Genomic and probiotic attributes of Lactobacillus strains from rice-based fermented foods of North Eastern India.

The probiotic attributes and genomic profiles of amylase-producing Lactobacillus strains from rice-based fermented foods of Meghalaya in the North-Eastern India were evaluated in the study. A preliminary screening of 17 lactic acid bacteria strains was performed based on their starch hydrolysis and glucoamylase activities. Out of 17 strains, 5 strains (L. fermentum KGL4, L. rhamnosus RNS4, L. fermentum WTS4, L. fermentum KGL2, and L. rhamnosus KGL3A) were selected for further characterization of different probiotic attributes. Whole-genome sequencing of two of the best strains was carried out using a shotgun sequencing platform based on their rich probiotic attributes. The EPS production was in the range of 2.89-3.92 mg/mL. KGL2 (41.5%) and KGL3A (41%) showed the highest antioxidant activity. The highest antibiotic susceptibility was exhibited by all the five Lactobacillus strains against ampicillin, ranging from 24.66 to 27.33 mm. The lactobacilli isolates used in the study could survive the simulated gastric/intestinal juices. Genomic characterization of KGL4 and KGL3A illustrated their possible adherence to the intestinal wall, specialized metabolic patterns, and possible role in boosting host immunity. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-022-05633-8.

RNA-Sequencing based analysis of bovine endometrium during the maternal recognition of pregnancy.

BACKGROUND: Maternal recognition is the crucial step for establishing pregnancy in cattle. This study aims to identify endometrial genes and biological pathways involved in the maternal recognition of pregnancy. Caruncular endometrial tissues were collected from Day 15-17 of gestation (pregnant), non-pregnant (absence of conceptus), and cyclic (non-bred) heifers. RESULTS: Total RNAs were isolated from the caruncular endometrial tissues of pregnant, non-pregnant, and cyclic heifers, and were subjected to high-throughput RNA-sequencing. The genes with at least two-fold change and Benjamini and Hochberg p-value ≤ 0.05 were considered differentially expressed genes and further confirmed with quantitative real-time PCR. A total of 107 genes (pregnant vs cyclic) and 98 genes (pregnant vs non-pregnant) were differentially expressed in the pregnant endometrium. The most highly up-regulated genes in the pregnant endometrium were MRS2, CST6, FOS, VLDLR, ISG15, IFI6, MX2, C15H11ORF34, EIF3M, PRSS22, MS4A8, and TINAGL1. Interferon signaling, immune response, nutrient transporter, synthesis, and secretion of proteins are crucial pathways during the maternal recognition of pregnancy. CONCLUSIONS: The study demonstrated that the presence of conceptus at Day 15-17 of gestation affects the endometrial gene expression related to endometrial remodeling, immune response, nutrients and ion transporters, and relevant signaling pathways in the caruncular region of bovine endometrium during the maternal recognition of pregnancy.

RNA sequencing-based analysis of the magnum tissues revealed the novel genes and biological pathways involved in the egg-white formation in the laying hen.

BACKGROUND: The mechanism of egg formation in the oviduct of laying hens is tightly controlled; each segment of the oviduct contributes a unique component of the egg. Several genes/proteins are involved in the synthesis of a completely healthy egg. This implies a time- and tissue-specific expression of genes and proteins in the different oviductal segments. We used hens at different physiological stages and time points to understand the transcriptional regulation of egg-white (albumen) synthesis and secretion onto the eggs in the magnum of laying hens. This study used Next-Generation Sequencing and quantitative real-time PCR (qPCR) to detect the novel genes and the cognate biological pathways that regulate the major events during the albumen formation. RESULTS: Magnum tissues collected from laying (n = 5 each at 3 h post-ovulation, p.o. and 15-20 h p.o.), non-laying (n = 4), and molting (n = 5) hens were used for differential gene expression analyses. A total of 540 genes (152 upregulated and 388 down-regulated) were differentially expressed at 3 h p.o. in the magnum of laying hens. Kyoto Encyclopedia of Genes and Genomes pathways analysis of the 152 upregulated genes revealed that glycine, serine, and threonine metabolism was the most-enriched biological pathway. Furthermore, the top two most enriched keywords for the upregulated genes were amino-acid biosynthesis and proteases. Nine candidate genes associated with albumen formation were validated with qPCR to have differential expression in laying, non-laying, and molting hens. Proteases such as TMPRSS9, CAPN2, MMP1, and MMP9 (protein maturation, ECM degradation, and angiogenesis); enzymes such as PSPH, PHGDH, and PSAT1 (amino-acid biosynthesis); RLN3, ACE, and REN (albumen synthesis, secretion and egg transport); and AVD, AvBD11, and GPX3 (antimicrobial and antioxidants) were recognized as essential molecules linked to albumen deposition in the magnum. CONCLUSIONS: This study revealed some novel genes that participate in the signaling pathways for egg-white synthesis and secretion along with some well-known functional genes. These findings help to understand the mechanisms involved in albumen biosynthesis.

