Production performance, haematological parameters, serum biochemistry, and expression of HSP-70 in broiler chickens fed dietary ascorbic acid during heat stress.

An experiment was carried out to assess the efficacy of supplemental ascorbic acid (AA) on broiler chicken production performance, blood haematological profile, biochemical profile, and carcass traits under heat stress conditions. A total of 192-day-old broiler chicks were divided into four groups, each with six replicates of eight each (4 × 6 × 8). Four corn-based dietary treatments were formulated: T1 (control diet), T2 (T1 + AA at 200 mg/kg), T3 (T1 + AA at 400 mg/kg), and T4 (T1 + AA at 600 mg/kg) for a period of 42 days. Despite the high temperature and humidity, the 600 mg AA supplemental groups (T4) gained significantly (P ≤ 0.05) more body weight and had a higher feed intake and better feed conversion ratio (FCR) than the control group (T1). After 28 days of feeding the three AA-supplemented diets, antibody titres (humoral immune response) were significantly higher (P ≤ 0.05). The response to intradermally injected phyto-haemagglutinin (PHA-P), an index of the in vivo cell-mediated immune response, was found to be increased (P ≤ 0.05) in the 400 and 600 mg AA-supplemented groups after 35 days. Higher levels of AA (T4) supplementation significantly (P ≤ 0.05) improved haematological values such as haemoglobin (Hb), total erythrocyte count (TEC), total leukocyte count (TLC), and differential leukocyte count (DLC), heterophil:lymphocyte (H:L) in comparison to the control group (T1). The supplemented group improved the serum biochemical profile of the birds, with an increase (P ≤ 0.05) in total serum protein, albumin, and globulin and a decrease in serum cholesterol and corticosterone levels in the T4 group compared to the control group. Heat shock protein-70 (HSP-70) was gradually elevated after increasing the ascorbic acid concentration (P ≤ 0.05) at 14 and 21 days. As a result, it can be concluded that supplementing ascorbic acid at 600 mg/kg is beneficial for improving the performance, immunity, and blood haematological biochemical profile and upregulating the HSP-70 gene of broiler chickens under heat stress conditions.

Salazinic Acid and Norlobaridone from the Lichen Hypotrachyna cirrhata: Antioxidant Activity, α-Glucosidase Inhibitory and Molecular Docking Studies.

The present study was intended for the identification of secondary metabolites in acetone extract of the lichen Hypotrachyna cirrhata using UPLC-ESI-QToF-MS/MS and the detection of bioactive compounds. This study led to the identification of 22 metabolites based on their MS/MS spectra, accurate molecular masses, molecular formula from a comparison of the literature database (DNP), and fragmentation patterns. In addition, potent antioxidant and α-glucosidase inhibitory potentials of acetone extract of H. cirrhata motivated us to isolate 10 metabolites, which were characterized as salazinic acid (11), norlobaridone (12), atranorin (13), lecanoric acid (14), lichesterinic acid (15), protolichesterinic acid (16), methyl hematommate (17), iso-rhizonic acid (18), atranol (19), and methylatratate (20) based on their spectral data. All these isolates were assessed for their free radicals scavenging, radical-induced DNA damage, and intestinal α-glucosidase inhibitory activities. The results indicated that norlobaridone (12), lecanoric acid (14), methyl hematommate (17), and atranol (19) showed potent antioxidant activity, while depsidones (salazinic acid (11), norlobaridone (12)) and a monophenolic compound (iso-rhizonic acid, (18)) displayed significant intestinal α-glucosidase inhibitory activities (p < 0.001), which is comparable to standard acarbose. These results were further correlated with molecular docking studies, which indicated that the alkyl chain of norlobaridione (12) is hooked into the finger-like cavity of the allosteric pocket; moreover, it also established Van der Waals interactions with hydrophobic residues of the allosteric pocket. Thus, the potency of norlobaridone to inhibit α-glucosidase enzyme might be associated with its allosteric binding. Also, MM-GBSA (Molecular Mechanics-Generalized Born Surface Area) binding free energies of salazinic acid (11) and norlobaridone (12) were superior to acarbose and may have contributed to their high activity compared to acarbose.

Comprehensive Lichenometabolomic Exploration of Ramalina conduplicans Vain Using UPLC-Q-ToF-MS/MS: An Identification of Free Radical Scavenging and Anti-Hyperglycemic Constituents.

