Genetically modified pigs:Emerging animal models for hereditary hearing loss

Hereditary hearing loss(HHL),a genetic disorder that impairs auditory function,significantly affects quality of life and incurs substantial economic losses for society.To investigate the underlying causes of HHL and evaluate therapeutic outcomes,appropriate animal models are necessary.Pigs have been extensively used as valuable large animal models in biomedical research.In this review,we highlight the advantages of pig models in terms of ear anatomy,inner ear morphology,and electrophysiological characteristics,as well as recent advancements in the development of distinct genetically modified porcine models of hearing loss.Additionally,we discuss the prospects,challenges,and recommendations regarding the use pig models in HHL research.Overall,this review provides insights and perspectives for future studies on HHL using porcine models.

Evaluation of the Pharmacological Potential of Strychnos camptoneura Trunk Bark Hydroethanol Extract on Reproductive Functions in Male Guinea Pigs (Cavia porcellus) Exposed to Cypermethrin

The main objective of this study was to evaluate the pharmacological potential of the hydroethanol extract of Strychnos camptoneura trunk bark on reproductive functions in male guinea pigs exposed to cypermethrin. The results showed that administration of the hydroethanolic extract (100 and 250 mg/kg) of Strychnos camptoneura trunk bark after exposure of the animals to cypermethrin induced a highly significant (p Strychnos camptoneura trunk bark produced a highly significant (p Strychnos camptoneura trunk bark may have protective effects against cypermethrin-induced male infertility due to its androgenic, spermatogenic and antiradical properties.

Preliminary study of metabonomic changes during the progression of atherosclerosis in miniature pigs

Background:To explore potential biomarkers for early diagnosis of atherosclerosis(AS)and provide basic data for further research on AS,the characteristics of serum metabolomics during the progression of AS in mini-pigs were observed dynamically.Methods:An AS model in Bama miniature pigs was established by a high-cholesterol and high-fat diet.Fasting serum samples were collected monthly for metabolomics and serum lipid detection.At the end of the treatment period,pathological analysis of the abdominal aorta and coronary artery was performed to evaluate the lesions of AS,thereby distinguishing the susceptibility of mini-pigs to AS.The metabolomics was de-tected using a high-resolution untargeted metabolomic approach.Statistical analysis was used to identify metabolites associated with AS susceptibility.Results:Based on pathological analysis,mini-pigs were divided into two groups:a susceptible group(n=3)and a non-susceptible group(n=6).A total of 1318 metabo-lites were identified,with significant shifting of metabolic profiles over time in both groups.Dynamic monitoring analysis highlighted 57 metabolites that exhibited an ob-vious trend of differential changes between two groups with the advance of time.The KEGG(Kyoto Encyclopedia of Genes and Genomes)pathway enrichment analysis in-dicated significant disorders in cholesterol metabolism,primary bile acid metabolism,histidine metabolism,as well as taurine and hypotaurine metabolism.Conclusions:During the progression of AS in mini-pigs induced by high-cholesterol/high-fat diet,the alterations in serum metabolic profile exhibited a time-dependent pattern,accompanied by notable disturbances in lipid metabolism,cholesterol me-tabolism,and amino acid metabolism.These metabolites may become potential bio-markers for early diagnosis of AS.

Associations of genome-wide structural variations with phenotypic differences in cross-bred Eurasian pigs

Background During approximately 10,000 years of domestication and selection,a large number of structural variations(SVs)have emerged in the genome of pig breeds,profoundly influencing their phenotypes and the ability to adapt to the local environment.SVs(≥50 bp)are widely distributed in the genome,mainly in the form of insertion(INS),mobile element insertion(MEI),deletion(DEL),duplication(DUP),inversion(INV),and translocation(TRA).While studies have investigated the SVs in pig genomes,genome-wide association studies(GWAS)-based on SVs have been rarely conducted.Results Here,we obtained a high-quality SV map containing 123,151 SVs from 15 Large White and 15 Min pigs through integrating the power of several SV tools,with 53.95%of the SVs being reported for the first time.These high-quality SVs were used to recover the population genetic structure,confirming the accuracy of genotyping.Potential functional SV loci were then identified based on positional effects and breed stratification.Finally,GWAS were performed for 36 traits by genotyping the screened potential causal loci in the F2 population according to their corresponding genomic positions.We identified a large number of loci involved in 8 carcass traits and 6 skeletal traits on chromosome 7,with FKBP5 containing the most significant SV locus for almost all traits.In addition,we found several significant loci in intramuscular fat,abdominal circumference,heart weight,and liver weight,etc.Conclusions We constructed a high-quality SV map using high-coverage sequencing data and then analyzed them by performing GWAS for 25 carcass traits,7 skeletal traits,and 4 meat quality traits to determine that SVs may affect body size between European and Chinese pig breeds.