Synergistic inhibition of csal1 and csal3 in granulosa cell proliferation and steroidogenesis of hen ovarian prehierarchical development†.

SALL1 and SALL3 are transcription factors that play an essential role in regulating developmental processes and organogenesis in many species. However, the functional role of SALL1 and SALL3 in chicken prehierarchical follicle development is unknown. This study aimed to explore the potential role and mechanism of csal1 and csal3 in granulosa cell proliferation, differentiation, and follicle selection within the prehierarchical follicles of hen ovary. Our data demonstrated that the csal1 and csal3 transcriptions were highly expressed in granulosa cells of prehierarchical follicles, and their proteins were mainly localized in the cytoplasm of granulosa cells and oocytes as well as in the ovarian stroma and epithelium. It initially revealed that both csal1 and csal3 may be involved in chicken prehierarchical follicle development via a translocation mechanism. Furthermore, our results showed an abundance of CCND1, Bcat, StAR, CYP11A1, and FSHR mRNA in granulosa cells, and the proliferation levels of granulosa cells from the prehierarchical follicles were significantly increased by siRNA-mediated knockdown of csal1 or/and csal3. Conversely, the overexpression of csal1 or/and csal3 in the granulosa cells led to a remarkably decreased of them. Moreover, csal1 and csal3 together exert a much stronger effect on the regulation than any of csal1 or csal3. These results indicated that csal1 and csal3 play synergistic inhibitory roles on granulosa cell proliferation, differentiation, and steroidogenesis during prehierarchical follicle development in vitro. The current data provide a basis of molecular mechanisms of csal1 and csal3 in controlling the prehierarchical follicle development and growth of hen ovary in vivo.

Differential Transcriptome Analysis of Early Postnatal Developing Longissimus Dorsi Muscle from Two Pig Breeds Characterized in Divergent Myofiber Traits and Fatness.

Meat quality traits (MQTs) are very important in the porcine industry, which are mainly determined by skeletal muscle fiber composition, extra-muscular and/or intramuscular fat content. To identify the differentially expressed candidate genes affecting the meat quality traits, first we compared the MQTs and skeletal muscle fiber characteristics in the longissimus dorsi muscle (LDM) of the Northeast Min pig (NM) and the Changbaishan wild boar (CW) with their body weight approaching 90 kg. The significant divergences in the skeletal muscle fiber phenotypes and fatness traits between the two porcine breeds established an ideal model system for further identifying potential key functional genes that dominated MQTs. Further, a transcriptome profile analysis was performed using the Illumina sequencing method in early postnatal developing LDM from the two breeds at the ages of 42 days. Comparative analysis between these two cDNA libraries showed that there were 17,653 and 22,049 unambiguous tag-mapped sense transcripts detected from NM and CW, respectively. 4522 differentially expressed genes (DEGs) were revealed between the two tissue samples, of them, 4176 genes were found as having been upregulated and 346 genes were identified as having been downregulated in the NM library. By pathway enrichment analysis, a set of significantly enriched pathways were identified for the DEGs, which are potentially involved in myofiber development, differentiation and growth, lipogenesis and lipolysis in porcine skeletal muscle. The expression levels of 30 out of the DEGs were validated by real-time quantitative reverse transcriptase PCR (qRT-PCR) and the observed result was consistent noticeably with the Illumina transcriptome profiles. The findings from this study can contribute to future investigations of skeletal muscle growth and development mechanism and to establishing molecular approaches to improve meat quality traits in pig breeding.

Biofunctional Attributes and Storage Study of Soy Milk Fermented by Lactobacillus rhamnosus and Lactobacillus helveticus.

Different soybean products are considered as traditional functional food among the Mongolian population in Northeast India. In the present study, the effect of different flavours (mango, orange, vanilla and white rose), inoculation rates of Lactobacillus rhamnosus K4E (KX950834) and Lactobacillus helveticus K14 (KU644578), and mass fractions of skimmed milk and sugar on the acceptability of soy yoghurts was studied. Physicochemical (pH, titratable acidity) and microbial analyses (total bacterial, total coliform, yeast and mould count) were conducted, and organoleptic (aroma, taste, colour, mouthfeel, texture and overall acceptability) and biofunctional properties (angiotensin-converting enzyme (ACE) inhibitory activity, antioxidant and antimicrobial activities and biotransformation of isoflavones) were evaluated during storage up to 10 days under refrigeration conditions (6-8 °C). Panellists preferred white rose soy yoghurt more than other flavours. The pH was from 5.65 to 4.20, the titratable acidity (expressed as mass fraction of lactic acid) was from 0.33 to 0.51% and total Lactobacillus count ranged from 6.81 to 8.69 log CFU/mL during storage. The ACE inhibitory activity increased from 21.17% on day 0 to 81.03% on day 5, followed by a decrease of the activity after 10 days (38.85%). The antioxidant activity was the highest on day 5 (87%). White rose soy yoghurt had the highest antimicrobial activity against Listeria monocytogenes, followed by Bacillus subtilis, Staphylococcus aureus, Salmonella typhi and Escherichia coli. RP-HPLC analysis showed that after 18 h, the production of soy isoflavone aglycones genistein and daidzein in yoghurt was 87.3 and 58.4%, respectively.

