Long-term music stimulating alleviated the inflammatory responses caused by acute noise stress on the immune organs of broilers by NF-κB signaling pathway.

As an environmental enrichment, music can positively influence the immune function, while noise has an adverse effect on the physical and mental health of humans and animals. However, whether music-enriched environments mitigate noise-induced acute stress remains unclear. To investigate the anti-inflammatory effects of music on the immune organs of broiler chickens under conditions of early-life acute noise stress, 140 one-day-old white feather broilers (AA) were randomly divided into four groups: control (C), the music stimulation (M) group, the acute noise stimulation (N) group, the acute noise stimulation followed by music (NM) group. At 14 days of age, the N and NM groups received 120 dB noise stimulation for 10 min for one week. After acute noise stimulation, the NM group and M group were subjected to continuous music stimulation for 14 days (6 h per day, 60 dB). At 28 days of age, the body temperature of the chicks, the histopathological changes, quantification of ROS-positive density and apoptosis positivity in tissues of spleen, thymus, and bursa of Fabricius (BF) were measured. The results showed that acute noise stimulation led to an increase in the number and area of splenic microsomes and the cortex/medulla ratio of the detected immune organs. The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) of immune tissues of broilers in N group were decreased compared to the broilers in C group, while the mRNA levels of malondialdehyde (MDA), TNF-α, IL-1, and IL-1ß increased. In addition, the gene and protein expression levels of IKK, NF-κB, and IFN-γ of three immune organs from broilers in the N group were increased. Compared to the C and N group, chickens from the NM group showed a decrease in the number and area of splenic follicles, an increase in the activities of SOD and GSH-Px, and a decrease in the expression levels of MDA, TNF-α, IL-1, and IL-1ß. Therefore, a music-enriched environment can attenuate oxidative stress induced by acute noise stimulation, inhibiting the activation of the NF-κB signaling pathway and consequently alleviating the inflammatory response in immune organs.

Lysinibacillus capsici 38,328 isolated from agricultural soils as a promising probiotic candidate for intestinal health.

There is an increasing interest in the use of spore-forming Bacillus spp. as probiotic ingredients on the market. However, probiotics Bacillus species are insufficient, and more safe Bacillus species were required. In the study, traditional fermented foods and soil samples were collected from more than ten provinces in China, and 506 Bacillus were selected from 109 samples. Using the optimized procedure, we screened nine strains, which successfully passed the acid, alkali, bile salt, and trypsin resistance test. Drug sensitivity test results showed that three Bacillus out of the nine isolates exhibited antibiotic sensitivity to more than 29 antibiotics. The three strains sensitive to antibiotics were identified by 16S ribosomal RNA, recA, and gyrB gene analysis, two isolates (38,327 and 38,328) belong to the species Lysinibacillus capsici and one isolate (37,326) belong to Bacillus halotolerans. Moreover, the three strains were confirmed safe through animal experiments. Finally, L. capsici 38,327 and 38,328 showed protections in the Salmonella typhimurium infection mouse model, which slowed down weight loss, reduced bacterial load, and improved antioxidant capacity. Altogether, our data demonstrated that selected L. capsici strains can be used as novel probiotics for intestinal health.

Effects of Catecholamine Stress Hormones Norepinephrine and Epinephrine on Growth, Antimicrobial Susceptibility, Biofilm Formation, and Gene Expressions of Enterotoxigenic Escherichia coli.

Enterotoxigenic Escherichia coli (ETEC) is a significant contributor to diarrhea. To determine whether ETEC-catecholamine hormone interactions contribute to the development of diarrhea, we tested the effects of catecholamine hormones acting on ETEC in vitro. The results showed that in the presence of norepinephrine (NE) and epinephrine (Epi), the growth of 9 out of 10 ETEC isolates was promoted, the MICs of more than 60% of the isolates to 6 antibiotics significantly increased, and the biofilm formation ability of 10 ETEC isolates was also promoted. In addition, NE and Epi also significantly upregulated the expression of the virulence genes feaG, estA, estB, and elt. Transcriptome analysis revealed that the expression of 290 genes was affected by NE. These data demonstrated that catecholamine hormones may augment the diarrhea caused by ETEC.

Ameliorative effects of dietary selenium against cadmium toxicity on production performance and egg quality in laying hens.