In this study, we propose ultra-performance liquid chromatography coupled with quadrupole/time-of-flight mass spectrometry (UPLC-QToF-MS/MS)-guided metabolite isolation as a choice analytical approach to the ongoing structure−activity investigations of chemical isolates from the edible lichen, Ramalina conduplicans Vain. This strategy led to the isolation and identification of a new depside (5) along with 13 known compounds (1−4, 6−14), most of which being newly described in this lichen species. The structures of the isolates were established by detailed analysis of their spectral data (IR, NMR, and Mass). The acetone extract was further analyzed by UPLC-Q-ToF-MS/MS in a negative ionization mode, which facilitated the identification and confirmation of 18 compounds based on their fragmentation patterns. The antioxidant capacities of the lichen acetone extract (AE) and isolates were measured by tracking DPPH and ABTS free radical scavenging activities. Most isolates displayed marked radical scavenging activities against ABTS while moderate activities were observed against DPPH radical scavenging. Except for atranol (14), oxidative DNA damage was limited by all the tested compounds, with a marked protection for the novel isolated compound (5), as previously noted for the acetone extract (p < 0.001). Furthermore, compound (4) and acetone extract (AE) have inhibited intestinal α-glucosidase enzyme significantly (p < 0.01). Although some phytochemical studies were already performed on this lichen, this study provided new insights into the isolation and identification of bioactive compounds, illustrating interest in future novel analytical techniques.

Comparing the efficiency of different Escherichia coli strains in producing recombinant capsid protein of porcine circovirus type 2.

The present study was aimed at comparing different E. coli strains in expressing the capsid protein of Porcine Circovirus 2 (PCV2). Full length capsid protein could be expressed only in Rosetta-gami 2 (DE3) pLysS strain using pET32b (+) vector. This confirmed that only those strains which possess tRNAs for rare codons can express the full length capsid protein. Purification of full length capsid protein could not be achieved even after several attempts using native and denaturing conditions. Subsequently, an attempt was made for expression of N-terminal truncated capsid protein using the same expression system. Truncated capsid protein was successfully expressed, purified and characterized by western blotting. The truncated capsid protein was also shown to be efficacious in testing serum samples using an optimized indirect ELISA, wherein a diagnostic sensitivity of 88.89% and specificity of 90.82% was obtained as compared to commercially available GreenSpring® porcine circovirus (PCV2) ELISA test kit. Thus, the expressed truncated capsid protein appears to be a promising diagnostic agent for PCV2. The comparative analysis suggests that cluster of arginine residues at N-terminal of capsid protein not only affects its expression in some E. coli strains but also its purification by Ni-NTA chromatography, when expressed as a histidine tagged fusion protein.

Stability of live attenuated classical swine fever cell culture vaccine virus in liquid form for developing an oral vaccine.

Classical swine fever (CSF) is an important viral disease of pigs and controlled by vaccination. Unorganised backyard and wild pigs are difficult to vaccinate by needle vaccination. Here we formulated liquid vaccines using an Indian CSF cell culture vaccine virus and four stabilisers and evaluated their stability at 4 °C, 25 °C and 37 °C up to 24 h for use as oral vaccine. The stabilisers were Lactalbumin hydrolysate-Trehalose, Lactalbumin hydrolysate-Trehalose-Gelatin, Lactalbumin hydrolysate-Lactose-Sucrose and Lactalbumin hydrolysate-Sucrose. The liquid vaccines, with or without stabilisers, were stable at 4 °C up to 24 h, whereas, a drop of one log10 titre was observed at 25 °C during the same period. At 37 °C, the virus titre diminished by only one log10 with the Lactalbumin hydrolysate-Trehalose (LT) stabiliser up to 24 h compared to two log10 losses in virus titre with other stabilisers and virus control. We therefore conclude that for developing a CSF oral vaccine, the vaccine virus in liquid form can be used directly during the winter, whereas for developing the oral vaccine for summer, the LT stabiliser would provide maximum stability to the virus to withstand the warm temperature while maintaining adequate therapeutic titre for inducing a protective immune response.

Molecular Characterization of Mx1 Gene in Native Indian Breeds of Chicken.