Very low doses of heavy oxygen ion radiation induce premature ovarian failure.

Astronauts are exposed to charged particles during space travel, and charged particles are also used for cancer radiotherapy. Premature ovarian failure is a well-known side effect of conventional, low linear energy transfer (LET) cancer radiotherapy, but little is known about the effects of high LET charged particles on the ovary. We hypothesized that lower LET (16.5 keV/µm) oxygen particles would be less damaging to the ovary than we previously found for iron (LET = 179 keV/µm). Adult female mice were irradiated with 0, 5, 30 or 50 cGy oxygen ions or 50 cGy oxygen plus dietary supplementation with the antioxidant alpha lipoic acid (ALA). Six-hour after irradiation, percentages of ovarian follicles immunopositive for γH2AX, a marker of DNA double strand breaks, 4-HNE, a marker of oxidative lipid damage and BBC3 (PUMA), a proapoptotic BCL-2 family protein, were dose dependently increased in irradiated mice compared to controls. One week after irradiation, numbers of primordial, primary and secondary follicles per ovary were dose dependently decreased, with complete absence of follicles in the 50 cGy groups. The ED50 for primordial follicle destruction was 4.6 cGy for oxygen compared to 27.5 cGy for iron in our previous study. Serum FSH and LH concentrations were significantly elevated in 50 cGy groups at 8 week. Supplementation with ALA mitigated the early effects, but not the ultimate depletion of ovarian follicles. In conclusion, oxygen charged particles are even more potent inducers of ovarian follicle depletion than charged iron particles, raising concern for premature ovarian failure in astronauts exposed to both particles during space travel.

Charged iron particles, components of space radiation, destroy ovarian follicles.

STUDY QUESTION: Do charged iron particles, components of space radiation, cause premature ovarian failure? SUMMARY ANSWER: Exposure to charged iron particles causes ovarian DNA damage, oxidative damage and apoptosis, resulting in premature ovarian failure. WHAT IS KNOWN ALREADY: The ovary is very sensitive to follicle destruction by low linear energy transfer (LET) radiation, such as X-rays and γ-rays. However, it is completely unknown whether high-LET radiation, such as charged iron particles, also destroys ovarian follicles. STUDY DESIGN, SIZE, DURATION: Twelve week old C57BL/6J female mice were exposed to single doses of 0, 5, 30 or 50 cGy (n = 8/group) charged iron particles (LET = 179 keV/µm) at energy of 600 MeV/u. Two groups were irradiated at the highest dose, one fed AIN-93M chow and the other fed AIN-93M chow supplemented with 150 mg/kg diet alpha lipoic acid (ALA). PARTICIPANTS/MATERIALS, SETTING, METHODS: We quantified the numbers of ovarian follicles, measured serum follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations, and analyzed histone H2AX phosphorylation, oxidative damage and apoptosis markers in the ovarian follicles. MAIN RESULTS AND THE ROLE OF CHANCE: H2AX phosphorylation, lipid peroxidation, protein nitration and apoptosis were highly induced in ovarian follicles at 6 h and remained increased 1 week after irradiation. As a result, numbers of healthy ovarian follicles were significantly and dose-dependently depleted at 1 and 8 weeks post-irradiation, with 57, 84 and 99% decreases in primordial follicles at 8 weeks at the 5, 30 and 50 cGy doses, respectively (P < 0.05 versus 0 cGy). Consistent with near-total depletion of ovarian follicles in the 50 cGy group, serum concentrations of FSH and LH were significantly elevated at 8 weeks. Dietary supplementation with ALA partially prevented the adverse ovarian effects of 50 cGy iron particles. LIMITATIONS, REASONS FOR CAUTION: About 21% of the estimated radiation dose from exposure to galactic cosmic rays during a multi-year Mars mission will be due to high-LET particles, of which iron is only one. The effects of galactic cosmic rays, which contain a mixture of multiple charged particles, as well as protons, neutrons, and helium ions, may differ from the effects of iron alone. WIDER IMPLICATIONS OF THE FINDINGS: We show for the first time that charged high-LET ions are highly damaging to the ovary even at low doses, causing premature ovarian failure. In addition to raising concerns for female astronauts, these findings raise concerns for ovarian damage due to clinical uses of high-LET particles for cancer treatment. In addition to causing infertility, premature ovarian failure has adverse implications for the functions of heart, brain, bone and muscle later in life. STUDY FUNDING/COMPETING INTERESTS: This work was supported by a National Aeronautics and Space Administration grant NNX14AC50G to U.L. B.M. was partially supported by a National Space Biomedical Research Institute First Award, PF04302. Additional support was received from the University of California Irvine Center for Occupational and Environmental Health. The authors have no conflicts of interests.

Embryonic thermal manipulation: a potential strategy to mitigate heat stress in broiler chickens for sustainable poultry production.