In order to reveal the influences of supplemented dietary selenium (Se) on the suppressive effect of cadmium (Cd) toxicity on performance and egg properties of laying hens, the effects of co-treatment Se and Cd on the performance, egg quality, levels of amino acids and the antioxidant capacity of egg and serum were investigated. A total of 128 31-week-old laying hens were randomly distributed in four treatments, which were fed with the basic diet (0.2 mg/kg Se and 0.08 mg/kg Cd), and the basic diet with Se (1.1 mg/kg Se and 0.08 mg/kg Cd), Cd (0.2 mg/kg Se and 92.1 mg/kg Cd) and Se+Cd for 13 weeks, respectively. Hens supplemented with Cd led to an impairment on production performance and egg quality with decreased egg production (EP), egg mass (EM), feed intake (FI), eggshell color, eggshell thickness, yolk color, albumen height and haugh unit and increased the feed conversion ratio (FCR) (p < 0.05). Cd treatment decreased the contents of cysteine (Cys), histidine (His), lithium (Li), aluminum (Al), chromium (Cr), manganese (Mn), nickel (Ni), zinc (Zn), arsenic (As), Se, strontium (Sr), stannum (Sn), mercury (Hg) and thallium (Tl) and increased the contents of isoleucine (Ile) and Cd (p < 0.05). Cd destroyed the egg yolk and serum redox states with the increased concentration of malondialdehyde (MDA) and the decreased activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx) (p < 0.05). The expression levels of ovarian apoptotic genes (protein 53, Caspase9, Cytochrome c and Bcl-2 associated X protein) increased, and B-cell lymphoma 2 (Bcl-2) expression decreased in the Cd group (p < 0.05). Feeding Se significantly alleviated Cd-induced toxicity on performance and egg quality. Se+Cd treatment restored the balance between oxidation and antioxidant systems and modulated the elements' homeostasis and alleviated the changes in apoptotic-related genes expression levels. Se could alleviate the Cd toxicity to laying hens and their eggs but could not counteract all negative effects of Cd.

Selenomethionine protects against ammonia-induced apoptosis through inhibition of endoplasmic reticulum stress in pig kidneys.

Ammonia (NH3) emission is a common threat to farm animals. Selenium (Se) is known for its antioxidant property and can resist several stressors affecting farm animals. The aims of this study were (Ⅰ) to determine how excess NH3 exert nephrotoxic effects in pigs and (Ⅱ) to investigate whether selenomethionine has an alleviative effect on NH3 toxicity. Two diets supplemented with different doses of Se (0.22 mg/kg or 0.50 mg/kg) and two concentrations of NH3 (< 5 mg/m3 or 89.8 mg/m3) were used in a 2 × 2 factorial design trial for a period of 30 days. The results showed that NH3 exposure caused apoptosis and increased the number of apoptotic cells in pig kidneys. Further, the activities of antioxidant enzymes were decreased, and the transcriptional and translational levels of endoplasmic reticulum stress-related genes, Bcl-2 and Caspase family members were increased under NH3 exposure. In addition, Wnt/ß-catenin signaling pathway was suppressed after NH3 treatment. Dietary supplement with selenomethionine appears to offer protection against NH3-induced kidney injury in pigs and the pathologic changes above were alleviated. Our findings provide additional insight into the mechanism of NH3 toxicity in pigs while elucidating the role of Se as a potential antidote against NH3 poisoning.

Exposure to the environmental pollutant ammonia causes changes in gut microbiota and inflammatory markers in fattening pigs.

