Parity changed fecal microbiota of sows and its correlation with milk long-chain fatty acid profiles.

The goal of this study was to characterize the fecal microbiota profiles of gestating sows, along with the fecal microbiota and milk fatty acid contents of lactating sows and their correlations with reproductive performance at different parities. The results showed that the microbiota of third parity gestating sows contained a greater abundance of Prevotella compared to the other two parity groups, while lactating sows exhibiting higher reproductive performance at fifth parity exhibited a greater abundance of Lactobacillus species. The lactating sows with higher reproductive performance also exhibited higher total monounsaturated fatty acid (MUFA) and higher total polyunsaturated fatty acid (PUFA) levels relative to sows with lower reproductive performance at all three analyzed parities, especially sows at fifth parity produced the lowest total saturated fatty acid (SFA) levels, and showed the highest C18:1n9c and C18:2n6c concentrations. In correlational analyses, the abundance of Oligella, Lactobacillus, and Corynebacterium was highly positively correlated with C18:1n9c, C18:2n6c, and C20:4n6. Overall, these results provide a rational basis for efforts to improve sow reproductive performance through the provision of precisely regulated nutrition. KEY POINTS: • Clear differences in the fecal microbiota were evident between sows of different parities. • Lactating sows with high reproductive performance showed distinct milk fatty acid profiles.

Sex-dependent changes in the microbiota profile, serum metabolism, and hormone levels of growing pigs after dietary supplementation with Lactobacillus.

Dietary supplementation with Lactobacillus can improve the intestinal microbial balance and exerts beneficial effects on pig health. However, whether these effects in pigs show differences between the sexes remains poorly understood. Therefore, we analyzed the sex-dependent patterns in the fecal microbiota after dietary supplementation with Lactobacillus acidophilus ZLA012 through high-throughput sequencing, determined the metabolomic profile of serum in barrows (immature castrated males) and gilts (sexually immature females) through nontargeted liquid chromatography-mass spectrometry, and measured the levels of various hormones, such as insulin, growth hormone, serotonin, triiodothyronine, and thyroxine. Lactobacillus supplementation resulted in more obvious effects on the microbial diversity and composition in barrows than in gilts. Specifically, supplementation with L. acidophilus ZLA012 significantly increased the abundance of the phylum Bacteroidetes and decreased that of Firmicutes in barrows. In contrast, more notable effects on metabolites, particularly those involved in lipid metabolism and glycerophospholipid metabolism, were observed in gilts than in barrows after Lactobacillus supplementation. Megasphaera, Dialister, Gemmiger, Faecalibacterium, Bulleidia, and Prevotella were the core functional genera associated with the significantly affected metabolites, which are involved in the biosynthesis, degradation, and elongation of fatty acids. L. acidophilus ZLA012 treatment increased the serum insulin, total bile acid, lipoprotein lipase, and nicotinamide adenine dinucleotide (NAD)+ levels in gilts, whereas higher serotonin levels were found in barrows than in gilts. The results indicated that the separate housing of barrows and gilts might be beneficial for targeted dietary supplementation and application of Lactobacillus in pig production. KEY POINTS: • L. acidophilus exerted obvious effects on microbiota profiles for barrows than gilts. • Gilts treated by L. acidophilus had a greater variety of lipid metabolism compared with barrows. • Lactobacillus regulated the dynamic balance among the microbiota, metabolism, and hormone in pigs.

Changes in the diversity and composition of gut microbiota of weaned piglets after oral administration of Lactobacillus or an antibiotic.

The gut microbiota plays important roles in the health and well-being of animals, and high-throughput sequencing facilitates exploration of microbial populations in the animal gut. However, previous studies have focused on fecal samples instead of the gastrointestinal tract. In this study, we compared the microbiota diversity and composition of intestinal contents of weaned piglets treated with Lactobacillus reuteri or chlortetracycline (aureomycin) using high-throughput sequencing. Nine weaned piglets were randomly divided into three groups and supplemented with L. reuteri, chlortetracycline, or saline for 10 days, and then the contents of three intestinal segments (jejunum, colon, and cecum) were obtained and used for sequencing of the V3-V4 hypervariable region of the 16S rRNA gene. The microbiota diversity and composition in the jejunum were different from those in the colon and cecum among the three treatments. In the jejunum, treatment with L. reuteri increased the species richness of the microbiota, as indicated by the ACE and Chao1 indexes, compared with the chlortetracycline group, in which several taxa were eliminated. In the colon and cecum, relative abundances of the phylum Firmicutes and the genus Prevotella were higher in the chlortetracycline group than in the other groups. Distances between clustered samples revealed that the L. reuteri group was closer to the chlortetracycline group than the control group for jejunum samples, while colon and cecum samples of the L. reuteri group were clustered with those of the control group. This study provides fundamental knowledge for future studies such as the development of alternatives to antibiotics.

