molecular detection and phylogenetic analysis of ovine herpesvirus-2 in subclinical infections of cattle and sheep.

Ovine herpesvirus-2 (OvHV-2) is the causative agent of malignant catarrhal fever (MCF), a serious and often fatal disease that affects cattle and other ruminants. This study aimed to investigate the molecular epidemiology and genetic diversity of OvHV-2 strains circulating in sheep and cattle populations in the Jammu and Kashmir region of India. Screening of 150 sheep and 57 cattle blood samples revealed the presence of the OvHV-2 polymerase (pol) gene in 8.6% of sheep, 10% of apparently healthy cattle, and 29.7% of cattle exhibiting MCF-like symptoms. The full-length glycoprotein B (gB) gene (2800 bp) and an 875 bp internal fragment were successfully amplified, cloned, and sequenced from pol-positive samples. Comparative sequence analysis of the deduced gB amino acid sequences identified seven substitutions at positions 278, 341, 390, 440, 468, 539, and 566 compared to reference strains. Phylogenetic analysis based on the gB nucleotide sequences clustered the OvHV-2 strains from this study within the Indian clade, distinct from strains reported in the UK and US. These findings provide insights into the genetic diversity of OvHV-2 strains circulating in Jammu and Kashmir, with the identified mutations potentially influencing virus-host interactions. Further investigations into the functional implications of these mutations are warranted to understand their role in viral pathogenesis and tropism.

Evidence of novel Treponema phylotypes implicated in contagious ovine digital dermatitis and association of treponemes with major lameness causing foot pathogens.

In this study 269 swabs collected from 254 ovine foot lesions and 15 apparently healthy ovine feet were screened by PCR for the presence of major lameness causing foot pathogens viz. Treponema species, D. nodosus, F. necrophorum and T. pyogenes with the presumption that ovine foot lesion positive for Treponema species alone or in association with other three pathogens were categorized as contagious ovine digital dermatitis (CODD). While samples positive for D. nodosus alone or its combination with F. necrophorum and T. pyogenes were considered as footrot (FR) and samples in which F. necrophorum or T. pyogenes was found either alone or in combination were considered as interdigital dermatitis (ID). The overall occurrence of Treponema sp. in ovine foot lesions was 48.0%, and ranged from 33 to 58%. In Treponema positive samples D. nodosus, F. necrophorum and T. pyogenes were present in 34 (27.4%), 66 (54.4%) and 84 (68.5%) in contrast to Treponema negative samples in which these were present in 15 (11.1%), 20 (14.12%) and 17 (12.6%) samples, respectively. The data signifies that Treponema sp. are significantly associated with these foot pathogens and their different combinations with Treponema sp. influence the severity of CODD lesion. The identification of Treponema phylotypes was done by sequencing the 16S rRNA gene fragment of ten representative samples. Out of ten sequences, four (Trep-2, Trep-4, Trep-7 and Trep-10) were identical to Treponema sp. phylotype 1 (PT1) that belongs to phylogroup T. refringens-like, one sequence (Trep-1) was genetically close (90% sequence homology) to Treponema brennaborense while five sequences (Trep-3, Trep-5, Trep-6, Trep-8 and Trep-9) matched with uncultured bacterium clones of treponemes forming separate monophyletic group in phylogenetic tree and could represent new digital dermatitis phylogroup presently containing five ovine specific phylotypes. This is the first report on the presence of Treponema phylotypes other than three digital dermatitis (DD) Treponema phylogroups viz. T. phagedenis-like, T. medium/T. vincentii-like, and T. pedis-like that are frequently detected in CODD lesions. Metagenomic analysis of two representative samples revealed the abundance of genus Treponema in CODD lesion while this genus was absent in swab collected from clinically healthy foot suggesting that it might play primary role in producing CODD. These findings may further aid in understanding the etiopathogenesis of CODD and could help to develop appropriate treatment and mitigation strategies to combat the disease.