Ammonia (NH3) is a major pollutant in livestock houses and atmospheric environment. It has been demonstrated that NH3 can cause a series of damage to animals and human. However, toxicity evaluation of NH3 on farm animals was rarely reported, especially in the intestinal microflora. Therefore, in this study, twenty-four 125-day-old fattening pigs were randomly divided into 4 groups: control group, NH3 group (88.2 mg m-3 < NH3 concentration < 90.4 mg m-3), Se group (Se content: 0.5 mg kg-1), and NH3 + Se group (88.2 mg m-3 < NH3 concentration < 90.4 mg m-3, Se content: 0.5 mg kg-1), and the effects of NH3 and L-Selenomethionine on the microbiota composition in the jejunum and the levels of inflammatory markers in feces of fattening pigs were examined by 16S rDNA and ELISA, respectively. Our results showed that the content of Matrix metalloproteinase-9 (MMP-9), Myeloperoxidase (MPO), Lactoferrin (LTF) and Calprotectin in the ammonia group (A group) were significantly elevated compared to the control group, and the content of MMP-9, MPO, LTF and Calprotectin in the A + Se group were significantly reduced. A significant difference in microbiota composition in the phylum, class, family and genus levels was found in the A group and the NH3 + Se group. There was a negative correlation between Streptococcus and Calprotectin. Our results indicated that excessive NH3 inhalation could cause changes in inflammatory markers and beta diversity of intestinal microflora in fattening pigs. We found there was a positive correlation between MPO and Pseudomonas. In addition, we first proposed that L-Selenomethionine could improve the imbalance of microbial flora and the inflammatory injury caused by NH3. Changes in intestinal microflora and inflammatory markers can be used as important indicators to evaluate NH3 toxicity, and studying changes in intestinal microflora is also an important mechanism to reveal NH3 toxicity.

Ammonia exposure causes the imbalance of the gut-brain axis by altering gene networks associated with oxidative metabolism, inflammation and apoptosis.

Ammonia is an acknowledged environment pollutant in atmosphere with irritating smell. Previous studies have shown that excessive ammonia has toxic effects on farm animals and humans. However, the detail toxicity mechanism of ammonia to pigs is still unknown so far. In order to clarify the mechanism of ammonia toxicity, we established a porcine exogenous ammonia poisoning model and assessed the effects of ammonia on the gut-brain axis by transcriptome sequencing, histological observation and chemical analysis. Our results showed that after 30 d of ammonia exposure, 578 differentially expressed genes (DEGs) and 407 DEGs were obtained in the hypothalamus and jejunum, respectively. These DEGs were enriched into Gene Ontology terms associated with inflammation, oxidative metabolism, apoptosis, and the highly expressed genes among these DEGs were verified by real-time quantitative PCR. The content of glutathione and the activities of glutathione peroxidase and superoxide dismutase were significantly decreased, while malondialdehyde content was increased after ammonia exposure. Corticotropin releasing factor, substance P, 5-hydroxytryptamine and ghrelin contents in serum elevated significantly. Furthermore, pathologic observation in the ammonia group revealed infiltration of lymphocytes in the hypothalamus and significant decrease of jejunal epithelial cells. Our results indicated that ammonia exposure mediated changes in transcriptional profiles, pathological damage, oxidative stress and brain-gut peptide of the pig jejunum and hypothalamus, and induced the imbalance of the brain-gut axis through the "oxidative stress-inflammation-apoptosis" interaction network. Our study not only provides a new perspective for the toxicity assessment of ammonia, but also enriches the toxicology mechanism of ammonia.

Pre-cold acclimation improves the immune function of trachea and resistance to cold stress in broilers.

Acclimation can alleviate the damage caused by adverse environmental factors. To investigate the effects of cold stimulation on immunity in tracheal of broilers, 360 one-day-old chicks were raised at normal temperatures during 1-7 days. From 8 day, G1 (control) continued to be raised at normal temperatures, whereas G2 and G3 (treatment groups) were cold-stimulated at 3°C and 12°C below the temperature of G1, respectively. At 42 day, all the groups were subjected to a 24-hr acute cold stress, designated as S1, S2, and S3. Tracheal tissues were collected to detect gene levels of immunoglobulins, antimicrobial peptides, Hsps, and cytokines, and oxidative stress-related indicators at 14 day, 42 day, and 43 day, and protein levels of Hsps and proinflammatory cytokines as well as morphology changes at 42 day and 43 day. The results showed that, compared with 42G1, tracheal structure of 42G2 was basically intact, and gene levels of immunoglobulins and antimicrobial peptides increased (p < 0.05), whereas tracheal structure of 42G3 was destroyed, with decreased levels of immunoglobulins ( p < 0.05), and increased levels of Hsps and proinflammatory cytokines ( p < 0.05). At 43 day, tracheal damage was visible and gene levels of immunoglobulins and antimicrobial peptides decreased in S1 ( p < 0.05). Tracheal structure was relatively intact and gene levels of antimicrobial peptides increased in S2 ( p < 0.05). Compared with S1 and S3, immune-related gene levels in S2 were higher, and Hsps and proinflammatory cytokines levels were lower. The results demonstrate that cold stimulation of lower 3°C from 8 to 42 day led to cold acclimation, which improved immunity of tracheal mucosa and resistance to cold stress in broilers.