Due to high environmental temperatures and climate change, heat stress is a severe concern for poultry health and production, increasing the propensity for food insecurity. With climate change causing higher temperatures and erratic weather patterns in recent years, poultry are increasingly vulnerable to this environmental stressor. To mitigate heat stress, nutritional, genetic, and managerial strategies have been implemented with some success. However, these strategies did not adequately and sustainably reduce the heat stress. Therefore, it is crucial to take proactive measures to mitigate the effects of heat stress on poultry, ensuring optimal production and promoting poultry well-being. Embryonic thermal manipulation (TM) involves manipulating the embryonic environment's temperature to enhance broilers' thermotolerance and growth performance. One of the most significant benefits of this approach is its cost-effectiveness and saving time associated with traditional management practices. Given its numerous advantages, embryonic TM  is a promising strategy for enhancing broiler production and profitability in the poultry industry. TM increases the standard incubation temperature in the mid or late embryonic stage to induce epigenetic thermal adaption and embryonic metabolism. Therefore, this review aims to summarize the available literature and scientific evidence of the beneficial effect of pre-hatch thermal manipulation on broiler health and performance.

Effect of in ovo feeding of xylobiose and xylotriose on plasma immunoglobulin, cecal metabolites production, microbial ecology, and metabolic pathways in broiler chickens.

BACKGROUND: Dietary supplementation of xylooligosaccharides (XOS) has been found to influence gut health by manipulating cecal microbiota and producing microbe-origin metabolites. But no study investigated and compared the effect of in ovo feeding of xylobiose (XOS2) and xylotriose (XOS3) in chickens. This study investigated the effect of in ovo feeding of these XOS compounds on post-hatch gut health parameters in chickens. A total of 144 fertilized chicken eggs were divided into three groups: a) non-injected control (CON), b) XOS2, and c) XOS3. On the 17th embryonic day, the eggs of the XOS2 and XOS3 groups were injected with 3 mg of XOS2 and XOS3 diluted in 0.5 mL of 0.85% normal saline through the amniotic sac. After hatching, the chicks were raised for 21 d. Blood was collected on d 14 to measure plasma immunoglobulin. Cecal digesta were collected for measuring short-chain fatty acids (SCFA) on d 14 and 21, and for microbial ecology and microbial metabolic pathway analyses on d 7 and 21. RESULTS: The results were considered significantly different at P < 0.05. ELISA quantified plasma IgA and IgG on d 14 chickens, revealing no differences among the treatments. Gas chromatography results showed no significant differences in the concentrations of cecal SCFAs on d 14 but significant differences on d 21. However, the SCFA concentrations were lower in the XOS3 than in the CON group on d 21. The cecal metagenomics data showed that the abundance of the family Clostridiaceae significantly decreased on d 7, and the abundance of the family Oscillospiraceae increased on d 21 in the XOS2 compared to the CON. There was a reduction in the relative abundance of genus Clostridium sensu stricto 1 in the XOS2 compared to the CON on d 7 and the genus Ruminococcus torques in both XOS2 and XOS3 groups compared to the CON on d 21. The XOS2 and XOS3 groups reduced the genes for chondroitin sulfate degradation I and L-histidine degradation I pathways, which contribute to improved gut health, respectively, in the microbiome on d 7. In contrast, on d 21, the XOS2 and XOS3 groups enriched the thiamin salvage II, L-isoleucine biosynthesis IV, and O-antigen building blocks biosynthesis (E. coli) pathways, which are indicative of improved gut health. Unlike the XOS3 and CON, the microbiome enriched the pathways associated with energy enhancement, including flavin biosynthesis I, sucrose degradation III, and Calvin-Benson-Bassham cycle pathways, in the XOS2 group on d 21. CONCLUSION: In ovo XOS2 and XOS3 feeding promoted beneficial bacterial growth and reduced harmful bacteria at the family and genus levels. The metagenomic-based microbial metabolic pathway profiling predicted a favorable change in the availability of cecal metabolites in the XOS2 and XOS3 groups. The modulation of microbiota and metabolic pathways suggests that in ovo XOS2 and XOS3 feeding improved gut health during the post-hatch period of broilers.

Pre-hatch thermal manipulation of embryos and post-hatch baicalein supplementation mitigated heat stress in broiler chickens.