An inexpensive phytoremediation system for treating 50,000 L/day of sewage.

The paper describes the setting up and long-term continuous operation of the first real-life, pilot scale, sewage treatment plant based on the recently patented phytoremediation technology, trademarked as SHEFROL®. The unit was about three times cheaper to install, operate and maintain than the least expensive of the other wetland-based technologies presently in vogue. Its semi-permanent version is 30 times cheaper. Monitoring of flow rates and levels of treatment intermittently over a 3 year course of continuous operation indicated the constancy and robustness of the reactor in treating total solids, suspended solids, chemical oxygen demand, biological oxygen demand, total Kjeldahl nitrogen, and soluble phosphorous to the average extents of 94, 84, 79, 70, 62 and 28% respectively. Earlier experience with bench-scale SHEFROL® units has indicated that removal of metals like Cu, Ni, Co, Zn, and Mn also takes place to the extent of 25-45% in these systems. These primary, secondary, and tertiary treatments occurred in a single unit process with no necessity of any pumping, aeration, or recycling. Models based on artificial intelligence were developed which enable forecasting the reactor performance in terms of secondary and tertiary treatment, respectively.

The paper describes the setting-up, long-term (3 years) performance evaluation at pilot scale, and modeling of a recently patented novel and unprecedented phytoremediation-based sewage treatment technology. The system is seen to be much more robust, versatile, space efficient, and economical, than existing macrophyte-based sewage treatment systems.

Land-use systems regulate carbon geochemistry in the temperate Himalayas, India.

The Himalayan ecosystem is critical for ecological security and environmental sustainability. However, continuous deforestation is posing a serious threat to Himalayan sustainability. Changing land-use systems exert a tenacious impact on soil carbon (C) dynamics and regulate C emissions from Himalayan ecosystem. Therefore, this study was conducted to determine the changes in different C pools and associated soil properties under diverse land-use systems, viz. natural forest, natural grassland, maize field converted from the forest, plantation, and paddy field of temperate Himalaya in the surface (0-20 cm) and subsurface (20-40 cm) soils. The highest total organic carbon (24.24 g kg-1) and Walkley-black carbon contents (18.23 g kg-1), total organic carbon (45.88 Mg ha-1), and Walkley-black carbon stocks (34.50 Mg ha-1) were recorded in natural forest in surface soil (0-20 cm depth), while soil under paddy field had least total organic carbon (36.45 Mg ha-1) and Walkley-black carbon stocks (27.40 Mg ha-1) in surface soil (0-20 cm depth). The conversion of natural forest into paddy land results in 47.36% C losses. Among the cultivated land-use system, minimum C losses (29.0%) from different pools over natural forest system were reported under maize-filed converted from forest system. Land conversion causes more C losses (21.0%) in surface soil (0-20 cm depth) as compared to subsurface soil. Furthermore, conversion of forest land into paddy fields increased soil pH by 5.9% and reduced total nitrogen contents and microbial population by 28.0% and 7.0%, respectively. However, the intensity of total nitrogen and microbial population reduction was the lowest under maize fields converted from the forest system. The study suggested that the conversion of natural forest to agricultural land must be discouraged in the temperate Himalayan region. However, to feed the growing population, converted forest land can be brought under conservation effective maize-based systems to reduce C loss from the intensive land use and contribute to soil quality improvements and climate change mitigation.

Delineating binding potential, stability of Sulforaphane-N-acetyl-cysteine in the active site of histone deacetylase 2 and testing its cytotoxicity against distinct cancer lines through stringent molecular dynamics, DFT and cell-based assays.