The protective role of selenium against cadmium-induced hepatotoxicity in laying hens: Expression of Hsps and inflammation-related genes and modulation of elements homeostasis.

The purpose of this study was to examine the potential role of high selenium (Se) diets in alleviating chronic cadmium (Cd) hepatic toxicity in laying hens. In the present study, 128 healthy 31-week-old laying hens were fed a diet supplemented with Se (Na2SeO3, 2 mg/kg), Cd (CdCl2, 150 mg/kg), or both Se and Cd (150 mg/kg of CdCl2 and 2 mg/kg of Na2SeO3) for 90 days. The expression levels of heat shock proteins (Hsps, including Hsp60, Hsp70 and Hsp90) and inflammation-related factors, including nuclear factor-kappa B p50 (NF-κB), cyclooxygenase-2 (COX-2), prostaglandin E synthases (PTGES), interleukin 1-beta (IL-1ß), and tumor necrosis factor-α (TNF-α) were investigated. The concentrations of 28 elements were also determined. The results indicated that Cd treatment significantly increased the mRNA and protein expression levels of Hsps and significantly improved the expression of inflammation-related genes. Moreover, Cd addition to the diets resulted in disturbances in the systemic balance of 13 elements, leading to decrease in the concentrations of Cr, Mn, Sr, Ba, and Hg and increase in Li, B, Ca, Ti, Fe, Cu, Mo, and Cd concentrations. Treatment with Se significantly alleviated Cd-induced hepatic toxicity, as evidenced by a reduction in Hsp60, Hsp70, Hsp90, NF-κB, COX-2, PTGES, TNF-α, and IL-1ß expression. Additionally, Se and Cd co-treatment alleviated the changes in Li, B, Ca, Fe, Ti, Cu, Mo, Cd, Cr, Se, Sr, Ba, and Hg concentrations, which was in contrast to that upon Cd induction. The study indicated that Se could help against the negative effects of Cd and may be related to the alleviation of Cd-induced Hsps stress and the inflammatory responses along with modulating the element homeostasis.

Exterior egg quality as affected by enrichment resources layout in furnished laying-hen cages.

OBJECTIVE: This study aimed to investigate the effects of enrichment resources (a perch, dustbath, and nest) layout in furnished laying-hen cages (FC) on exterior quality of eggs. METHODS: One hundred and sixty-eight (168) Hy-Line Brown laying hens at 16 weeks of age were randomly distributed to four treatments: small furnished cages (SFC), medium furnished cages type I (MFC-I), medium furnished cages type II (MFC-II), and medium furnished cages type III (MFC-III). Each treatment had 4 replicates or cages with 6 hens for SFC (24 birds for each SFC) and 12 hen/cage for MFC-I, -II, and -III (48 birds for each MFC-I, -II and -III). Following a 2-week acclimation, data collection started at 18 weeks of age and continued till 52 weeks of age. Dirtiness of egg surface or cracked shell as indicators of the exterior egg quality were recorded each week. RESULTS: The results showed that the proportion of cracked or dirty eggs was significantly affected by the FC type (p<0.01) in that the highest proportion of cracked or dirty eggs was found in MFC-I and the lowest proportion of dirty eggs in SFC. The results of this showed that furnished cage types affected both dirty eggs and cracked eggs (p<0.01). The results also indicated that not nest but dustbath lead to more dirty eggs. Only MFC-I had higher dirty eggs at nest than other FC (p< 0.01). The results of dirty eggs in MFC-I and MFC-II compared with SFC and MFC-III seemed suggest that a low position of dustbath led to more dirty eggs. CONCLUSION: SFC design affected exterior egg quality and the low position of dustbath in FC resulted in higher proportion of dirty eggs.

Isolation and complete nucleotide sequence of a Batai virus strain in Inner Mongolia, China.