BACKGROUND: High environmental temperatures induce heat stress in broiler chickens, affecting their health and production performance. Several dietary, managerial, and genetics strategies have been tested with some success in mitigating heat stress (HS) in broilers. Developing novel HS mitigation strategies for sustaining broiler production is critically needed. This study investigated the effects of pre-hatch thermal manipulation (TM) and post-hatch baicalein supplementation on growth performance and health parameters in heat-stressed broilers. RESULTS: Six hundred fertile Cobb 500 eggs were incubated for 21 d. After candling on embryonic day (ED) 10, 238 eggs were thermally manipulated at 38.5 °C with 55% relative humidity (RH) from ED 12 to 18, then transferred to the hatcher (ED 19 to 21, standard temperature) and 236 eggs were incubated at a controlled temperature (37.5 °C) till hatch. After hatch, 180-day-old chicks from both groups were raised in 36 pens (n = 10 birds/pen, 6 replicates per treatment). The treatments were: 1) Control, 2) TM, 3) control heat stress (CHS), 4) thermal manipulation heat stress (TMHS), 5) control heat stress supplement (CHSS), and 6) thermal manipulation heat stress supplement (TMHSS). All birds were raised under the standard environment for 21 d, followed by chronic heat stress from d 22 to 35 (32-33 °C for 8 h) in the CHS, TMHS, CHSS, and TMHSS groups. A thermoneutral (22-24 °C) environment was maintained in the Control and TM groups. RH was constant (50% ± 5%) throughout the trial. All the data were analyzed using one-way ANOVA in R and GraphPad software at P < 0.05 and are presented as mean ± SEM. Heat stress significantly decreased (P < 0.05) the final body weight and ADG in CHS and TMHS groups compared to the other groups. Embryonic TM significantly increased (P < 0.05) the expression of heat shock protein-related genes (HSP70, HSP90, and HSPH1) and antioxidant-related genes (GPX1 and TXN). TMHS birds showed a significant increment (P < 0.05) in total cecal volatile fatty acid (VFA) concentration compared to the CHS birds. The cecal microbial analysis showed significant enrichment (P < 0.05) in alpha and beta diversity and Coprococcus in the TMHSS group. CONCLUSIONS: Pre-hatch TM and post-hatch baicalein supplementation in heat-stressed birds mitigate the detrimental effects of heat stress on chickens' growth performance, upregulate favorable gene expression, increase VFA production, and promote gut health by increasing beneficial microbial communities.

Effects of microalgae, with or without xylanase supplementation, on serum immunoglobulins, cecal short-chain fatty acids, microbial diversity, and metabolic pathways of broiler chickens.

Modern broilers are highly susceptible to environmental and pathogenic threats, leading to gut disorders and poor nutrient utilization if not managed properly. Nutritional programming using several feedstuffs and coproducts to manage gut health has been studied. This study used microalgae as a functional compound and xylanase enzyme in broilers' diets as a strategy to manage gut health. A total of 162 one-day-old unsexed Cobb 500 broiler chicks were randomly assigned to 1 of the 3 dietary treatments: a) corn-soybean meal-based control diet (CON), b) 3% microalgae (MAG), and c) MAG with xylanase enzyme (MAG+XYN). The chicks were reared for 35 days (d) on a floor pen system maintaining standard environment conditions to evaluate the effects of microalgae, with or without xylanase supplementation, on serum immunoglobulins, cecal short-chain fatty acids (SCFA) production, cecal microbial diversity, and metabolic pathways. No significant differences were found for serum immunoglobulin and cecal SCFA among the treatment groups (P > 0.05). Relative microbial abundance at the genus level showed that MAG and MAG+XYN groups had a diverse microbial community on d 3 and d 35. However, no bacterial genus had a significant difference (P > 0.05) in their relative abundance on d 3, but 16 genera showed significant differences (P < 0.05) in their relative abundance among the dietary treatments on d 35. Most of these bacteria were SCFA-producing bacteria. Moreover, MAG and MAG+XYN-fed broilers had better responses than CON groups for metabolic pathways (D-mannose degradation, pectin degradation I and II, ß-1-4-mannan degradation, tetrahydrofolate biosynthesis, glutathione biosynthesis, glutathione-peroxide redox reactions, lactate fermentation to propionate, acetate, and hydrogen, etc.) both on d 3 and d 35. The results suggest that using microalgae, with or without xylanase, had no statistical impact on serum immunoglobulins and cecal SCFA production in broilers. However, an improvement in the cecal microbial diversity and metabolic pathways, which are essential indicators of gut health and nutrient utilization, was observed. Most of the improved metabolic pathways were related to fiber utilization and oxidative stress reduction.

Scanning electron microscope-based evaluation of eggshell quality.

The eggshell is the outermost covering of an egg that provides physical and chemical protection. It is a major source of calcium and minerals for the growing embryo during incubation. The egg industry suffers from a considerable economic loss due to poor eggshell quality. Therefore, developing an accurate and precise method of determining eggshell quality is crucial in improving eggshells in subsequent generations of breeding stock. Hence, this study aimed to develop a method to accurately and precisely determine 1) eggshell thickness using a scanning electron microscope (SEM) and 2) eggshell mineral components using an SEM-Energy Dispersive Spectrometry (EDS) system. Four types of table eggs (N = 48; 12 eggs/group): Cage-Free Organic from the US Mainland (CFO-M) and Hawaii (CFO-H), Caged Non-Organic from the US Mainland (CNO-M), and Hawaii (CNO-H) were sourced from the grocery store. Approximately 0.5 mm2 pieces of eggshells from the equator region of the egg were taken and processed for visualization under the SEM. Three distinct layers of eggshell were identified under SEM: the outermost cuticle, the middle palisade, and the innermost mammillary region. The results showed that CFO-H eggs have a greater eggshell thickness (380.43 ± 2.69 µm) and effective thickness (306.28 ± 4.15 µm). Similarly, the mammillary knob count was denser in CNO-H eggs (186 ± 23.02 knobs/0.5 mm2). Calcium (97.36 ± 0.17%) was the highest among minerals in lower palisades (LP). The magnesium concentration was lowest in the LP region, whereas the phosphorus concentration was highest in the upper palisades. Our study established a scientific method to assess the eggshell quality and biochemical characteristics of eggs through SEM and EDS. This method can be used as a marker for selecting superior parent stock to improve eggshell quality in subsequent generations of breeding stock.