Histone deacetylase 2 (HDAC2), an isozyme of Class I HDACs has potent imputations in actuating neurodegenerative signaling. Currently, there are sizeable therapeutic disquiets with the use of synthetic histone deacetylase inhibitors in disease management. This strongly suggests the unfulfilled medical necessity of plant substitutes for therapeutic intervention. Sulforaphane-N-acetyl-cysteine (SFN-N-acetylcysteine or SFN-NAC), a sulforaphane metabolite has shown significantly worthier activity against HDACs under in vitro conditions. However, the atomistic studies of SFN-NAC against HDAC2 are currently lacking. Thus, the present study employed a hybrid strategy including extra-precision (XP) grid-based flexible molecular docking, molecular mechanics generalized born surface area (MM-GBSA), e-Pharmacophores method, and molecular dynamics simulation for exploring the binding strengh, mode of interaction, e-Pharmacophoric features, and stability of SFN-NAC towards HDAC2. Further, the globally acknowledged density functional theory (DFT) study was performed on SFN-NAC and entinostat individually in complex state with HDAC2. Apart from this, these inhibitors were tested against three distinct cancer cell models and one transformed cell line for cytotoxic activity. Moreover, double mutant of HDAC2 was generated and the binding orientation and interaction of SFN-NAC was scrutinized in this state. On the whole, this study unbosomed and explained the comparatively higher binding affinity of entinostat for HDAC2 and its wide spectrum cytotoxicity than SFN-NAC.

Identification of Immunodominant Outer Membrane Proteins of Fusobacterium necrophorum from Severe Ovine Footrot By MALDI-TOF Mass Spectrometry.

The aim of this study was to identify the immunodominant outer membrane proteins (OMPs) of Fusobacterium necrophorum from sheep affected with severe foot-rot. The OMP profile of ovine strains of F. necrophorum has not been well studied. We analyzed the OMP profile of the most frequent lktA variant JKS-F3 of F. necrophorum associated with severe ovine foot-rot with lesion score 4 in order to identify its major immunodominant OMPs. Electrophoretic separations of extracted OMPs showed a number of spots in two-dimensional electrophoretic gels. Two immunoreactive proteins of size around 43 kDa were identified through western blotting using hyperimmune sera raised in rabbits. These two immunogenic OMPs were analyzed by Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF/MS) which revealed that these two OMPs of lktA variant JKS-F3 of F. necrophorum showed 46 and 42 percent protein sequence coverage and scores of 125 and 114, respectively, with the reported 43 kDa outer membrane protein of F. necrophorum strain H05, a putative porin having properties similar to pore-forming proteins of anaerobic Gram-negative bacteria. These identified immunogenic OMPs will contribute to our understanding of the pathogenic role played by this organism in ovine foot-rot and could be exploited to devise an effective control strategy through development of an OMP-based recombinant vaccine to mitigate foot-rot in sheep and goats.

Genotyping and antibiotic resistance patterns of Campylobacter fetus subsp.venerealis from cattle farms in India.

Bovine genital campylobacteriosis caused by Campylobacter fetus subsp. venerealis (Cfv) is of considerable economic importance to the cattle industry worldwide. Cfv causes syndrome of temporary infertility in female cattle, early embryonic mortality, aberrant oestrus cycles, delayed conception, abortions and poor calving rates. In the present study, a total of 200 samples obtained from vaginal swabs, cervicovaginal mucous (CVM), preputial washes and semen straws were investigated that were obtained from organized cattle farm of MLRI, Manasbal and unorganized sectors. Out of a total of 200 samples, 49 (47·57%) vaginal swabs, 1 (3·33%) preputial wash and 8 (25%) carried out CVM samples were positive for Cfv, whereas none of the semen straws were positive for Cfv. A total of eleven isolates of Cfv were recovered. PFGE (Pulse field gel electrophoresis) analysis revealed four different pulsotypes (I-IV) circulating in the screened farms. A common pulsotype circulating among farms could not be established. Insertion element (ISCfe1), a 233 bp amplicon of Cfv, was sequenced and the sequence was deposited in GenBank (accession no: MK475662).

Determination of prevalence, serological diversity, and virulence of Dichelobacter nodosus in ovine footrot with identification of its predominant serotype as a potential vaccine candidate in J&K, India.