Effects of Lactobacillus-fermented low-protein diets on the growth performance, nitrogen excretion, fecal microbiota and metabolomic profiles of finishing pigs.

This study investigated the effects of Lactobacillus-fermented low-protein diet on the growth performance, nitrogen balance, fecal microbiota, and metabolomic profiles of finishing pigs. A total of 90 finishing pigs were assigned to one of three dietary treatments including a normal protein diet (CON) as well as two experimental diets in which a low-protein diet supplemented with 0 (LP) or 1% Lactobacillus-fermented low-protein feed (FLP). In comparison with CON, the LP and FLP significantly increased average daily gain (P = 0.044), significantly decreased feed to gain ratio (P = 0.021), fecal nitrogen (P < 0.01), urine nitrogen (P < 0.01), and total nitrogen (P < 0.01), respectively. The LP group exhibited increased abundances of unclassified_f_Selenomonadaceae, Coprococcus, Faecalibacterium, and Butyricicoccus, while the abundances of Verrucomicrobiae, Verrucomicrobiales, Akkermansiaceae, and Akkermansia were enriched in the FLP group. Low-protein diet-induced metabolic changes were enriched in sesquiterpenoid and triterpenoid biosynthesis and Lactobacillus-fermented low-protein feed-induced metabolic changes were enriched in phenylpropanoid biosynthesis and arginine biosynthesis. Overall, low-protein diet and Lactobacillus-fermented low-protein diet improved the growth performance and reduce nitrogen excretion, possibly via altering the fecal microbiota and metabolites in the finishing pigs. The present study provides novel ideas regarding the application of the low-protein diet and Lactobacillus-fermented low-protein diet in swine production.

Maternal Malic Acid May Ameliorate Oxidative Stress and Inflammation in Sows through Modulating Gut Microbiota and Host Metabolic Profiles during Late Pregnancy.

Sows suffer oxidative stress and inflammation induced by metabolic burden during late pregnancy, which negatively regulates reproductive and lactating performances. We previously found that L-malic acid (MA) alleviated oxidative stress and inflammation and improved reproductive performances in sows. However, the mechanism underlying the MA's positive effects remains unexplored. Here, twenty Large White × Landrace sows with similar parity were randomly divided into two groups and fed with a basal diet or a diet supplemented with 2% L-malic acid complex from day 85 of gestation to delivery. The gut microbiome, fecal short-chain fatty acids, and untargeted serum metabolome were determined. Results showed that Firmicutes, Bacteroidota, and Spirochaetota were the top abundant phyla identified in late pregnancy for sows. Maternal MA supplementation modulated the composition but not the richness and diversity of gut microbiota during late pregnancy. Correlation analysis between gut microbiota and antioxidant capacity (or inflammation indicators) revealed that unclassified_f_Ruminococcaceae, unclassified_f_Lachnospiraceae, UCG-002, norank_f_norank_o_RF3, and Lactobacillus might play a role in anti-oxidation, and Lachnospiraceae_XPB1014_group, Lachnospiraceae_NK4A136_group, UCG-002, unclassified_f_Ruminococcaceae, Candidatus_Soleaferrea, norank_f_UCG-010, norank_f_norank_o_RF39, and unclassified_f_Lachnospiraceae might be involved in the anti-inflammatory effect. The improved antioxidant and inflammation status induced by MA might be independent of short chain fatty acid changes. In addition, untargeted metabolomics analysis exhibited different metabolic landscapes of sows in the MA group from in the control group and revealed the contribution of modified amino acid and lipid metabolism to the improved antioxidant capacity and inflammation status. Notably, correlation results of gut microbiota and serum metabolites, as well as serum metabolites and antioxidant capacity (or inflammation indicators), demonstrated that differential metabolism was highly related to the fecal microorganisms and antioxidant or inflammation indicators. Collectively, these data demonstrated that a maternal dietary supply of MA can ameliorate oxidative stress and inflammation in sows through modulating gut microbiota and host metabolic profiles during late pregnancy.

Wheat bran fermented by Lactobacillus regulated the bacteria-fungi composition and reduced fecal heavy metals concentrations in growing pigs.