The genetic polymorphism of Mx1 gene was explored in Indian chicken breeds. PCR-RFLP analysis in 102 bp fragment of partial intron 13 and partial exon 14 of Mx1 gene revealed two genotypes viz. RS and SS with two alleles viz. R and S both in Naked Neck and Tellicherry breeds of chicken. The homozygous genotype RR was not identified. When deduced amino acid sequences were compared, the asparagine amino acid was found to be substituted in "R" allele for serine in "S" allele. PCR-SSCP analysis of 284 bp fragment in 5'-UTR and partial promoter region revealed three genotypes viz. CC, CG, and CH with three different alleles viz. C, G, and H in Naked Neck breed of chicken and five genotypes viz. DI, JK, KK, KL, and KM with six different alleles viz. D, I, J, K, L, and M in Tellicherry breed of chicken. The homozygous genotypes viz. GG and HH in Naked Neck and DD, II, JJ, LL, and MM in Tellicherry chicken was not identified. The nucleotide substitution rate estimated to be in the range of 0.004-0.011. The identified genetic variation can be helpful for better insight to disease resistance property of the Mx1 gene.

Expression kinetics of ISG15, IRF3, IFNγ, IL10, IL2 and IL4 genes vis-a-vis virus shedding, tissue tropism and antibody dynamics in PPRV vaccinated, challenged, infected sheep and goats.

Here, we studied the in vivo expression of Th1 (IL2 and IFN gamma) and Th2 (IL4 and IL10) - cytokines and antiviral molecules - IRF3 and ISG15 in peripheral blood mononuclear cells in relation to antigen and antibody dynamics under Peste des petits ruminants virus (PPRV) vaccination, infection and challenge in both sheep and goats. Vaccinated goats were seropositive by 9 days post vaccination (dpv) while in sheep idiosyncratic response was observed between 9 and 14 dpv for different animals. Expression of PPRV N gene was not detected in PBMCs of vaccinated and vaccinated challenged groups of both species, but was detected in unvaccinated infected PBMCs at 9 and 14 days post infection. The higher viral load at 9 dpi coincided with the peak clinical signs of the disease. The peak in viral replication at 9 dpi correlated with significant expression of antiviral molecules IRF3, ISG15 and IFN gamma in both the species. With the progression of disease, the decrease in N gene expression also correlated with the decrease in expression of IRF3, ISG15 and IFN gamma. In the unvaccinated infected animals ISG15, IRF3, IFN gamma and IL10 expression was higher than vaccinated animals. The IFN gamma expression predominated over IL4 in both vaccinated and infected animals with the infected exhibiting a stronger Th1 response. The persistent upregulation of this antiviral molecular signature - ISG15 and IRF3 even after 2 weeks post vaccination, presumably reflects the ongoing stimulation of innate immune cells.

Genome sequencing of an Indian peste des petits ruminants virus isolate, Izatnagar/94, and its implications for virus diversity, divergence and phylogeography.

Peste des petits ruminants is an important transboundary disease infecting small ruminants. Genome or gene sequence analysis enriches our knowledge about the evolution and transboundary nature of the causative agent of this disease, peste des petits ruminants virus (PPRV). Although analysis using whole genome sequences of pathogens leads to more precise phylogenetic relationships, when compared to individual genes or partial sequences, there is still a need to identify specific genes/genomic regions that can provide evolutionary assessments consistent with those predicted with full-length genome sequences. Here the virulent Izatnagar/94 PPRV isolate was assembled and compared to all available complete genome sequences (currently in the NCBI database) to estimate nucleotide diversity and to deduce evolutionary relationships between genes/genomic regions and the full length genomes. Our aim was to identify the preferred candidate gene for use as a phylogenetic marker, as well as to predict divergence time and explore PPRV phylogeography. Among all the PPRV genes, the H gene was identified to be the most diverse with the highest evolutionary relationship with the full genome sequences. Hence it is considered as the most preferred candidate gene for phylogenetic study with 93% identity set as a nucleotide cutoff. A whole genome nucleotide sequence cutoff value of 94% permitted specific differentiation of PPRV lineages. All the isolates examined in the study were found to have a most recent common ancestor in the late 19th or in the early 20th century with high posterior probability values. The Bayesian skyline plot revealed a decrease in genetic diversity among lineage IV isolates since the start of the vaccination program and the network analysis localized the ancestry of PPRV to Africa.