The aim of this study was to determine the prevalence, serological diversity, and virulence of Dichelobacter nodosus in footrot lesions of sheep and identification of its predominant serotype as a potential vaccine candidate. The overall prevalence of footrot in sheep was 16.19%, and ranged from 13.69 to 19.71%, respectively. A total of 759 flocks with 22,698 sheep were investigated for footrot and 2374 clinical samples were collected from naturally infected sheep exhibiting footrot lesions. Of the 2374 samples collected, 1446 (60.90%) were positive for D. nodosus by polymerase chain reaction (PCR). These positive samples when subjected to serogroup-specific multiplex PCR, 1337 (92.46%) samples carried serogroup B, 247 (17.08%) possessed serogroup E, 86 (5.94%) serogroup I, and one (0.069%) serogroup G of D. nodosus. While mixed infection of serogroups B and E was detected in 127 (8.78%), B and I in 46 (3.18%) and B, E, and I in 26 (1.79%) samples, respectively. The serogroup B of D. nodosus was the predominant (92.47%) serogroup affecting sheep population with footrot followed by serogroup E (19.91%) and serogroup I (4.57%), respectively. Virulent status of D. nodosus strains were confirmed by presence of virulence-specific integrase A (intA) gene and the production of thermostable proteases. The intA gene was detected in 709 (72.79%) samples while gelatin gel test carried out on 246 representative isolates all positive for intA gene produced thermostable proteases, confirming their virulence nature. The PCR-restriction fragment length polymorphism (PCR-RFLP) of whole fimA gene of serogroup B revealed the predominance of serotype B5 (82.97%) of serogroup B. This information suggests that serotype B5 is the predominant serotype of D. nodosus associated with severe footrot lesions in sheep in Jammu & Kashmir (J&K), India. Hence, this serotype can be a potential vaccine candidate for the effective control and treatment of ovine footrot.

Elevated oxidative DNA damage in patients with coronary artery disease and its association with oxidative stress biomarkers.

OBJECTIVE: The objective of the present study was to evaluate oxidative DNA damage in peripheral blood leukocytes (PBLs) of patients with coronary artery disease (CAD) and to explore the relationship of oxidised purine and pyrimidine with oxidative stress. METHODS: The study participants (n = 100) included 50 patients and unrelated 50 age-, sex- and population-subgroup (Jat Sikhs)-matched healthy controls. Oxidative DNA damage using the modified enzymatic comet in PBLs, and malondialdehyde (MDA) levels, total oxidant status (TOS) and total antioxidant status (TAS) in blood serum samples using spectrophotometric methods was determined. RESULTS: The basal DNA damage of percent tail DNA (T-DNA%) was increased as were tail moment (TM) and olive tail moment (OTM). Oxidative DNA damage in terms of oxidised purines and oxidised pyrimidines was also significantly (p < .001) elevated in patients. Rather the advanced stages of CAD, unstable angina and acute myocardial infarction had significantly more basal and oxidative DNA damage (p < .05) compared to stable angina. MDA levels (p < .01) and TOS (p < .001) were increased significantly in patients with significant (p < .001) decrease in TAS. There was positive correlation of oxidised purines (T-DNA% r = 0.399, p = .004; TM r = 0.623, p = .001; OTM r = 0.456, p= .001) and of total oxidative damage (TM r = 0.515, p = .001; OTM r = 0.463, p = .001) with disease severity, and, with TOS (r = 0.279, p = .050) and negative with TAS (r = -0.341, p = .015). Multiple linear regression analysis revealed TOS and disease severity as independent predictors of oxidative DNA damage. CONCLUSIONS: There was significant increase in oxidative DNA damage and oxidative stress in CAD patients compared to levels in healthy controls.

CYP2D6 (C2850T, G1846A, C100T) polymorphisms, haplotypes and MDR analysis in predicting coronary artery disease risk in north-west Indian population: A case-control study.