Excess dietary Lys reduces feed intake, stimulates jejunal CCK secretion and alters essential and non‑essential blood AA profile in pigs

Background Commercial diets are frequently formulated to meet or exceed nutrient levels including those of lim-iting essential amino acids(AA)covering potential individual variations within the herd.However,the provision of dietary excess of AA,such as Lys,may lead to reduced appetite and growth in pigs.The mechanisms modulat-ing these responses have not been extensively investigated.This study evaluated the effect of Lys dietary excesses on performance and satiety biomarkers in post weaning pigs.Methods Twenty-four pigs aged 21 d and weighing 6.81±0.12kg(mean±SEM)were individually housed and offered 1 of 4 dietary treatments for 3weeks:a diet containing a standardized ileal digestible Lys reaching 100%(T0),120%(T1),150%(T2)or 200%(T3)of the NRC(2012)requirements.At the end of the experiment,blood samples from the cephalic vein of the T0 and T3 groups were obtained for AA analysis.In addition,primary intestinal cultures from T0 pigs were used,following their humane killing,to evaluate the effect of Lys on gut hormone secretion and AA sensors gene expression under ex vivo conditions.Results Feed intake was linearly reduced(P<0.001)and the weight gain to feed ratio reduced(P<0.10)with increased dietary levels of Lys during the third-and first-week post weaning,respectively.Cholecystokinin con-centration(P<0.05)and the metabotropic glutamate receptor 1 and the solute carrier family 7 member 2(P<0.10)gene expression was enhanced in proximal jejunum tissues incubated with Lys at 20mmol/L when compared to the control(Lys 0mmol/L).Plasma Lys and Glu(P<0.05)concentration increased in the T3 compared to T0 pigs.In contrast,plasma levels of His,Val,Thr,Leu(P<0.05)and Gln(P<0.10)were lower in T3 than T0 pigs.Conclusion The present results confirm that excess dietary Lys inhibits hunger in pigs.Moreover,the results provide evidence of pre-and post-absorptive mechanisms modulating these responses.Lys dietary excesses should be nar-rowed,when possible,to avoid negative effects of the AA on appetite in pigs.

Investigation of the nutritional and functional roles of a combinational use of xylanase and β-glucanase on intestinal health and growth of nursery pigs

Background Xylanase andβ-glucanase combination(XG)hydrolyzes soluble non-starch polysaccharides that are anti-nutritional compounds.This study aimed to evaluate the effects of increasing levels of XG on intestinal health and growth performance of nursery pigs.Methods Forty pigs(6.5±0.4 kg)were assigned to 5 dietary treatments and fed for 35 d in 3 phases(11,9,and 15 d,respectively).Basal diets mainly included corn,soybean meal,and corn distiller's dried grains with solubles,contained phytase(750 FTU/kg),and were supplemented with 5 levels of XG at(1)0,(2)280 TXU/kg xylanase and 125 TGU/kgβ-glucanase,(3)560 and 250,(4)840 and 375,or(5)1,120 and 500,respectively.Growth performance was measured.On d 35,all pigs were euthanized and jejunal mucosa,jejunal digesta,jejunal tissues,and ileal digesta were collected to determine the effects of increasing XG levels and XG intake on intestinal health.Results Increasing XG intake tended to quadratically decrease(P=0.059)viscosity of jejunal digesta(min:1.74 m Pa·s at 751/335(TXU/TGU)/kg).Increasing levels of XG quadratically decreased(P<0.05)Prevotellaceae(min:0.6%at 630/281(TXU/TGU)/kg)in the jejunal mucosa.Increasing XG intake quadratically increased(P<0.05)Lactobacillaceae(max:40.3%at 608/271(TXU/TGU)/kg)in the jejunal mucosa.Increasing XG intake quadratically decreased(P<0.05)Helicobacteraceae(min:1.6%at 560/250(TXU/TGU)/kg)in the jejunal mucosa.Increasing levels of XG tended to linearly decrease(P=0.073)jejunal Ig G and tended to quadratically increase(P=0.085)jejunal villus height to crypt depth ratio(max:2.62 at 560/250(TXU/TGU)/kg).Increasing XG intake tended to linearly increase the apparent ileal digestibility of dry matter(P=0.087)and ether extract(P=0.065).Increasing XG intake linearly increased(P<0.05)average daily gain.Conclusions A combinational use of xylanase andβ-glucanase would hydrolyze the non-starch polysaccharides fractions,positively modulating the jejunal mucosa-associated microbiota.Increased intake of these enzyme combination possibly reduced digesta viscosity and humoral immune response in the jejunum resulting in improved intestinal structure,and ileal digestibility of nutrients,and finally improving growth of nursery pigs.The beneficial effects were maximized at a combination of 550 to 800 TXU/kg xylanase and 250 to 360 TGU/kgβ-glucanase.