Synthesis and evaluation of anticancer and antiobesity activity of 1-ethoxy carbonyl-3,5-bis (3'-indolyl methylene)-4-pyperidone analogs.

A series of eleven novel bisindole derivatives were synthesized and screened for anticancer and antiobesity potentials in in vitro mode. The reaction of 1-ethoxy carbonyl 4-pyperidone 1a with indole-3-carboxaldehyde 1b in presence of catalytic amount of piperidine gave 2 which was N-alkylated with different benzyl halides in the presence of potassium carbonate to afford compounds 3a-3k in quantitative yields. Among the compounds tested for anticancer activity against different human cancer cell lines, 3f significantly inhibited HepG2 cell line (IC50 7.33 µM) when compared with standard doxorubicin (IC50 10.15 µM). Compounds 3e (IC50 2.75 µM), 3f (IC50 4.21 µM) and 3i (IC50 15.98 µM) showed better activity than the standard curcumin (IC50 23.54 µM) against A549 cell line. Also, among the synthesized compounds, 3g (IC50 14.89 µM), 3c (IC50 56.41 µM) and 3i (IC50 30.88 µM) have potentially inhibited enzyme lipase when compared to standard Orlistat (IC50 62.25 µM). In in silico docking assays, piperidones 3e, 3f, 3i, 3c and 3a showed higher binding affinity towards anti-cancer target of A549 (3e: -11.1, 3f: -10.3, 3c: -11.3, 3i: -11.2 kcal/mol), HepG2 (3f: -10.5 kcal/mol), HeLa (3d: -10.0 kcal/mol) and SKOV3 (3f: -8.4 kcal/mol) cell lines better than standard drug doxorubicin. Docking to lipase protein for compounds 3i, 3g and 3c showed scores of -11.1, -10.7 and -10.5 kcal/mol when compared to that of standard drug Orlistat with -6.9 kcal/mol.

Protective immune-response of aluminium hydroxide gel adjuvanted phage lysate of Brucella abortus S19 in mice against direct virulent challenge with B. abortus 544.

The prophylactic efficacies of plain and alum adsorbed lysate were evaluated by direct virulent challenge in mice model. A recently isolated brucellaphage 'ϕLd' was used for generation of lysates. Twenty four h incubated Brucella abortus S19 broth cultures standardized to contain approximately 10(8) CFU/ml were found suitable for generation of lysates. Three lysate batches produced through separate cycles did not show any significant variation with respect to protein and polysaccharide contents, endotoxin level and phage counts, indicating that compositionally stable lysate preparations can be generated through an optimized production process. Three polypeptides of ∼16, 19 and 23 kDa could be identified as immuno-dominant antigens of the lysate which induced both humoral and cell-mediated immune responses in a dose dependent manner. Results of efficacy evaluation trial confirmed dose-dependent protective potencies of lysate preparation. The lysate with an antigenic dose of 0.52 µg protein and 60 µg CHO adsorbed on aluminium gel (0.1 percent aluminium concentration) exhibited the highest protective potency which was greater than that induced by standard S19 vaccine. Phage lysate methodology provides a very viable option through which an improved immunizing preparation with all desirable traits can be developed against brucellosis, and integrated with immunization programmes in a more efficient manner.

The cage-free egg sector: perspectives of Indian poultry producers.

India is ranked as the 2nd largest egg producer in the world. Despite the prevalence of backyard poultry (free range), a majority of the commercial egg-laying hens in the country are still housed in battery cages. There is a global shift toward cage-free eggs, due to regulations and increased demand from conscious consumers and food corporations. However, there are very few commercial cage-free facilities in India to meet this demand. The aim of this study was to undertake a needs-assessment survey of Indian egg producers on cage-free production, and understand what support is needed to build the capacities of the cage-free egg production sector to develop it into a viable and sustainable alternative to battery cage eggs. The results showed that nearly all producers agreed on the need for additional support in shifting to, and operating in, the cage-free sector. This included support in the form of financial assistance, technical training, and promotion of the cage-free sector. The results of this study highlight the pressing need for government and private support, in the absence of which cage-free producers are compelled to compete with battery cage poultry producers on prices, which will result in increased losses and failure of the sector, since they have not yet achieved economies of scale.