AIMS: The present study was aimed to evaluate the association of C2850T, G1846A and C100T polymorphisms of the CYP2D6 with coronary artery disease (CAD) in North-West Indian population. METHODS: In this case-control study, 200 patients with CAD and 200 age-, gender- and ethnicity-matched healthy controls were genotyped for C2850T, G1846A and C100T polymorphisms of CYP2D6 using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. RESULTS: Genotype and allele distributions of C2850T and G1846A polymorphisms of the CYP2D6 were significantly different between cases and controls (p = 0.038, p = 0.021; p = 0.048, p = 0.012, respectively) whereas the distribution of genotype and allele for C100T polymorphism did not differ significantly (p = 0.098, p = 0.117, respectively). The 2850T and1846A variants were significantly associated with the increased risk of developing CAD, as observed from the odds ratios for the 2850 T/T and 1846 G/A genotypes (OR: 2.44, 95% CI: 1.99-4.99, p = 0.015 and OR: 1.62, 95% CI: 1.02-2.56, p = 0.041, respectively). Moreover, the recessive model in C2850T and the dominant model in G1846A are the best fit inheritance models to predict the susceptible gene effects (OR: 2. 07, 95% CI: 1.05-4.08, p = 0.031 and OR: 1.70, 95% CI: 1.10-2.62, p = 0.016, respectively). On gender stratification, these associations were observed only in females in addition to the C/T genotype of C2850T (OR: 2.52, 95% CI: 1.42-4.38, p = 0.001) and C100T (OR: 3.18, 95% CI: 1.52-6.67, p = 0.002). Furthermore, it is also observed that the CAT (OR: 2. 61, 95% CI: 1.07-6.34, p = 0.035) and TAC (OR: 15. 22, 95% CI: 1.97-117.58, p = 0.009) are high-risk haplotypes for CAD in the total group, whereas, the TGC (OR: 1. 64, 95% CI: 1.02-2.62, p = 0.042) and CAT (OR: 4. 21, 95% CI: 1.12-15.59, p = 0.035) haplotypes provide gender-specific risk in females. CONCLUSIONS: Our results indicate significant association of C2850T, G1846A and C100T polymorphisms of CYP2D6 with CAD especially in females of North-West Indian population.

The detection and prevalence of leukotoxin gene variant strains of Fusobacterium necrophorum in footrot lesions of sheep in Kashmir, India.

The objective of this study was to determine the prevalence and identification of leukotoxin gene, lktA, variant strains of Fusobacterium necrophorum in the footrot lesions of sheep. The detection of F. necrophorum was carried out by PCR targeting the lktA gene fragment and identification of lktA variant strains was done by PCR-single stranded conformational polymorphism (PCR-SSCP) and gene sequencing. Of the 450 swabs collected from footrot lesions of sheep, 117 were lktA-positive for F. necrophorum. Of the 50 swabs collected from apparently asymptomatic sheep, only one was lktA-positive for F. necrophorum. The overall prevalence of F. necrophorum in footrot affected sheep in Kashmir valley was 26%, and ranged from 20 to 34.8%, respectively. PCR-SSCP of lktA gene fragment analysis revealed three lktA variants, designated as JKS-F1/F2/F3, while two samples (1.7%) showed multiple lktA variant strains of F. necrophorum in a single footrot-affected sheep hoof. This appears to be the first report on the presence of more than one lktA variant of F. necrophorum in a footrot lesion of sheep. The JKS-F3 lktA variant was the most frequent (75.4%), followed by JKS-F2 (14.4%) and JKS-F1 (8.4%), respectively. Among the three lktA variants identified, JKS-F3 was detected in 74 (86.0%) samples from severe footrot affected sheep with a lesion score of 4. The data suggest that JKS-F3 is the predominant lktA variant of F. necrophorum and is associated with severe footrot in sheep. Hence, JKS-F3 may be a significant variant contributing to the severity and duration of the disease in sheep.