Genome-wide association study identifies 12 new genetic loci associated with growth traits in pigs

Growth traits are among the most important economic traits in pigs and are regulated by polygenes with complex regulatory mechanisms.As the major indicators of growth performance,the backfat thickness(BFT),loin eye area(LEA),and days to 100 kg(D100)traits are commonly used to the genetics improvement in pigs.However,the available genetic markers for these traits are limited.To uncover novel loci and candidate genes associated with growth performance,we collected the phenotypic information of BFT,LEA,and D100 in 1,186 pigs and genotyped all these individuals using the Neogen GGP porcine 80K BeadChip.We performed a genome-wide association study(GWAS)using 4 statistical models,including mixed linear models(MLM),fixed and random model circulating probability unification(FarmCPU),settlement of MLM under progressively exclusive relationships(SUPER),Bayesian-information and linkage-disequilibrium Iteratively nested keyway(Blink),and identified 5,3,and 6 high-confidence single nucleotide polymorphisms(SNPs)associated with BFT,LEA,and D100,respectively.Variant annotation and quantitative trait locus(QTL)mapping analysis suggested that6 genes(SKAP2,SATB1,PDE7B,PPP1R16B,WNT3,and WNT9B)were potentially associated with growth performance in pigs.Transcriptome analysis suggested that the expression of Src Kinase Associated Phosphoprotein 2(SKAP2)was higher in prenatal muscles than in postnatal muscles,and the expression of Phosphodiesterase 7B(PDE7B)continuously increased during the prenatal stages and gradually decreased after birth,implying their potential roles in prenatal skeletal muscle development.Overall,this study provides new candidate loci and genes for the genetic improvement of pigs.

Amino acid and mineral digestibility,bone ash,and plasma inositol is increased by including microbial phytase in diets for growing pigs

Background The effect of microbial phytase on amino acid and energy digestibility is not consistent in pigs,which may be related to the phytase dosage or the adaptation length to the diet.Therefore,an experiment was conducted to test the hypotheses that increasing dietary phytase after an 18-day adaptation period:1)increases nutrient and energy digestibility;2)increases plasma P,plasma inositol,and bone ash of young pigs;and 3)demonstrates that maximum phytate degradation requires more phytase than maximum P digestibility.Results Data indicated that increasing inclusion of phytase[0,250,500,1,000,2,000,and 4,000 phytase units(FTU)/kg feed]in corn-soybean meal-based diets increased apparent ileal digestibility(AID)of Trp(quadratic;P<0.05),and of Lys and Thr(linear;P<0.05),and tended to increase AID of Met(linear;P<0.10).Increasing dietary phytase also increased AID and apparent total tract digestibility(ATTD)of Ca and P(quadratic;P<0.05)and increased ATTD of K and Na(linear;P<0.05),but phytase did not influence the ATTD of Mg or gross energy.Concentrations of plasma P and bone ash increased(quadratic;P<0.05),and plasma inositol also increased(linear;P<0.05)with increasing inclusion of phytase.Reduced concentrations of inositol phosphate(IP)6 and IP5(quadratic;P<0.05),reduced IP4 and IP3(linear;P<0.05),but increased inositol concentrations(linear;P<0.05)were observed in ileal digesta as dietary phytase increased.The ATTD of P was maximized if at least 1,200 FTU/kg were used,whereas more than 4,000 FTU/kg were needed to maximize inositol release.Conclusions Increasing dietary levels of phytase after an 18-day adaptation period increased phytate and IP ester degradation and inositol release in the small intestine.Consequently,increasing dietary phytase resulted in improved digestibility of Ca,P,K,Na,and the first 4 limiting amino acids,and in increased concentrations of bone ash and plasma P and inositol.In a corn-soybean meal diet,maximum inositol release requires approximately 3,200 FTU/kg more phytase than that required for maximum P digestibility.