Epigen enhances the developmental potential of in vitro fertilized embryos by improving cytoplasmic maturation.

Numerous growth factors contribute to oocyte maturation and embryonic development in vivo; however, only a few are understood. One such factor is epigen, a new member of the epidermal growth factor (EGF) family that is secreted by the granulosa cells of immature oocytes. We hypothesized that epigen may play a role in oocyte maturation, specifically in the nuclear and cytoplasmic aspects. This study aimed to investigate the effects of epigen on porcine oocyte maturation and embryo development in vitro. In this study, three different concentrations of epigen (3, 6, and 30 ng/mL) were added to tissue culture medium-199 (TCM-199) during in vitro maturation of porcine oocytes. A control group that did not receive epigen supplementation was also included. Mature porcine oocytes were fertilized, and the resulting zygotes were cultured until day 7. The levels of intracellular glutathione (GSH) and reactive oxygen species (ROS) were measured in the in vitro matured oocytes. At the same time, the expression patterns of genes related to apoptosis were detected in day 7 blastocysts (BLs) using real-time quantitative PCR Apoptosis was detected by annexin-V assays in mature oocytes. Data were analyzed using ANOVA and Duncan's test on SPSS, and results are presented as mean ± SEM. The group that received 6 ng/mL epigen had a significantly lower rate of germinal vesicle breakdown (GVBD) than the control group without affecting the nuclear maturation among the experimental groups. Among the treatment groups, the 6 ng/mL epigen group showed significantly higher levels of intracellular GSH and lower ROS production. Supplementation with 6 ng/mL epigen significantly improved blastocyst (BL) formation rates compared to those in the control and 3 ng/mL groups. Additionally, the blastocyst expansion rate was significantly higher with epigen supplementation (6 ng/mL). In the fertilization experiment, the group supplemented with 6 ng/mL epigen exhibited significantly higher levels of monospermy and fertilization efficiency and lower levels of polyspermy than the control group. This study indicated that adding epigen at a concentration of 6 ng/mL can significantly enhance the developmental potential of porcine oocytes fertilized in vitro. Specifically, the study found that epigen improves cytoplasmic maturation, which helps prevent polyspermy and emulates monospermic penetration.

Embryonic thermal manipulation reduces hatch time, increases hatchability, thermotolerance, and liver metabolism in broiler embryos.

The broilers' health and growth performance are affected by egg quality, incubation conditions, and posthatch management. Broilers are more susceptible to heat stress because they have poor thermoregulatory capacity. So, it is crucial to develop a strategy to make chicks thermotolerant and cope with heat stress in post-hatch life. This study investigated the effects of embryonic thermal manipulation (TM) on different hatching parameters (hatch time, hatchability, and hatch weight), brain thermotolerance, and liver metabolism. Six hundred fertile Cobb 500 eggs were incubated for 21 d. After candling on embryonic day (ED) 10, 238 eggs were thermally manipulated at 38.5°C with 55% relative humidity (RH) from ED 12 to 18, then transferred to the hatcher (ED 19-21, standard temperature, 37.5°C) and 236 eggs were incubated at a standard temperature (37.5°C) till hatch. The samples were collected from the Control and TM groups on ED 15 and 18 of the embryonic periods. Hatchability was significantly higher (P < 0.05) in the TM group (94.50%) than in the control group (91.0%). Hatch weight did not differ significantly between the TM group (50.54 g) and the Control group (50.39 g). Most importantly, hatch time was significantly lower (P < 0.05) in the TM group than in the Control. In the D15 embryo brain, the mRNA expression of TRPV1,TRPV2, TRPV3, and the epigenetic marker H3K27 were significantly lower (P < 0.05) in the TM group compared to the Control group. However, in the D18 brain, the expression of TRPV1, TRPV2, and CRHR1 was significantly higher (P < 0.05) in the TM group than in the Control group. In the liver, the mRNA expression of SLC6A14 was significantly lower (P < 0.05) in the D15 TM group than in the D15 Control group. Conversely, the DIO3 mRNA expression was significantly higher (P < 0.05) in the D15 TM group than in the D15 Control group. The expression of GPX3, FOXO1, IGF2, and GHR in the liver was significantly higher in the D18 TM group compared to the D18 Control group (P < 0.05). In conclusion, increased expression of the aforementioned markers during the later embryonic period has been linked to reduced hatch time by increasing liver metabolism and thermotolerance capacity in the brain.