Postpartum Acute Kidney Injury: Experience of a Tertiary Care Center.

Pregnancy-related-acute kidney injury (PR-AKI) had decreased from 40% to 20% in 1960 to <10% in recent series, mostly due to meticulous antenatal management. Postpartum-AKI (PP-AKI) resulting from late obstetric complications has become more apparent after improvement in antenatal care and legalization of medical termination of pregnancy. Women with renal injury in peripartum period admitted to our hospital over a period of 2 years (April 2013 to May 2015) were studied. Of 713 patients of AKI admitted, 61 had PR-AKI with an incidence of 4.27%. Out of the 61 patients, 28 had PP-AKI with an incidence of 1.96%. The mean age of patients with PP-AKI was 26.10 ± 4.3 years. Sepsis was the most common cause accounting for 11 (39.28%) cases followed by postpartum hemorrhage (PPH) in 7 (25%) cases. Renal biopsy was done in nine patients, out of whom four were having cortical necrosis. Patients having diffuse cortical necrosis remained dialysis-dependent. High contribution of sepsis and PPH to PP-AKI in our setting makes it an ideal target for rectification. Protocolized peripartum monitoring and standard clinical practices of asepsis will go long way in decreasing the incidence of PP-AKI and maternal morbidity in our valley.

Erythropoietin monotherapy in perinatal asphyxia with moderate to severe encephalopathy: a randomized placebo-controlled trial.

OBJECTIVE: Erythropoietin (EPO) is neuroprotective after asphyxia in animal studies. The efficacy and safety of EPO monotherapy in term neonates with hypoxic ischemic encephalopathy (HIE) is uncertain. STUDY DESIGN: Hundred term neonates with moderate or severe HIE were randomized by random permuted block algorithm to receive either EPO 500 U kg-1 per dose in 2 ml saline intravenously (50 neonates) on alternate days for a total of five doses with the first dose given by 6 h of age (treatment group) or 2 ml of normal saline (50 neonates) similarly for a total of five doses (placebo group) in a double-blind study. No hypothermia was given. The primary outcome was combined end point of death or moderate or severe disability at mean age of 19 months (s.d., 0.61). RESULTS: Death or moderate or severe disability occurred in 40% of neonates in the treatment group vs 70% in the placebo group (risk ratio, 0.57; 95% confidence interval (CI) 0.38 to 0.85; P=0.003). Death occurred in 16% of patients in both the groups (risk ratio, 1.0; 95% CI 0.33 to 2.9; P=0.61). The risk of cerebral palsy was lower among survivors in the treatment group (risk ratio, 0.52; 95% CI 0.25 to 1.03; P=0.04) and lesser number of babies were on anticonvulsants at assessment (risk ratio, 0.47; 95% CI 0.20 to 1.01; P=0.03). Neonatal brain magnetic resonance imaging showed more abnormalities in the placebo group (relative risk, 0.66; 95% CI 0.42 to 1.03; P=0.04)). Improvement in other neurological outcomes was not significant. CONCLUSION: EPO monotherapy reduces the risk of death or disability in term neonates with moderate or severe encephalopathy.

Glutathione S-transferase P1 gene polymorphisms and susceptibility to coronary artery disease in a subgroup of north Indian population.