Adequate Levels of Zinc and Copper on Turkey Poult Performance, Immunological Response, and Skeletal Health.

In this study, the impact of varying dietary zinc (Zn) and copper (Cu) levels on the growth, immunological response, and skeletal health of developing turkey poults was investigated. For 0-4 and 5-8 weeks of age, respectively, three Zn levels of 50, 70, and 90 mg/kg and 45, 65, and 85 mg/kg were employed. Three Cu levels, namely 8, 12, and 16 mg/kg for 0-8 weeks of age, were also utilized. There were 288 (9 × 4 × 8) day-old turkey poults with equal body weight that were randomly assigned to 9 treatments, each consisting of 4 replicates and 8 poults per replicate. In comparison to other dietary combinations, there was a significant (P ≤ 0.05) increase in body weight gain observed in the 90 and 85 mg Zn/kg with 16 mg Cu/kg diet during 0-4 and 0-8 weeks of age, respectively, and the 70 and 65 mg Zn with 16 mg Cu /kg diet during 0-4 and 0-8 weeks of age, respectively. When compared to low levels of zinc in the diet at 5-8 and 0-8 weeks of age, respectively, feed conversion ratio was shown to be significantly (P ≤ 0.01) better with 90 and 85 mg Zn/kg diet. In a similar pattern, feed utilization efficiency was considerably (P ≤ 0.01) higher at 16 mg Cu/kg diet than it was at lower Cu levels in the diets given to the animals over the 0-4 week period. Compared to other dietary combinations, there was a substantial (P ≤ 0.01) improvement in cell-mediated immune response (foot web index to PHAP) and humoral immune response (HA titer to SRBC) when 85 mg zinc and 16 mg copper/kg diet was consumed at 0-4 and 5-8 weeks of age. At greater dietary Zn and Cu levels than its lower values, the weight of the spleen and thymus was considerably (P ≤ 0.05) higher. In a dietary combination of 90 mg Zn with 16 mg Cu/kg during 0-4 and 85 mg Zn with 16 mg Cu/kg diet during 5-8 weeks of age, respectively, there were significantly (P ≤ 0.05) greater bone width (proximal and distal), tibia bone ash, calcium, and phosphorus detected respectively. Significantly (P ≤ 0.01) greater Zn and Fe contents were found in the tibia bone at 90 and 85 mg Zn/kg diet, respectively, compared to values obtained at other Zn levels in the diet throughout 0-4 and 5-8 weeks of age. During the first 8 weeks of life, a diet containing 12 mg of copper per kilogram was shown to have a significantly (P ≤ 0.05) increased Zn and Fe content in the tibia bone compared to other levels. It is possible to draw the conclusion from the data that, for growing turkey poults, dietary combinations of 90 mg Zn/kg with 16 mg Cu/kg diet and 85 mg Zn with 16 mg Cu/kg diet between 0 and 4 and 5-8 weeks of age, respectively, were sufficient for optimum development, immunity, and skeletal health indices.

Effects of Different Concentration of Copper on Performance, Immunity and Carcass Traits in Broiler Japanese Quails.

In this study, the effects of dietary copper supplements on broiler Japanese quail growth performance, immune response, blood biochemistry, and carcass quality were examined. Two copper sources (copper sulphate-CuS, and copper methionine-CuM), each at five distinct dietary dosages of 5, 10, 15, 100, and 150 mg/kg, were used. A total of 280 (10 × 4 × 7) day-old quail chicks of uniform body weight were randomly distributed into 10 treatments with 4 replicates each and having 7 chicks in each replicate. In comparison to CuS-supplemented diets, CuM-supplemented diets (100 mg Cu/kg diet) considerably (P ≤ 0.01) increased body weight gain and improved feed conversion ratio (FCR). In the 150-mg CuM/kg diet, the cell-mediated immune response (foot web index to PHAP) was considerably (P ≤ 0.01) greater. The humoral immune response (HA titre to SRBC) was substantially (P ≤ 0.01) lower with CuS-supplemented meals than with CuM-supplemented diets. When compared to CuS source, the weight of the bursa and spleen from CuM source was considerably (P ≤ 0.01) higher. The 100- and 150-mg CuM/kg diets considerably (P ≤ 0.01) reduced serum cholesterol levels. Thus, it may be concluded that dietary supplementation of copper methionine as a source of Cu @ 100 mg Cu/kg diet to broiler Japanese quails was more effective in improving growth performance, immunological response, carcass quality features, and serum cholesterol reduction.