BACKGROUND: Batai virus (BATV) is a member of the Orthobunyavirus genus of the family Bunyaviridae, and a vector-borne pathogen. Genomic variations of BATV exist in different regions of the world, due to genetic reassortment. Whole-genome sequencing of any isolate is necessary for a phylogenetic analysis. In 1998, a BATV strain was isolated from an Anopheles philippines mosquito in Yunnan Province, China. This strain has not been found to infect any other host. We investigated BATV infection in cattle in Inner Mongolia, China and performed deep sequencing of the genome of the BATV isolate. FINDINGS: Ninety-five blood samples were collected from cattle in Inner Mongolia, China in 2012. The BATV infection rate was 2.1%. Previously, BATV strain NM/12 was isolated from two cattle in Inner Mongolia, China, and the whole genomic sequence of the strain has been available. We determined the complete genomic nucleotide sequences of the small (S), medium (M), and large (L) genome segments using bovine blood obtained in 2012, and the nucleotide homologies of these segments with those from GenBank were 88.7%-97%, 84%-95.4%, and 72.6%-95.8%, respectively. The deduced amino acid identities were 87.2-99.7%, 64.2-96.8%, and 81.1-98.6%. Phylogenetic analyses based on full-length genomic sequences indicated that the M and L segments, and a portion of the S segment, of NM/12 are most closely related to the BATV strains isolated in Asia. The S and M segments of NM/12 were independent of phylogenetic lineages. The L segment was the most closely related to Chittoor/IG-20217 (isolated in India), and distantly related to isolated strains in Italy. Nucleotide substitution rates in the nucleotide sequences that code for the nucleocapsid, envelope glycoprotein, and polymerase protein of NM/12 strain were 2.56%, 4.69%, and 4.21%, respectively, relative to the original strain of MM2222. CONCLUSION: A novel BATV NM/12 strain from bovine serum collected in Inner Mongolia was isolated from cattle in China for the first time. Our findings elucidate the evolutionary status of the BATV NM/12 strain among different orthobunyavirus strains and may provide some clues to prevent the emergence of BATV infection in cattle and humans.

Research Note: Effects of environmental sound stimulus on behavioral responses, cortisol levels, and horizontal immunity of transferred pullets.

The aim of this study was to investigate the effects of prolonged exposure to varying levels of music or noise on the behavioral, physiological, and immune responses of pullets following their transfer to an egg-laying facility. A total of 240 one-day-old Hy-Line Brown pullets were randomly assigned to five groups: 0 dB sound stimulation, low-decibel music (65-75 dB), high-decibel music (85-95 dB), low-decibel noise (65-75 dB), and high-decibel noise (85-95 dB) stimuli. Pullets received music or noise stimuli 10 h per d from 1-day-old to 16-wk-old and were then transferred to the egg-laying facility. The results indicated that feeding and drinking behaviors significantly decreased (P < 0.05), whereas feather pecking, aggression, and preening behaviors significantly increased (P < 0.05) in the pullets after transfer. Pullets also had higher serum cortisol (COR) levels (P < 0.05), whereas immunoglobulin Y (IgY), tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ), interleukin-1ß (IL-1ß), interleukin-2 (IL-2) and interleukin-6 (IL-6) levels significantly increased (P < 0.05). Low-decibel sound stimuli increased aggressive behavior and decreased pecking behavior (P < 0.05). High-decibel sound stimuli decreased feather pecking, cage pecking, aggression, and sham dustbathing behaviors (P < 0.05). In addition, a low-decibel sound stimulus decreased the serum COR content, and increased the serum IL-6 level in the transferred pullets. A high-decibel sound stimulus also induced shorter tonic immobility (TI) durations in pullets on d 7 after transport stress. Meanwhile, high-decibel sound stimulus decreased the serum IL-6 and TNF-α levels of pullets. In conclusion, the transfer has detrimental effects on the pullets. Long-term sound stimulation effectively mitigated the negative impact of transportation stress on pullets. Among them, the high-decibel sound stimulus showed more promise in relieving transport stress.

The inflammatory immunity and gut microbiota are associated with fear response differences in laying hens.