Application of botanical products as nutraceutical feed additives for improving poultry health and production.

Poultry is one of the most consumed sources of animal protein around the world. To meet the global demands for poultry meat and eggs, it is necessary to improve their nutrition to sustain the poultry industry. However, the poultry industry faces several challenges, including feedstuff availability, the banning of antibiotics as growth promoters, and several environmental stressors. Therefore, there is a critical need to include available nutraceuticals in the diet to sustain the poultry industry. Nutraceuticals are natural chemical substances that positively influence animal physiological and productive traits. Botanical products (such as fenugreek seeds, ginger roots, and olive leaves) are among the most commonly used nutraceuticals and are gradually gaining popularity in the poultry industry due to their immense benefits in nutrition and therapeutic properties. They can be added to the diet separately or in combination (as a natural antioxidant and immunostimulant) to improve poultry health and production. Botanical products are rich in essential oils and essential fatty acids, which have multiple benefits on the animal's digestive system, such as activating the digestive enzymes and restoring microbiota balance, enhancing poultry health, and production. These nutraceuticals have been shown to stimulate the expression of several genes related to growth, metabolism, and immunity. In addition, the essential oil supplementation in poultry diets up-regulated the expression of some crucial genes associated with nutrient transportation (such asglucose transporter-2 and sodium-glucose cotransporter-1). Previous studies have suggested that supplementation of botanical compounds increased broiler body weight and hen egg production by approximately 7% and 15%, respectively. Furthermore, the supplementation of botanical compounds enhanced the reproductive efficiency of hens and the semen quality of roosters by 13%. This review article discusses the significant effects of some botanical products in the poultry industry and how they can benefit poultry, especially in light of the ban on antibiotics as growth promoters.

Effects of microalgae, with or without xylanase supplementation, on growth performance, organs development, and gut health parameters of broiler chickens.

Microalgae are becoming potential sustainable feed ingredients, whereas terrestrial feedstuffs are becoming scarce and costly. They are rich in nutritional and functional values but have lower digestibility. This study evaluated the effects of microalgae with or without xylanase supplementation on growth performance and gut health of broiler chickens. A total of 162-day-old Cobb 500 chicks were raised for 35 d. Birds were fed with either 1 of the 3 dietary treatments: 1) corn-soybean meal-based diet (CON), 2) CON + 3% microalgae (MAG), and 3) MAG + xylanase (MAG+XYN) in 2 phases (starter: d 0-21 and finisher: d 22-35) in mash form. Each dietary treatment had 6 replicates, with 9 birds in each replicate. The level of significance was considered at the P value <0.05. The BW, ADG, and ADFI were significantly higher in MAG by 50%, 52.5%, and 42.4%, respectively, and MAG+XYN by 44.1%, 49.7%, and 38.6%, respectively, compared to the CON group. No significant difference was observed for FCR; however, FCR was reduced by 6.3% in both MAG and MAG+XYN groups compared to the CON group. The carcass and organ weight relative to the total body weight were not significantly different among the treatments. The expressions of Zonula occludens 1 (ZO1), Cluster of differentiation 56 (CD56), and Solute carrier family 7 member 7 (SLC7A7) were significantly modulated, for example, by 3.7, 3.9, and 3.3 folds, respectively, in the MAG group compared to CON and 0.8, 0.6, and 1.1 folds, respectively, in the MAG group compared to MAG+XYN groups on d 35. Villi surface area (VSA) of ileum tended to increase on d 3 (P = 0.0725) and d 35 (P = 0.0785) in the MAG and MAG+XYN groups, compared to the CON group. The results suggest that adding microalgae with or without xylanase to broiler's diet could promote growth performance and show a tendency to improve gut health parameters. The nutrient profile and its functional properties make microalgae a valuable resource to the poultry industry as a part substitution of corn and soybean meal and a functional feed supplement to modulate the gut health of broilers.

Dietary supplementation of microalgae mitigates the negative effects of heat stress in broilers.

Heat stress in poultry is a serious concern, affecting their health and productivity. To effectively address the issue of heat stress, it is essential to include antioxidant-rich compounds in the poultry diet to ensure the proper functioning of the redox system. Microalgae (Spirulina platensis) are rich in antioxidants and have several health benefits in humans and animals. However, its role in health and production and the underlying mechanism in heat-stressed broilers are poorly understood. This study aimed to determine the effect of microalgae supplementation on the health and production of heat-stressed broilers. Cobb500 day-old chicks (N = 144) were raised in litter floor pens (6 pens/treatment and 8 birds/pen). The treatment groups were: 1) no heat stress (NHS), 2) heat stress (HS), and 3) heat stress + 3% microalgae (HS+MAG). The broilers in the HS+MAG group were fed a diet supplemented with 3% microalgae, whereas NHS and HS groups were fed a standard broiler diet. Broilers in the NHS were raised under standard temperature (20°C-24°C), while HS and HS+MAG broilers were subjected to cyclic heat stress from d 22 to 35 (32°C-33°C for 8 h). Heat stress significantly decreased the final body weight, whereas the supplementation of microalgae increased the final body weight of broilers (P < 0.05). The expressions of ileal antioxidant (GPX3), immune-related (IL4), and tight-junction (CLDN2) genes were increased in microalgae-supplemented broilers compared to heat-stressed broilers (P < 0.05). The ileal villus height to crypt depth ratio was improved in microalgae-supplemented broilers (P < 0.05). In addition, microbial alpha, and beta diversities were higher in the HS+MAG group compared to the HS group (P < 0.05). There was an increase in volatile fatty acid-producing bacteria at the genus level, such as Ruminococcus, Ocillospira, Lactobacillus, Oscillobacter, Flavonifractor, and Colidextribacter in the group that received microalgae supplementation. In conclusion, dietary supplementation of microalgae improved the growth performances of heat-stressed broilers by improving their physiogenomics. Thus, the dietary inclusion of microalgae can potentially mitigate heat stress in broilers.