The present study aimed to investigate the association of g.313A>G and g.341C>T polymorphisms of GSTP1 with coronary artery disease (CAD) in a subgroup of north Indian population. In the present case-control study, CAD patients (n = 200) and age-matched, sex-matched and ethnicity-matched healthy controls (n = 200) were genotyped for polymorphisms in GSTP1 using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Genotype distribution of g.313A>Gand g.341C>T polymorphisms of GSTP1 gene was significantly different between cases and controls (P = 0.005 and 0.024, respectively). Binary logistic regression analysis showed significant association of A/G (odds ratio (OR): 1.6, 95% CI: 1.08-2.49, P = 0.020) and G/G (OR: 3.1, 95% CI: 1.41-6.71, P = 0.005) genotypes of GSTP1 g.313A>G, and C/T (OR: 5.8, 95% CI: 1.26-26.34, P = 0.024) genotype of GSTP1 g.341C>T with CAD. The A/G and G/G genotypes of g.313A>G and C/T genotype of g.341C>T conferred 6.5-fold increased risk for CAD (OR: 6.5, 95% CI: 1.37-31.27, P = 0.018).Moreover, the recessive model of GSTP1 g.313A>G is the best fit inheritance model to predict the susceptible gene effect (OR: 2.3, 95% CI: 1.11-4.92, P = 0.020). In conclusion, statistically significant associations of GSTP1 g.313A>G (A/G, G/G) and g.341C>T (C/T) genotypes with CAD were observed.

Targeting leukemic side population cells by isatin derivatives of nicotinic acid amide.

Side population (SP) cells mediate chemoresistance in leukemia. However, chemical inhibition approach to target SP cells has been poorly studied. Herein, we report the discovery of isatin derivatives of nicotinic acid amide as potent side population cell inhibitors. The selected derivatives showed superior potency over the reference drug verapamil. Furthermore, the treatment increased chemosensitivity and inhibited the cell proliferation on three different leukemic cell lines, K562, THP-1 and U937, suggesting that both SP and the bulk of leukemic cells are affected. Moreover, treatment with the most potent compound Nic9 reduced the expression of ABCG2, demonstrating that side population inhibition effect of the target derivatives is at least via ABCG2 inhibition. Importantly, the target derivatives induced erythrocyte/dendritic differentiation to leukemic cells mainly through Musashi/Numb pathway modulation.

Cationic antimicrobial peptide, magainin down-regulates secretion of pro-inflammatory cytokines by early placental cytotrophoblasts.

BACKGROUND: Human placental villous cytotrophoblasts exhibit relative externalization of negatively charged moieties to the outer leaflet of the plasma membrane during the time of syncytialization rendering their reactivity to positively charged cationic antimicrobial peptides (CAMPs) during the window of implantation and early placentation. Vaginal administration of a synthetic CAMP, Ala(8,13,18)-magainin II amide (AMA) inhibited blastocyst implantation and early placentation in monkeys. Furthermore, the administration of AMA resulted in significant inhibition of cell differentiation, enhancement in apoptosis and loss of viability in first trimester placental villous cytotrophoblasts in primary culture. The present study examines the effect of in vitro application of different doses (0, 1, 10, 100, 1000 ng/ml) of AMA on the secreted cytokine profiles of cytotrophoblasts obtained from placental villi samples (n = 13) collected during 8-9 weeks of gestation and grown on three-dimensional collagen matrix in vitro. METHODS: A panel of forty-eight (48) cytokines in conditioned medium was analysed using multiplex immunoassays technique. Further, the steady state transcript levels of four cytokines (CCL4, CCL5, IL1B, IL6), the concentrations of which were affected by AMA in the isolated cytotrophoblasts, as well as, two cytokines (IL1A and TNF) which were not affected by AMA were estimated. Input list of cytokines secreted by cytotrophoblasts and showing differential secretion in response to AMA were used in enrichment analysis for the generation of biological networks. RESULTS: Placental cytotrophoblasts secreted 27 cytokines, 13 of which are affected by AMA in vitro with significantly decreased secretion of CCLs-2, 3, 4, 5, CXCLs-1 and 8, FGF2 and MCSF and that of IL1B, IL6 and MIF, and increased secretion of IL16 and IL-2RA. Of the above cytokines showing differential secretion, only IL-2RA, IL16 and MIF showed significant correspondence in the steady state expression of their respective transcript levels. Post-hoc Enrichment analysis revealed Toll-like receptor (TLR) mediated pathways were the top-scored target pathways that were affected by AMA. CONCLUSIONS: Administration of a CAMP causes shift in the balance of immune-inflammatory responses involving downstream pathways of TLRs in cytotrophoblast function. Further verification of functions of placental trophoblasts on administration of CAMP with pregnancy outcome is necessary.