Lysine 2-hydroxyisobutyrylation levels determined adipogenesis and fat accumulation in adipose tissue in pigs

Background Excessive backfat deposition lowering carcass grade is a major concern in the pig industry,especially in most breeds of obese type pigs.The mechanisms involved in adipogenesis and fat accumulation in pigs remain unclear.Lysine 2-hydroxyisobutyrylation(Khib),is a novel protein post-translational modification(PTM),which play an important role in transcription,energy metabolism and metastasis of cancer cells,but its role in adipogenesis and fat accumulation has not been shown.Results In this study,we first analyzed the modification levels of acetylation(Kac),Khib,crotonylation(Kcr)and succinylation(Ksu)of fibro-adipogenic progenitors(FAPs),myogenic precursors(Myo)and mesenchymal stem cells(MSCs)with varied differentiation potential,and found that only Khib modification in FAPs was significantly higher than that in MSCs.Consistently,in parallel with its regulatory enzymes lysine acetyltransferase 5(KAT5)and histone deacetylase 2(HDAC2)protein levels,the Khib levels increased quadratically(P<0.01)during adipogenic differentiation of FAPs.KAT5 knockdown in FAPs inhibited adipogenic differentiation,while HDAC2 knockdown enhanced adipogenic differentiation.We also demonstrated that Khib modification favored to adipogenic differentiation and fat accumulation by comparing Khib levels in FAPs and backfat tissues both derived from obese-type pigs(Laiwu pigs)and lean-type pigs(Duroc pigs),respectively.Accordingly,the expression patterns of KAT5 and HDAC2 matched well to the degree of backfat accumulation in obese-and lean-type pigs.Conclusions From the perspective of protein translational modification,we are the first to reveal the role of Khib in adipogenesis and fat deposition in pigs,and provided new clues for the improvement of fat accumulation and distribution as expected via genetic selection and nutritional strategy in obese-type pigs.

Genetically modified pigs with CD163 point mutation are resistant to HP-PRRSV infection

Porcine reproductive and respiratory syndrome(PRRS)is a globally prevalent contagious disease caused by the positive-strand RNA PRRS virus(PRRSV),resulting in substantial economic losses in the swine industry.Modifying the CD163 SRCR5 domain,either through deletion or substitution,can eff1ectively confer resistance to PRRSV infection in pigs.However,large fragment modifications in pigs inevitably raise concerns about potential adverse effects on growth performance.Reducing the impact of genetic modifications on normal physiological functions is a promising direction for developing PRRSV-resistant pigs.In the current study,we identified a specific functional amino acid in CD163 that influences PRRSV proliferation.Viral infection experiments conducted on Marc145 and PK-15CD163 cells illustrated that the mE535G or corresponding pE529G mutations markedly inhibited highly pathogenic PRRSV(HP-PRRSV)proliferation by preventing viral binding and entry.Furthermore,individual viral challenge tests revealed that pigs with the E529G mutation had viral loads two orders of magnitude lower than wild-type(WT)pigs,confirming effective resistance to HP-PRRSV.Examination of the physiological indicators and scavenger function of CD163 verified no significant differences between the WT and E529G pigs.These findings suggest that E529G pigs can be used for breeding PRRSV-resistant pigs,providing novel insights into controlling future PRRSV outbreaks.

Evaluating the performance of genomic selection on purebred population by incorporating crossbred data in pigs

Genomic selection(GS)has been widely used in livestock,which greatly accelerated the genetic progress of complex traits.The population size was one of the significant factors affecting the prediction accuracy,while it was limited by the purebred population.Compared to directly combining two uncorrelated purebred populations to extend the reference population size,it might be more meaningful to incorporate the correlated crossbreds into reference population for genomic prediction.In this study,we simulated purebred offspring(PAS and PBS)and crossbred offspring(CAB)base on real genotype data of two base purebred populations(PA and PB),to evaluate the performance of genomic selection on purebred while incorporating crossbred information.The results showed that selecting key crossbred individuals via maximizing the expected genetic relationship(REL)was better than the other methods(individuals closet or farthest to the purebred population,CP/FP)in term of the prediction accuracy.Furthermore,the prediction accuracy of reference populations combining PA and CAB was significantly better only based on PA,which was similar to combine PA and PAS.Moreover,the rank correlation between the multiple of the increased relationship(MIR)and reliability improvement was 0.60-0.70.But for individuals with low correlation(Cor(Pi,PA or B),the reliability improvement was significantly lower than other individuals.Our findings suggested that incorporating crossbred into purebred population could improve the performance of genetic prediction compared with using the purebred population only.The genetic relationship between purebred and crossbred population is a key factor determining the increased reliability while incorporating crossbred population in the genomic prediction on pure bred individuals.