Lactobacillus fermentation can increase the value of wheat bran, but the benefits of fermented wheat bran for pig production are poorly understood. We evaluated the phenolic acid content of wheat bran fermented with Lactobacillus. The bacterial and fungal compositions, short-chain fatty acids, and heavy metals concentrations in the feces of growing pigs were determined, and the correlations between the bacterial and fungal compositions and short-chain fatty acid and heavy metals concentrations were also assessed. The concentrations of phenolic acids (caffeic acid, catechinic acid, and gallic acid) were higher in fermented bran than in control wheat bran. The diversity of feces bacterial species was significantly higher, whereas the diversity of fungi was lower in fermented wheat bran treatment than those in the control group, and pigs consuming fermented and control wheat bran with different bacterial and fungal compositions had different growth rates. The abundance of genera in fungi that were less abundant in the fermented group samples than in the control samples (including Wallemia, Trichosporon, Candida, Aspergillus, and unclassified_f__Microascaceae) was positively correlated with heavy metals concentrations in pig feces, and the abundances of these fungi were negatively correlated with caffeic acid, catechinic acid, and gallic acid concentrations. Metagenomic function predictions indicated that larger amounts of secondary metabolites were synthesized in the fermented group than in the control group. The results provide new insights into the roles of bacterial-fungal interactions in the growth and decreasing environmental pollution of pigs consuming fermented wheat bran.

Lactobacillus-driven feed fermentation regulates microbiota metabolism and reduces odor emission from the feces of pigs.

IMPORTANCE: Our present study showed that dietary supplementation with feed fermented by Lactobacillus could promote the growth performance of pigs, regulate the microbiota, and inhibit the growth of harmful bacteria. It could prevent the accumulation of toxic substances and reduce odor emission from pig feces, thereby reducing environmental pollution. In addition, one key triumph of the present study was the isolation of Weissella cibaria ZWC030, and the strain could inhibit the production of skatole in vitro in our present results.

Chenodeoxycholic Acid Improves Embryo Implantation and Metabolic Health through Modulating Gut Microbiota-Host Metabolites Interaction during Early Pregnancy.

Fetus loss in early pregnancy is of major concern to both humans and animals, and this issue is largely influenced by embryo implantation. Chenodeoxycholic acid (CDCA), a primary bile acid, contributes to metabolic improvements and protects against intrahepatic cholestasis of pregnancy. However, the effect of CDCA on embryo implantation during early pregnancy has not been investigated. The present study demonstrated that CDCA administration during early pregnancy improved embryo implantation in sows and rats, thereby improving the pregnancy outcomes of sows. CDCA significantly reduced inflammation, oxidative stress, and insulin resistance. The metabolomics analysis indicated significant differences in the fecal metabolome, especially regarding the level of secondary bile acids, between the control and CDCA-treated sows. CDCA also influenced the serum metabolite profiles in sows, and the serum L-Histidine level was significantly correlated with the abundance of 19 differential fecal metabolites. Importantly, L-Histidine administration improved embryo implantation and metabolic health in rats during early pregnancy. Moreover, CDCA administration during early pregnancy also led to long-term metabolic improvements in sows. Our data indicated that CDCA improved embryo implantation by alleviating inflammation and oxidative stress, improving insulin sensitivity, and modulating the interaction between the gut microbiota and host metabolites. Therefore, CDCA intervention is a potential therapeutic strategy regarding embryo loss during pregnancy.

An Economic Model Predictive Control for Knowledge Transmission Processes in Multilayer Complex Networks.

In this article, we study the optimal feedback control problems of knowledge dissemination processes in multilayer complex networks. First, a node-based model is established in multilayer complex networks and two collaborative control strategies are exerted to increase the scope and speed of knowledge dissemination, forming a closed-loop control system. Then, we develop a two-layer optimal control framework. At the upper level, the optimal solution of the control system is solved and sent to the lower layer. At the lower level, a model predictive controller (MPC) receives input information from the upper level and is formulated to decide on the network and then transmits it to its heterogeneous networks which can reduce control resources and computation complexity. Finally, numerical simulations are conducted to confirm the theoretical results.

Identification of Phytogenic Compounds with Antioxidant Action That Protect Porcine Intestinal Epithelial Cells from Hydrogen Peroxide Induced Oxidative Damage.