Performance of rapid influenza diagnostic tests (QuickVue) for influenza A and B Infection in India.

BACKGROUND: Rapid point-of-care (POC) tests provide an economical alternative for rapid diagnosis and treatment of influenza, especially in public health emergency situations. OBJECTIVES: To test the performance of a rapid influenza diagnostic test, QuickVue (Quidel) as a POC test against a real-time polymerase chain reaction (RT-PCR) assay for detection of influenza A and B in a developing country setting. STUDY DESIGN: In a prospective observational design, 600 patients with influenza-like illness (ILI) or with severe acute respiratory illness (SARI) who were referred to the Influenza Clinic of a tertiary care hospital in Srinagar, India from September 2012 to April 2013, were enrolled for diagnostic testing for influenza using QuickVue or RT-PCR. All influenza A-positive patients by RT-PCR were further subtyped using primers and probes for A/H1pdm09 and A/H3. RESULTS: Of the 600 patients, 186 tested positive for influenza A or B by RT-PCR (90 A/H1N1pdm09, 7 A/H3 and 89 influenza B), whereas only 43 tested positive for influenza (influenza A=22 and influenza B=21) by QuickVue. Thus, the sensitivity of the QuickVue was only 23% (95% confidence interval, CI: 17.3-29.8) and specificity was 100% (95% CI: 99.1-100) with a positive predictive value (PPV) of 100% (95% CI 91.8-100) and a negative predictive value (NPV) of 74.3% (95% CI: 70.5-77.9) as compared to RT-PCR. CONCLUSIONS: The high specificity of QuickVue suggest that this POC test can be a useful tool for patient management or triaging during a public health crisis but a low sensitivity suggests that a negative test result need to be further tested using RT-PCR.

Group A rotavirus and bacterial agents associated with diarrhoea-induced hospitalisations in children below 5 years of age in Jammu.

Out of 210 faecal samples collected from children below 5 years attending different hospitals in Jammu and exhibiting clinical signs of diarrhoea, 41.9% samples were found positive for group A rotavirus by RNA-PAGE. Escherichia coli isolated in the study belonged to nine serogroups, out of which O69 was most frequent, being present in 12.38% samples. E. coli serogroups well recognised as enteropathogens viz. O69, O20 and O153 were present in 27.6% samples. Other bacterial pathogens associated with diarrhoea were present in 8.09% samples, out of which Shigella spp. was found in 4.76% samples followed by Salmonella spp. (2.38%) and Pseudomonas spp. (0.95%).

Non-specificity of primers used for PCR based serogrouping of Dichelobacter nodosus and identification of a novel D. nodosus strain.

The present study records the first case of non-specificity of typing primers developed by Dhungyel et al. A strain of Dichelobacter nodosus (JKS-20G) isolated from ovine footrot in Kashmir, India, showed specificity for serogroup C and G primers. The fimA sequence of the strain turned out to be closer to serogroup G than C. The nucleotide sequence showed maximum homology of 92% with that of serotype G1 strain 238 and 95% with partial sequence available for serotype G2 strain VCS 1004. However, the deduced amino acid sequence of the fimbrial subunit gene of JKS-20G differed from strain 238 by 16 amino acids and by four amino acids from that of partial sequence of strain VCS 1004. This variation indicates towards declaring this isolate as a new serotype (G3) but just insufficient to classify this into a new serogroup. Some of the amino acid substitutions were located within three hypervariable regions a characteristic of different serogroups. However, to ascertain whether this isolate deserves a new serotype status, there is a need to go for antigenic characterisation of this isolate using the tube and cross tube agglutination test.