Core and variable antimicrobial resistance genes in the gut microbiomes of Chinese and European pigs

Monitoring the prevalence of antimicrobial resistance genes(ARGs)is vital for addressing the global crisis of antibiotic-resistant bacterial infections.Despite its importance,the characterization of ARGs and microbiome structures,as well as the identification of indicators for routine ARG monitoring in pig farms,are still lacking,particularly concerning variations in antimicrobial exposure in different countries or regions.Here,metagenomics and random forest machine learning were used to elucidate the ARG profiles,microbiome structures,and ARG contamination indicators in pig manure under different antimicrobial pressures between China and Europe.Results showed that Chinese pigs exposed to high-level antimicrobials exhibited higher total and plasmid-mediated ARG abundances compared to those in European pigs(P<0.05).ANT(6)-Ib,APH(3')-IIIa,and tet(40)were identified as shared core ARGs between the two pig populations.Furthermore,the core ARGs identified in pig populations were correlated with those found in human populations within the same geographical regions.Lactobacillus and Prevotella were identified as the dominant genera in the core microbiomes of Chinese and European pigs,respectively.Forty ARG markers and 43 biomarkers were able to differentiate between the Chinese and European pig manure samples with accuracies of 100%and 98.7%,respectively.Indicators for assessing ARG contamination in Chinese and European pigs also achieved high accuracy(r=0.72-0.88).Escherichia flexneri in both Chinese and European pig populations carried between 21 and 37 ARGs.The results of this study emphasize the importance of global collaboration in reducing antimicrobial resistance risk and provide validated indicators for evaluating the risk of ARG contamination in pig farms.

Supplementation of vitamin E or a botanical extract as antioxidants to improve growth performance and health of growing pigs housed under thermoneutral or heat-stressed conditions

Background Heat stress has severe negative consequences on performance and health of pigs,leading to significant economic losses.The objective of this study was to investigate the effects of supplemental vitamin E and a botanical extract in feed or drinking water on growth performance,intestinal health,and oxidative and immune status in grow-ing pigs housed under heat stress conditions.Methods Duplicate experiments were conducted,each using 64 crossbred pigs with an initial body weight of 50.7±3.8 and 43.9±3.6 kg and age of 13-week and 12-week,respectively.Pigs(n=128)were housed individually and assigned within weight blocks and sex to a 2×4 factorial arrangement consisting of 2 environments(thermo-neutral(21.2℃)or heat-stressed(30.9℃))and 4 supplementation treatments(control diet;control+100 IU/L of D-α-tocopherol in water;control+200 IU/kg of DL-α-tocopheryl-acetate in feed;or control+400 mg/kg of a botanical extract in feed).Results Heat stress for 28 d reduced(P≤0.001)final body weight,average daily gain,and average daily feed intake(-7.4 kg,-26.7%,and-25.4%,respectively)but no effects of supplementation were detected(P>0.05).Serum vitamin E increased(P<0.001)with vitamin E supplementation in water and in feed(1.64 vs.3.59 and 1.64 vs.3.24),but not for the botanical extract(1.64 vs.1.67 mg/kg)and was greater when supplemented in water vs.feed(P=0.002).Liver vitamin E increased(P<0.001)with vitamin E supplementations in water(3.9 vs.31.8)and feed(3.9 vs.18.0),but not with the botanical extract(3.9 vs.4.9 mg/kg).Serum malondialdehyde was reduced with heat stress on d 2,but increased on d 28(interaction,P<0.001),and was greater(P<0.05)for antioxidant supplementation compared to control.Cellular proliferation was reduced(P=0.037)in the jejunum under heat stress,but increased in the ileum when vitamin E was supplemented in feed and water under heat stress(interaction,P=0.04).Tumor necrosis factor-αin jejunum and ileum mucosa decreased by heat stress(P<0.05)and was reduced by vitamin E sup-plementations under heat stress(interaction,P<0.001).Conclusions The addition of the antioxidants in feed or in drinking water did not alleviate the negative impact of heat stress on feed intake and growth rate of growing pigs.

Whole-genome analysis reveals distinct adaptation signatures to diverse environments in Chinese domestic pigs

Background Long-term natural and artificial selection has resulted in many genetic footprints within the genomes of pig breeds across distinct agroecological zones.Nevertheless,the mechanisms by which these signatures contribute to phenotypic diversity and facilitate environmental adaptation remain unclear.Results Here,we leveraged whole-genome sequencing data from 82 individuals from 6 domestic pig breeds originating in tropical,high-altitude,and frigid regions.Population genetic analysis suggested that habitat isolation significantly shaped the genetic diversity and contributed to population stratification in local Chinese pig breeds.Analysis of selection signals revealed regions under selection for adaptation in tropical(55.5 Mb),high-altitude(43.6 Mb),and frigid(17.72 Mb)regions.The potential functions of the selective sweep regions were linked to certain complex traits that might play critical roles in different geographic environments,including fat coverage in frigid environments and blood indicators in tropical and high-altitude environments.Candidate genes under selection were significantly enriched in biological pathways involved in environmental adaptation.These pathways included blood circulation,protein degradation,and inflammation for adaptation to tropical environments;heart and lung development,hypoxia response,and DNA damage repair for high-altitude adaptation;and thermogenesis,cold-induced vasodilation(CIVD),and the cell cycle for adaptation to frigid environments.By examining the chromatin state of the selection signatures,we identified the lung and ileum as two candidate functional tissues for environmental adaptation.Finally,we identified a mutation(chr1:G246,175,129A)in the cis-regulatory region of ABCA1 as a plausible promising variant for adaptation to tropical environments.Conclusions In this study,we conducted a genome-wide exploration of the genetic mechanisms underlying the adaptability of local Chinese pig breeds to tropical,high-altitude,and frigid environments.Our findings shed light on the prominent role of cis-regulatory elements in environmental adaptation in pigs and may serve as a valuable biological model of human plateau-related disorders and cardiovascular diseases.