The fear response is a crucial adaptive mechanism for coping with environmental changes, and the individuals have different levels of fearfulness. The purpose of this study was to determine the status of the immune response and gut health in hens with different fear responses. A total of 80 healthy 75-wk-old native Lindian chickens were individually housed in conventional cages and categorized into high (TH) and low (TL) levels of fearfulness using the tonic immobility (TI) test. The immunological status and intestinal health of the laying hens were assessed, and the intestinal microbial community was sequenced using 16S rRNA testing. The results showed that the immune-related genes of interleukin (IL)-1ß, IL-4, IL-6, and IgG were significantly upregulated in the spleen of TH hens compared with hens in the TL group (P < 0.01). The inflammatory immune-related genes Toll-like receptor (TLR)2, TLR4, nuclear factor (NF)-κB, inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, IL-10, and IgG were significantly increased in the intestinal tract, whereas IL-4, IgA, and the intestinal barrier gene claudin-4 were significantly decreased in TH hens (P < 0.05). In addition, serum concentrations of IL-1ß, IL-6, IL-10, interferon (IFN)-α and IgG were significantly higher in TH hens (P < 0.01). A high fear response also led to changes in gut microbial diversity, with a higher Simpson's index and lower ß-diversity similarity than hens with a low-fear response (P < 0.05). The TH group showed an increase in 8 genera, including Bacillaceae and Coprococcus, whereas the genus Anaerorhabdus decreased (P < 0.05). The gut microbiota has also been associated with gut barrier genes, and inflammatory cytokines. Bartonella stimulates IL-1ß and IgG secretion, whereas Lactobacillus inhibits IL-6 secretion, and Coprococcus and Subdoligranulum are associated with the maintenance of intestinal barrier function. The results of this study suggest that laying hens with high fear response levels have a more sensitive immune response and a more enriched gut microbiota, which may have positive effects on adapting to a complex environment.

Characterization of the optimized C2 domain of protein G: finding its additional chicken IgY-binding ability.

The C2 domain of streptococcal protein G is a small (55 residue) peptide with immunoglobulin-binding activity. Following codon optimization, the gene was divided into four oligonucleotide fragments and amplified by overlap PCR. The recombinant plasmid pET30a-C2 was transformed into Escherichia coli Rosetta (DE3) PLysS for expression. After purification by Ni-NTA, the fusion protein was identified by western-blotting, Dot-ELISA and ELISA. His-tagged C2 bound to human, rabbit, cattle, pig, goat, mouse or guinea pig IgG had no affinity for goose, duck, wild duck, wild turkey and red-crowned crane IgY. Its affinity for chicken IgY, however, was comparable to that of guinea pig IgG. The C2 domain may therefore provide an ideal material for the purification and detection of immunoglobulin G from various mammals.

The Effect of Short-term Classical Music stimulus on Behavior and Tonic Immobility Reaction of Pullets.

The purpose of this study was to investigate the effect of short-term stimulation of classical music on the behavior of pullets and to explore whether classical music as an auditory enrichment factor reduces the fear level in pullets. One hundred and twenty 10-weeks'old Roman white pullets were randomly divided into two treatments of which one group was exposed to classic music (M), and another one was the control group (N). The music was played during 8:00-18:00 every day for 7 consecutive days. The behavior of the focal animals was observed from 11:00 to13:00 every day for 7 days. The results showed that during the observation period, the pullets in group M had more comforting (P < .05) and preening (P < .05), but less aggressive (P < .01) and feather-pecking (P < .01) behaviors than those in group N. No significant difference was found in other behaviors and the duration of tonic immobility between the two groups. Therefore, auditory enrichment as a means of environmental enrichment can increase the welfare level of pullets to a certain extent.

Serum bone remodeling parameters and transcriptome profiling reveal abnormal bone metabolism associated with keel bone fractures in laying hens.