Effects of Various Copper Sources and Concentrations on Performance, Skeletal Growth, and Mineral Content of Excreta in Broiler Chickens.

The experiment was designed to study the effect of supplemental sources and concentrations of copper on the performance and development and mineralization of tibia bones in broiler chickens. A 42-day feeding experiment was conducted utilising three copper sources, including copper sulphate (CuS), copper chloride (CuCl), and copper propionate (CuP), each with four different concentrations, i.e. 8, 100, 150, and 200 mg/kg. The body weight gain with 200 mg Cu/kg food was noticeably higher during the first 4-6 weeks of age. Due to the interaction between Cu sources and levels, there was no significant change in the body weight gained. The feed intake during various growing phases did differ significantly neither the main effect nor the interaction between different copper sources and levels. A CuP-supplemented diet (200 mg/kg food) considerably (P ≤ 0.05) improved the feed conversion ratio between 4-6 and 0-6 weeks. At the end of the experiment, a total of 72 tibia bones, i.e. six for each treatment were collected. A metabolic trial was conducted to look into mineral retention in broiler chickens on the final 3 days of the trial (40-42 days). Increased tibia bone zinc (Zn) levels were seen with the addition of 8 mg Cu/kg of Cu chloride, 100 mg Cu/kg of Cu propionate, 8 mg Cu/kg of Cu sulphate, and 8 mg/kg of Cu propionate to the diet. At higher levels of Cu (150 and 200 mg/kg diet), there was a significantly (P ≤ 0.01) reduced tibia Zn content. Cu sulphate treatment group had higher (P ≤ 0.01) tibia Cu content (8 mg Cu/kg diet). Cu sulphate supplemented diet had a greater excreta Zn content (P ≤ 0.01) than Cu chloride supplemented diet, and Cu propionate supplemented diet had the lowest excreta Zn content. Excreta with a higher Fe concentration were found in diets supplemented with copper sulphate and copper chloride (P ≤ 0.05) than in diets supplied with copper propionate. Thus, it may be concluded that feeding dietary Cu concentrations up to 200 mg Cu/kg diet, regardless of the different sources, had no negative effects on bone morphometry and mineralization parameters with the exception of a decrease in the tibia's zinc content.

First report of whole genome sequence of septicemic Pasteurella multocida serovar B:2 'Soron' strain isolated from swine.

Pig pasteurellosis, caused by Pasteurella multocida, is an acute infection that also has economic implications for pig farmers. We report the complete genome sequence of a P. multocida, serovar B:2 'Soron' strain isolated from the blood of a pig that had died of pasteurellosis in India. The isolate was not found to be haemorrhagic septicaemia (HS) specific B:2 by the PCR assay. The genome of 'Soron' strain is a single circular chromosome of 2,272,124 base pairs in length and contains 2014 predicted coding regions, 4 ribosomal RNA operons, and 52 tRNAs. It has 1812 protein-coding genes that were also found in reference sequence PmP52Vac. Phylogenetic analysis revealed that Pm_P52VAc and P. multocida 'Soron' serovar B:2 were clustered in different clades. Pasteurella multocida 'Soron' serovar B:2 was found to cluster with the same ancestor of Pm70, which is of avian origin. The genome was found to contain regions that encode proteins which may confer resistance to various antibiotics including cephalosporin, which is used to treat pasteurellosis. The isolate was also found to harbour a phage region. This strain represents a novel multi-locus sequence type (MLST) that has not been previously identified, as all of the alleles used for MLST were found, but did not match any of the alleles in the database with 100% nucleotide identity. The most closely related ST was ST221. This is the first whole-genome sequence from P. multocida serovar B:2 of pig origin.

Changes in m6A RNA methylation of goat lung following PPRV infection.