Impact of an oligosaccharide-based polymer on the metabolic profiles and microbial ecology of weanling pigs experimentally infected with a pathogenic E.coli

Background Our previous study has reported that supplementation of oligosaccharide-based polymer enhances gut health and disease resistance of pigs infected with enterotoxigenic E.coli(ETEC)F18 in a manner similar to carbadox.The objective of this study was to investigate the impacts of oligosaccharide-based polymer or antibiotic on the host metabolic profiles and colon microbiota of weaned pigs experimentally infected with ETEC F18.Results Multivariate analysis highlighted the differences in the metabolic profiles of serum and colon digesta which were predominantly found between pigs supplemented with oligosaccharide-based polymer and antibiotic.The relative abundance of metabolic markers of immune responses and nutrient metabolisms,such as amino acids and carbohydrates,were significantly differentiated between the oligosaccharide-based polymer and antibiotic groups(q<0.2 and fold change>2.0).In addition,pigs in antibiotic had a reduced(P<0.05)relative abundance of Lachnospiraceae and Lactobacillaceae,whereas had greater(P<0.05)Clostridiaceae and Streptococcaceae in the colon digesta on d 11 post-inoculation(PI)compared with d 5 PI.Conclusions The impact of oligosaccharide-based polymer on the metabolic and microbial profiles of pigs is not fully understood,and further exploration is needed.However,current research suggest that various mechanisms are involved in the enhanced disease resistance and performance in ETEC-challenged pigs by supplementing this polymer.

Elimination of GGTA1,CMAH,β4GalNT2 and CIITA genes in pigs compromises human versus pig xenogeneic immune reactions

Background:Pig organ xenotransplantation is a potential solution for the severe organ shortage in clinic,while immunogenic genes need to be eliminated to improve the immune compatibility between humans and pigs.Current knockout strategies are mainly aimed at the genes causing hyperacute immune rejection(HAR)that occurs in the first few hours while adaptive immune reactions orchestrated by CD4 T cell thereafter also cause graft failure,in which process the MHCⅡmolecule plays critical roles.Methods:Thus,we generate a 4-gene(GGTA1,CMAH,β4GalNT2,and CIITA)knockout pig by CRISPR/Cas9 and somatic cell nuclear transfer to compromise HAR and CD4 T cell reactions simultaneously.Results:We successfully obtained 4KO piglets with deficiency in all alleles of genes,and at cellular and tissue levels.Additionally,the safety of our animals after gene editing was verified by using whole-genome sequencing and karyotyping.Piglets have survived for more than one year in the barrier,and also survived for more than 3 months in the conventional environment,suggesting that the piglets without MHCⅡcan be raised in the barrier and then gradually mated in the conventional environment.Conclusions:4KO piglets have lower immunogenicity,are safe in genomic level,and are easier to breed than the model with both MHCⅠandⅡdeletion.

Low crude protein formulation with supplemental amino acids for its impacts on intestinal health and growth performance of growing-finishing pigs