Keel bone fractures affect welfare, health, and production performance in laying hens. A total of one hundred and twenty 35-wk-old Hy-line Brown laying hens with normal keel (NK) bone were housed in furnished cages and studied for ten weeks to investigate the underlying mechanism of keel bone fractures. At 45 wk of age, the keel bone state of birds was assessed by palpation and X-ray, and laying hens were recognized as NK and fractured keel (FK) birds according to the presence or absence of fractures in keel bone. The serum samples of 10 NK and 10 FK birds were collected to determine bone metabolism-related indexes and slaughtered to collect keel bones for RNA-sequencing (RNA-seq), Micro-CT, and histopathological staining analyses. The results showed that the concentrations of Ca, phosphorus, calcitonin, 25-hydroxyvitamin D3, and osteocalcin and activities of alkaline phosphatase and tartrate-resistant acid phosphatase (TRAP) in serum samples of FK birds were lower than those of NK birds (P < 0.05), but the concentrations of parathyroid hormone, osteoprotegerin, and corticosterone in serum samples of FK birds were higher than those of NK birds (P < 0.05). TRAP staining displayed that FK bone increased the number of osteoclasts (P < 0.05). Micro-CT analysis indicated that FK bone decreased bone mineral density (P < 0.05). Transcriptome sequencing analysis of NK and FK bones identified 214 differentially expressed genes (DEGs) (|log2FoldChange| > 1, P < 0.05), among which 88 were upregulated and 126 downregulated. Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) analysis indicated that 14 DEGs related to skeletal muscle movement and bone Ca transport (COL6A1, COL6A2, COL6A3, PDGFA, MYLK2, EGF, CAV3, ADRA1D, BDKRB1, CACNA1S, TNN, TNNC1, TNNC2, and RYR3) were enriched in focal adhesion and Ca signaling pathway, regulating bone quality. This study suggests that abnormal bone metabolism related to keel bone fractures is possibly responded to fracture healing in laying hens.

Effect of Noise and Music on Neurotransmitters in the Amygdala: The Role Auditory Stimuli Play in Emotion Regulation.

Stress caused by noise is becoming widespread globally. Noise may lead to deafness, endocrine disorders, neurological diseases, and a decline in mental health. The mechanism behind noise-induced neurodevelopmental abnormalities is unclear, but apoptosis and pro-inflammatory signals may play an important role. In this study, weaned piglets were used as a model to explore noise-induced neurodevelopmental abnormalities. We hypothesized that long-term noise exposure would induce anxiety and cause acute stress, exhibited by alterations in neurotransmission in the amygdala. A total of 72 hybrid piglets (Large White × Duroc × Min Pig) were randomly divided into three groups, including noise (exposed to mechanical noise, 80-85 dB), control (blank, exposed to natural background sound, <40 dB), and music (positive control, exposed to Mozart K.448, 60-70 dB) groups. The piglets were exposed to 6 h of auditory noise daily (10:00-16:00) for 28 days. Compared with the control group, piglets exposed to noise showed more aggressive behavior. The expression of Caspase3, Caspase9, Bax, NF-κB (p56), TLR4, MYD88, I κ B α, IL-1 ß, TNF-α, and IL-12RB2 was significantly upregulated in the amygdala, while the expression of Nrf2, HO-1, CAT, and SOD was downregulated in piglets in the noise group. Cell death occurred, and numerous inflammatory cells accumulated in the amygdala of piglets in the noise group. Targeted metabolomics showed that the content of inhibitory neurotransmitter GABA was higher in the amygdala of piglets in the noise group. Compared with the noise group, piglets in the music group displayed more positive emotion-related behaviors. Compared with the noise group, the expression of genes related to apoptosis, inflammation, and oxidative damage was lower in the music group. Cells of the amygdala in the music group were also of normal morphology. Our results show that noise-induced stress causes apoptosis and neuroinflammation in the amygdala and induces anxiety during the early neonatal neural development of piglets. In contrast, to some extent, music alleviates noise-induced anxiety.

Biological Activity of Optimized Codon Bovine Type III Interferon Expressed in Pichia pastoris.

Type III interferons (IFN-λs) exhibit potent antiviral activity and immunomodulatory effects in specific cells. Nucleotide fragments of the bovine ifn-λ (boifn-λ) gene were synthetized after codon optimization. The boifn-λ gene was then amplified by splicing using overlap extension PCR (SOE PCR), resulting in the serendipitous acquisition of the mutated boIFN-λ3V18M. The recombinant plasmid pPICZαA-boIFN-λ3/λ3V18M was constructed, and the corresponding proteins were expressed in Pichia pastoris with a high-level extracellular soluble form. Dominant expression strains of boIFN-λ3/λ3V18M were selected by Western blot and ELISA and cultured on a large scale, and the recombinant proteins purified by ammonium sulfate precipitation and ion exchange chromatography yielded 1.5g/L and 0.3 g/L, with 85% and 92% purity, respectively. The antiviral activity of boIFN-λ3/λ3V18M exceeded 106 U/mg, and they were neutralized with IFN-λ3 polyclonal antibodies, were susceptible to trypsin, and retained stability within defined pH and temperature ranges. Furthermore, boIFN-λ3/λ3V18M exerted antiproliferative effects on MDBK cells without cytotoxicity at 104 U/mL. Overall, boIFN-λ3 and boIFN-λ3V18M did not differ substantially in biological activity, except for reduced glycosylation of the latter. The development of boIFN-λ3 and comparative evaluation with the mutant provide theoretical insights into the antiviral mechanisms of boIFN-λs and provide material for therapeutic development.