Anti-Inflammatory, ACE Inhibitory, Antioxidative Activities and Release of Novel Antihypertensive and Antioxidative Peptides from Whey Protein Hydrolysate with Molecular Interactions.

OBJECTIVE: The aim of the study was to evaluate the whey protein hydrolysate with bio-functional attributes viz. antioxidative, anti-inflammatory and ACE inhibition efficacy and release of bioactive peptides with antioxidative and ACE-inhibitory activity by employing Pepsin. METHOD: The antioxidant, Anti-inflammatory, ACE inhibitory and proteolytic activities of the whey protein hydrolysates were studied followed by SDS-PAGE analysis and IEF. Anti-inflammatory activity of whey protein hydrolysate was also studied on RAW 264.7 cell line. The separation of the bioactive peptides from whey protein hydrolysate was achieved by RP-HPLC. The purified bioactive peptides were identified and characterized using RPLC/MS. RESULTS: WPC (Whey protein concentrate) hydrolysate with pepsin showed proteolytic activity ranging between 14.46 and 18.87 mg/ml. Using the ABTS assay, the highest antioxidative activity was observed in 10 kDa retentate (84.50%) and 3 kDa retentate (85.96%), followed by the highest proteolytic activity (13.83 mg/ml) and ACE inhibitory activity (58.37%) in a 5% WPC solution at 65 °C after 8 h of pepsin hydrolysis. When the protein hydrolysate concentration was low, the production of proinflammatory cytokines by lipopolysaccharide-treated murine macrophages (RAW 264.7) was reduced. SDS-PAGE results exhibited very little protein bands when comparing with WPC hydrolysates to insoluble WPC. There were no protein spots on 2 D gel electrophoresis and "in-solution trypsin digestion" technique have been utilized to digest protein samples directly from WPC hydrolysates. Novel antioxidative peptides and ACE inhibitory peptides were also observed by comparing two databases, i.e., BIOPEP and AHTPDB respectively. The peptide sequences used in this study were found to have excellent potential to be used as inhibitors of hACE as all of them were able to show substantial interactions against the enzyme's active site. CONCLUSIONS: The antihypertensive and antioxidative peptides from whey protein hydrolysates may be beneficial for the future development of physiologically active functional foods. Further, in vivo investigations are required to establish the health claim for each individual bioactive peptide from whey protein hydrolysate.Supplemental data for this article is available online at.

The role of extracellular matrix metalloproteinase inducer (EMMPRIN) in the regulation of bovine endometrial cell functions.

Extracellular matrix metalloproteinase inducer (EMMPRIN) is a cell surface glycoprotein that stimulates the production of several matrix metalloproteinases (MMPs) for tissue remodeling. Previously, we detected EMMPRIN in the bovine endometrium, and it is mainly expressed in the luminal and glandular epithelium whereas MMPs are expressed in the underlying stroma. From this expression pattern, we hypothesized that EMMPRIN may regulate stromal MMPs in endometrial cell functions. To test this hypothesis, a coculture of epithelial and stromal cells was performed using a transwell system. In the coculture, epithelial cells were cultured on the insert membrane and stromal cell on the surface of well plates. Expression of stromal MMP-2 and MMP-14 was significantly higher in coculture with epithelial cell. Further, with the addition of anti-EMMPRIN antibody into the epithelial cell compartment, the expression of stromal EMMPRIN and MMP-2 and MMP-14 was decreased. To identify the active site of EMMPRIN for the augmentation of MMPs, EMMPRIN synthetic peptides that correspond to the extracellular loop domain-I (EM1, EM2, EM3, and EM4) were added into the epithelial cell compartment, and only EM2 at a higher dose interfered with EMMPRIN-mediated expression of MMP-14. Next, we examined the effects of progesterone and/or estrogen on the expression of EMMPRIN, MMP-2, and MMP-14. Progesterone (300 nM) significantly stimulated the expression of EMMPRIN but had no effects on any of the MMPs. These results suggest that EMMPRIN derived from epithelial cells regulates MMPs in the endometrium under progesterone-rich conditions and may thereby modulate bovine endometrial cell functions during gestation.