Background Low crude protein(CP)formulations with supplemental amino acids(AA)are used to enhance intestinal health,reduce costs,minimize environmental impact,and maintain growth performance of pigs.However,extensive reduction of dietary CP can compromise growth performance due to limited synthesis of non-essential AA and limited availability of bioactive compounds from protein supplements even when AA requirements are met.Moreover,implementing a low CP formulation can increase the net energy(NE)content in feeds causing excessive fat deposition.Additional supplementation of functional AA,coupled with low CP formulation could further enhance intestinal health and glucose metabolism,improving nitrogen utilization,and growth performance.Three experiments were conducted to evaluate the effects of low CP formulations with supplemental AA on the intestinal health and growth performance of growing-finishing pigs.Methods In Exp.1,90 pigs(19.7±1.1 kg,45 barrows and 45 gilts)were assigned to 3 treatments:CON(18.0%CP,supplementing Lys,Met,and Thr),LCP(16.0%CP,supplementing Lys,Met,Thr,Trp,and Val),and LCPT(16.1%CP,LCP+0.05%SID Trp).In Exp.2,72 pigs(34.2±4.2 kg BW)were assigned to 3 treatments:CON(17.7%CP,meeting the requirements of Lys,Met,Thr,and Trp);LCP(15.0%CP,meeting Lys,Thr,Trp,Met,Val,Ile,and Phe);and VLCP(12.8%CP,meeting Lys,Thr,Trp,Met,Val,Ile,Phe,His,and Leu).In Exp.3,72 pigs(54.1±5.9 kg BW)were assigned to 3 treatments and fed experimental diets for 3 phases(grower 2,finishing 1,and finishing 2).Treatments were CON(18.0%,13.8%,12.7%CP for 3 phases;meeting Lys,Met,Thr,and Trp);LCP(13.5%,11.4%,10.4%CP for 3 phases;meeting Lys,Thr,Trp,Met,Val,Ile,and Phe);and LCPG(14.1%,12.8%,11.1%CP for 3 phases;LCP+Glu to match SID Glu with CON).All diets had 2.6 Mcal/kg NE.Results In Exp.1,overall,the growth performance did not differ among treatments.The LCPT increased(P<0.05)Claudin-1 expression in the duodenum and jejunum.The LCP and LCPT increased(P<0.05)CAT-1,4F2hc,and B0AT expressions in the jejunum.In Exp.2,overall,the VLCP reduced(P<0.05)G:F and BUN.The LCP and VLCP increased(P<0.05)the backfat thickness(BFT).In Exp.3,overall,growth performance and BFT did not differ among treatments.The LCPG reduced(P<0.05)BUN,whereas increased the insulin in plasma.The LCP and LCPG reduced(P<0.05)the abundance of Streptococcaceae,whereas the LCP reduced(P<0.05)Erysipelotrichaceae,and the alpha diversity.Conclusions When implementing low CP formulation,CP can be reduced by supplementation of Lys,Thr,Met,Trp,Val,and Ile without affecting the growth performance of growing-finishing pigs when NE is adjusted to avoid increased fat deposition.Supplementation of Trp above the requirement or supplementation of Glu in low CP formulation seems to benefit intestinal health as well as improved nitrogen utilization and glucose metabolism.

Oils with different degree of saturation:effects on ileal digestibility of fat and corresponding additivity and bacterial community in growing pigs

Background Oils are important sources of energy in pig diets.The combination of oils with different degree of saturation contributes to improve the utilization efficiency of the mixed oils and may reduce the cost of oil supplemented.An experiment was conducted to evaluate the effects of oils with different degree of saturation on the fat digestibility and corresponding additivity and bacterial community in growing pigs.Methods Eighteen crossbred(Duroc×Landrace×Yorkshire)barrows(initial body weight:29.3±2.8 kg)were surgically fitted with a T-cannula in the distal ileum.The experimental diets included a fat-free basal diet and 5 oil-added diets.The 5 oil-added diets were formulated by adding 6%oil with different ratio of unsaturated to saturated fatty acids(U:S)to the basal diet.The 5 oils were palm oil(U:S=1.2),canola oil(U:S=12.0),and palm oil and canola oil were mixed in different proportions to prepare a combination of U:S of 2.5,3.5 and 4.5,respectively.Results The apparent and standardized ileal digestibility(AID and SID)of fat and fatty acids increased linearly(P<0.05)as the U:S of dietary oils increased except for SID of fat and C18:2.The AID and SID of fat and fatty acids differed among the dietary treatments(P<0.05)except for SID of unsaturated fatty acids(UFA)and C18:2.Fitted one-slope broken-line analyses for the SID of fat,saturated fatty acids(SFA)and UFA indicated that the breakpoint for U:S of oil was 4.14(R^(2)=0.89,P<0.01),2.91(R^(2)=0.98,P<0.01)and 3.84(R^(2)=0.85,P<0.01),respectively.The determined SID of fat,C18:1,C18:2 and UFA in the mixtures was not different from the calculated SID of fat,C18:1,C18:2 and UFA.However,the determined SID of C16:0,C18:0 and SFA in the mixtures were greater than the calculated SID values(P<0.05).The abundance of Romboutsia and Turicibacter in pigs fed diet containing palm oil was greater than that in rapeseed oil treatment group,and the two bacteria were negatively correlated with SID of C16:0,C18:0 and SFA(P<0.05).Conclusions The optimal U:S for improving the utilization efficiency of mixed oil was 4.14.The SID of fat and UFA for palm oil and canola oil were additive in growing pigs,whereas the SID of SFA in the mixture of two oils was greater than the sum of the values of pure oils.Differences in fat digestibility caused by oils differing in degree of saturation has a significant impact on bacterial community in the foregut.