Gut Microbiota-Derived 5-Hydroxyindoleacetic Acid Alleviates Diarrhea in Piglets via the Aryl Hydrocarbon Receptor Pathway.

With the improvement in sow prolificacy, formula feeding has been increasingly used in the pig industry. Diarrhea remains a serious health concern in formula-fed (FF) piglets. Fecal microbiota transplantation (FMT) is an efficacious strategy to reshape gut microbiota and the metabolic profile for treating diarrhea. This study aims to investigate whether FMT from breast-fed piglets could alleviate diarrhea in FF piglets. The piglets were randomly assigned to the control (CON) group, FF group, and FMT group. Our results showed that FF piglets exhibited a higher diarrhea incidence, damaged colonic morphology, and disrupted barrier function. In contrast, FMT treatment normalized the morphology and barrier function. FMT suppressed the JNK/MAPK pathway and production of proinflammatory cytokines. Additionally, FF piglets had a lower abundance of the beneficial bacterial genus Bifidobacterium compared to CON piglets. Following FMT administration, Bifidobacterium was restored. Meanwhile, 5-HIAA, a metabolite of tryptophan, and AHR-responsive CYP1A1 and CYP1B1 were upregulated. Importantly, integrated multiomics analysis revealed a strong positive correlation between Bifidobacterium and 5-HIAA. In vitro, 5-HIAA supplementation reversed the LPS-induced disruption of tight junctions and production of proinflammatory cytokines in IPEC-J2 cells. In conclusion, FMT reduced diarrhea incidence and improved growth performance. The alleviative effect of FMT on diarrhea was associated with Bifidobacterium and 5-HIAA.

Effects of dustbathing environment on gut microbiota and expression of intestinal barrier and immune-related genes of adult laying hens housed individually in modified traditional cage.

Litters, the composition of sand and peat rich in microbiota, are essential to trigger the dustbathing behavior of chickens. To investigate the effects of a dustbathing environment (DE) on the intestinal health, gut microbiota, and immune responses of laying hens, a total of 72 healthy Hy-Line Brown laying hens at 69 wk of age (WOA) were housed individually in modified traditional cages and randomly divided into 2 groups: one group had free access to litters (CT), while the other one was restricted from litters (CC). The experiment lasted for 42 d. At the end of the experiment, the intestinal histomorphology and immune status of laying hens were determined, and the 16S rRNA sequencing method was used to assess the composition of the intestinal microbial community of birds. Intestinal histomorphology changed, including villus height and villus-to-crypt ratio significantly increased in the CT group (P < 0.01). DE reshaped the microbial community and increased the microbial richness with the higher indicators of Chao1 and observed species and the comparatively abundant beta diversity (P < 0.05). Ten genera, including Faecalibacterium and Coprococcus, declined in laying hens from the CT group (P < 0.05), while Alistipes increased in CT hens (P < 0.05) compared to those hens from the CC group. The expression levels of intestinal barrier-related genes of claudin-1, claudin-4, occludin, ZO-1, and ZO-2 and immune-related genes of IL-4, IL-6, IL-8, IFN-γ, IgA, TLR-2, and TLR-4 were significantly upregulated in the intestine of laying hens in CT group (P < 0.05). DE also increased the serum levels of IL-4, IL-6, IL-8, IFN-γ, and IgA (P < 0.01). The alteration of the gut microbiota by DE is closely related to host immune responses, including Lactobacillus positively correlated with IL-4 and IgA. Thus, a dustbathing environment can improve the welfare of laying hens by changing the intestinal histomorphology, immune response, and the gut microbial community.