Peste des petits ruminants (PPR) is an acute, highly contagious viral disease of goats and sheep, caused by the Peste des petits ruminants virus (PPRV). Earlier studies suggest the involvement of diverse regulatory mechanisms in PPRV infection. Methylation at N6 of Adenosine called m6A is a type RNA modification that influences various physiological and pathological phenomena. As the lung tissue represents the primary target organ of PPRV, the present study explored the m6A changes and their functional significance in PPRV disease pathogenesis. m6A-seq analysis revealed 1289 m6A peaks to be significantly altered in PPRV infected lung in comparison to normal lung, out of which 975 m6A peaks were hypomethylated and 314 peaks were hypermethylated. Importantly, hypomethylated genes were enriched in Interleukin-4 and Interleukin-13 signaling and various processes associated with extracellular matrix organization. Further, of the 843 differentially m6A-containing cellular transcripts, 282 transcripts were also found to be differentially expressed. Functional analysis revealed that these 282 transcripts are significantly enriched in signaling by Interleukins, extracellular matrix organization, cytokine signaling in the immune system, signaling by receptor tyrosine kinases, and Toll-like Receptor Cascades. We also found m6A reader HNRNPC and the core component of methyltransferase complex METTL14 to be highly upregulated than the m6A readers - HNRNPA2B1 and YTHDF1 at the transcriptome level. These findings suggest that alteration in the m6A landscape following PPRV is implicated in diverse processes including Interleukin signaling.

Effects of paraprobiotic as replacements for antibiotic on performance, immunity, gut health and carcass characteristics in broiler chickens.

This study sought to determine the effects of dietary paraprobiotic (PPB) on broiler chicken performance, immunity, gut health, and carcass traits. A total of 240 day-old CARIBRO Vishal commercial broiler chicks of identical body weight randomly divided into six treatment groups, each with five replicates and eight chicks in each replicate. Six dietary treatments were preapared: T1 = (control diet), T2 = T1 + 0.02% (w/v) chlortetracycline (CTC), T3 = T1 + 0.2% (w/v) PPB, T4 = T1 + 0.4% (w/v) PPB, T5 = T1 + 0.6% (w/v) PPB and T6 = T1 + 0.8% (w/v) PPB, respectively. Body weight gain (BWG) significantly (P ≤ 0.05) increased in the T5 (0.6% PPB) and T6 (0.8% PPB) group. At the same time the feed intake significantly (P ≤ 0.05) decreased and the feed conversion ratio (FCR) significantly (P ≤ 0.05) improved in T5 and T6 group. There was a significant (P ≤ 0.05) increase in cell-mediated immunity and haem-agglutination titre (HA titre) in the 0.6% and 0.8% PPB supplemented groups compare to the control group (T1). The percentage of carcass traits and organ weights did not significantly differ between the PPB-supplemented and control groups, but the percentage of live weight in cut up parts showed a significant improvement (P ≤ 0.05) in the PPB-supplemented group. At 42 days, villus height, width, and crypt depth all significantly (P ≤ 0.05) increased in the groups supplemented with 0.6 and 0.8% para-probiotics (T5 and T6). The results show that para-probiotics can be added to broiler diets at a rate of 0.6% (w/v) to enhance performance, immunity, gut health, and breast yield. The para-probiotic may therefore be a useful substitution for antibiotic growth promoters in the diet of chickens.

Chemical profiling of marine seaweed Halimeda gracilis using UPLC-ESI-Q-TOF-MSE and evaluation of anticancer activity targeting PI3K/AKT and intrinsic apoptosis signaling pathway.

Marine seaweeds are predominant for nutraceuticals and have been food source since ancient times and are currently being used in Chinese medicines and Japanese traditional medicines. In the present study, the chemical profile analysis of Halimeda gracilis was performed using UPLC-ESI-Q-TOF-MSE analysis and assessed for its anticancer activity. A cursory investigation of the total ion chromatograms of the both methanol (MHG) and ethyl acetate (EAHG) of H. gracilis extracts reveals that both extracts have different kind of metabolites including phenols and its derivatives, diterpenes, cinnamic acids derivatives etc. The in vitro anticancer activity of EAHG and MHG exhibiting significant activity and induced apoptosis against skin cancer cells by generating excessive ROS, damaging the mitochondrial membrane and nuclear components. Further, the western blot analysis showed that the EAHG and MHG downregulates the oncoproteins PI3K, AKT, p-AKT, and BcL2 and upregulates the apoptotic proteins Bax, Cyto C, p21, p53, Caspase 9 and Caspase 3. To best of our knowledge, this is the first report which implies the UPLC-ESI-Q-TOF-MSE based chemical profiling of H. gracilis and can be used for the treatment of cancer.