基于PigsTrack跟踪器的群养生猪多目标跟踪

基于视频的生猪行为跟踪和识别对于实现精细化养殖具有重要价值。为了应对群养生猪多目标跟踪任务中由猪只外观相似、遮挡交互等因素带来的挑战,研究提出了基于PigsTrack跟踪器的群养生猪多目标跟踪方法。PigsTrack跟踪器利用高性能YOLOX网络降低目标误检与漏检率,采用Transformer模型获取具有良好区分特性的目标外观特征;基于OC-SORT(observation-centric sort)的思想,通过集成特征匹配、IoU匹配和遮挡恢复匹配策略实现群养生猪的准确跟踪。基于PBVD(pigs behaviours video dataset)数据集的试验结果表明,PigsTrack跟踪器的HOTA(higher order tracking accuracy),MOTA(multiple object tracking accuracy)和IDF1得分(identification F1 score)分别为85.66%、98.59%和99.57%,相较于现有算法的最高精度,分别提高了3.71、0.03和2.05个百分点,证明了PigsTrack跟踪器在解决外观相似和遮挡交互引起的跟踪过程中身份跳变问题方面的有效性。随后,利用Slowfast网络对PigsTrack跟踪器的跟踪结果进行了典型行为统计,结果显示PigsTrack在群养生猪个体行为统计方面更准确。此外,通过在ABVD(aggressive-behavior video)数据集上的试验,PigsTrack跟踪器的HOTA、MOTA和IDF1得分分别为69.14%、94.82%和90.11%,相对于现有算法的最高精度,提高了5.33、0.57和8.60个百分点,验证了PigsTrack跟踪器在群养生猪跟踪任务中的有效性。总而言之,PigsTrack跟踪器能够有效应对外观相似和遮挡交互等挑战,实现了准确的生猪多目标跟踪,并在行为统计方面展现出更高的准确性,为生猪养殖领域的研究和实际应用提供了有价值的指导。

Comparative effects of soy protein concentrate,enzyme‑treated soybean meal,and fermented soybean meal replacing animal protein supplements in feeds on growth performance and intestinal health of nursery pigs

Background Soy protein supplements,with high crude protein and less antinutritional factors,are produced from soybean meal by different processes.This study evaluated the comparative effects of various soy protein supplements replacing animal protein supplements in feeds on the intestinal immune status,intestinal oxidative stress,mucosaassociated microbiota,and growth performance of nursery pigs.Methods Sixty nursery pigs(6.6±0.5 kg BW)were allotted to five treatments in a randomized complete block design with initial BW and sex as blocks.Pigs were fed for 39 d in 3 phases(P1,P2,and P3).Treatments were:Control(CON),basal diet with fish meal 4%,2%,and 1%,poultry meal 10%,8%,and 4%,and blood plasma 4%,2%,and 1%for P1,P2,and P3,respectively;basal diet with soy protein concentrate(SPC),enzyme-treated soybean meal(ESB),fermented soybean meal with Lactobacillus(FSBL),and fermented soybean meal with Bacillus(FSBB),replacing 1/3,2/3,and 3/3 of animal protein supplements for P1,P2,and P3,respectively.Data were analyzed using the MIXED procedure in SAS 9.4.Results The SPC did not affect the BW,ADG,and G:F,whereas it tended to reduce(P=0.094)the ADFI and tended to increase(P=0.091)crypt cell proliferation.The ESM did not affect BW,ADG,ADFI,and G:F,whereas tended to decrease(P=0.098)protein carbonyl in jejunal mucosa.The FSBL decreased(P<0.05)BW and ADG,increased(P<0.05)TNF-α,and Klebsiella and tended to increase MDA(P=0.065)and IgG(P=0.089)in jejunal mucosa.The FSBB tended to increase(P=0.073)TNF-α,increased(P<0.05)Clostridium and decreased(P<0.05)Achromobacter and alpha diversity of microbiota in jejunal mucosa.Conclusions Soy protein concentrate,enzyme-treated soybean meal,and fermented soybean meal with Bacillus could reduce the use of animal protein supplements up to 33%until 7 kg body weight,up to 67%from 7 to 11 kg body weight,and entirely from 11 kg body weight without affecting the intestinal health and the growth performance of nursery pigs.Fermented soybean meal with Lactobacillus,however,increased the immune reaction and oxidative stress in the intestine consequently reducing the growth performance.