Bombax ceiba calyx displays antihyperglycemic activity via improving insulin secretion and sensitivity: Identification of bioactive phytometabolomes by UPLC-QTof-MS/MS.

Vegetables are considered good food for the management of hyperglycemia. Bombax ceiba L. (family: Bombacaceae) calyces are part of traditional vegetables. This study evaluated its usefulness on various parameters responsible for the development of hyperglycemia and conducted phytometabolomic analysis to identify phytochemicals responsible for the observed activities. It was found that the aqueous methanol extract of its calyces (B. ceiba calyx extract, BCE) reduced (12.4%) significantly (p < 0.05) the development of sucrose-induced postprandial hyperglycemic load in rats. In-vitro studies revealed that BCE improved glucose-stimulated insulin secretory activity in MIN6 cells plausibly by decreasing ADP/ATP ratio. BCE also augmented concentration-dependent (5 µg, 10 µg, and 20 µg) increase in glucose uptake in hyperglycemic L6 myotubes both by non-insulin-dependent manner (35%, 68%, and 132%, respectively) and insulin-dependent manner (42%, 59%, and 172%, respectively). The insulin-stimulated GLUT4 translocation was compromised (34%) significantly (p < 0.05) under hyperglycemic condition; however, it was improved by 23% and 72% (p < 0.001) when L6 myotubes were primed with 10 and 20 µg of BCE, respectively. Hyperglycemia aggravated reactive oxygen species (ROS) generation in L6 myotubes. The ROS generation was significantly (p < 0.001) reduced by priming myotubes with BCE before challenging myotubes to hyperglycemic environment, possibly by preserving cellular antioxidant enzymes catalase, glutathione peroxidase, and reduced glutathione levels. Phytometabolomic analysis disclosed a number of phytochemicals present in B. ceiba calyces known to display these activities. This is the first study reporting antihyperglycemic activity in B. ceiba calyces, its mechanisms of action, and phytometabolomic profile applying UPLC-QTof-MS/MS technique. PRACTICAL APPLICATION: B. ceiba calyces are part of traditional vegetables. Our study finds that B. ceiba calyces contain phytochemicals possessing antihyperglycemic, insulin secretory, insulin sensitization properties, and potentials for preserving hyperglycemia-induced vitiations in cellular antioxidant defense. These observations provide foundation for exploring further possibilities of B. ceiba calyces to become valuable dietary inclusion in the diet of people suffering from metabolic disorders.

A novel quantitative assay for analysis of GLUT4 translocation using high content screening.

Insulin resistance is associated with obesity and can lead to several metabolic disorders including type II diabetes, nonalcoholic fatty liver disease and cardiovascular problems. Search for the small molecules which can either induce or mimic the insulin action are of great interest and can be utilized to manage insulin resistance. There are several dietary phytochemicals which can potentially have insulinomimetic action. Nevertheless, high throughput screening methods to test efficiency of small molecules to act as an insulinomimetic are not fully established. In this paper we have performed chemical screen analysis based on GLUT4 translocation using a cell line CHO-HIRC-myc-GLUT4 eGFP that expresses GLUT4-GFP in association with human Insulin receptor. We have established a high content screening-based method which can track and quantify the GLUT4 translocation from perinuclear area to the cell membrane. The assay involves measuring fluorescence intensity in a defined perinuclear area and a defined area along the cell membrane; and the results are expressed as the ratio of fluorescence intensity in the perinuclear to membrane area. The assay could collect real time data of GLUT4 translocation from thousand of cells/ sample and from many such samples in one experiment. We validated the assay using Insulin, insulin mimics/sensitizers and insulin inhibitors. The agonist or antagonists were analyzed for their ability to enhance or block the GLUT4 translocation independent of insulin. The outcome of the assay was correlated by performing glucose uptake assay using differentiated 3T3L1 cells. Using this platform we further identified several plant extracts which had the insulin mimetic action. We confirmed that these plant extracts were non-toxic to the beta cells using RIN mf5cells and 3T3L1 cells. We have identified plant extracts with the potential insulinomimetic action using novel high-content screening approach; these can be further tested for their efficiency in-vivo in pre-clinical trials.