Oxidative stress contributes to intestinal dysfunction. Plant extracts can have antioxidant action; however, the specific phytogenic active ingredients and their potential mechanisms are not well known. We screened 845 phytogenic compounds using a porcine epithelial cell (IPEC-J2) oxidative stress model to identify oxidative-stress-alleviating compounds. Calycosin and deoxyshikonin were evaluated for their ability to alleviate H2O2-induced oxidative stress by measuring their effects on malondialdehyde (MDA) accumulation, reactive oxygen species (ROS) generation, apoptosis, mitochondrial membrane potential (MMP), and antioxidant defense. Nrf2 pathway activation and the effect of Nrf2 knockdown on the antioxidative effects of hit compounds were investigated. Calycosin protected IPEC-J2 cells against H2O2-induced oxidative damage, likely by improving the cellular redox state and upregulating antioxidant defense via the Nrf2-Keap1 pathway. Deoxyshikonin alleviated the H2O2-induced decrease in cell viability, ROS production, and MMP reduction, but had no significant effect on MDA accumulation and apoptosis. Nrf2 knockdown did not weaken the effect of deoxyshikonin in improving cell viability, but it weakened its effect in suppressing ROS production. These results indicate that the mechanisms of action of natural compounds differ. The newly identified phytogenic compounds can be developed as novel antioxidant agents to alleviate intestinal oxidative stress in animals.

Lactobacillus plantarum Exhibits Antioxidant and Cytoprotective Activities in Porcine Intestinal Epithelial Cells Exposed to Hydrogen Peroxide.

Probiotics are widely used for protection against stress-induced intestinal dysfunction. Oxidative stress plays a critical role in gastrointestinal disorders. It is established that probiotics alleviate oxidative stress; however, the mechanism of action has not been elucidated. We developed an in vitro intestinal porcine epithelial cells (IPEC-J2) model of oxidative stress to explore the antioxidant effect and potential mode of action of Lactobacillus plantarum ZLP001. The IPEC-J2 cells were preincubated with and without L. plantarum ZLP001 for 3 h and then exposed to hydrogen peroxide (H2O2) for 4 h. Pretreatment with L. plantarum ZLP001 protected IPEC-J2 cells against H2O2-induced oxidative damage as indicated by cell viability assays and significantly alleviated apoptosis elicited by H2O2. L. plantarum ZLP001 pretreatment decreased reactive oxygen species production and the cellular malondialdehyde concentration and increased the mitochondrial membrane potential compared with H2O2 treatment alone, suggesting that L. plantarum ZLP001 promotes the maintenance of redox homeostasis in the cells. Furthermore, L. plantarum ZLP001 regulated the expression and generation of some antioxidant enzymes, thereby activating the antioxidant defense system. Treatment with L. plantarum ZLP001 led to nuclear erythroid 2-related factor 2 (Nrf2) enrichment in the nucleus compared with H2O2 treatment alone. Knockdown of Nrf2 significantly weakened the alleviating effect of L. plantarum ZLP001 on antioxidant stress in IPEC-J2 cells, suggesting that Nrf2 is involved in the antioxidative effect of L. plantarum ZLP001. Collectively, these results indicate that L. plantarum ZLP001 is a promising probiotic bacterium that can potentially alleviate oxidative stress.

Effects of dietary supplementation with Pediococcus acidilactici ZPA017 on reproductive performance, fecal microbial flora and serum indices in sows during late gestation and lactation.

OBJECTIVE: This study was conducted to determine the effects of dietary supplementation with Pediococcus acidilactici (P. acidilactici) ZPA017 as a probiotic on reproductive performance, fecal microbial flora and serum indices in sows during late gestation and lactation. METHODS: A total of 94 sows (Large White×Yorkshire, average 4.50 parities) were randomly allotted to two dietary treatments: control diet and the diet supplemented with P. acidilactici ZPA017 (2.40×109 colony-forming unit/kg of diets). The study started at d 90 of gestation and conducted until d 28 of lactation. RESULTS: Compared to sows fed the control diet, supplementation of P. acidilactici ZPA017 increased the number of weaning piglets, weaning weight of litter and piglets, survival rate of piglets at weaning (p<0.05), and decreased diarrhea rate of piglets in lactation (p<0.05). Dietary P. acidilactici ZPA017 increased fecal Lactobacillus populations (p = 0.030) and reduced fecal Escherichia coli and Staphylococcus aureus populations (p<0.05) of sows at weaning. Moreover, the supplementation of P. acidilactici ZPA017 increased serum concentrations of immunoglobulin G, immunoglobulin A and total protein (p<0.05), while decreased serum haptoglobin concentration and alanine aminotransferase activity (p<0.05) of sows at weaning. CONCLUSION: Administration of P. acidilactici ZPA017 in diets during late gestation and lactation had positive effects on the reproductive performance, intestinal microflora balance and immunity of sows.

Complete Genome Sequencing and Comparative Genome Characterization of Lactobacillus johnsonii ZLJ010, a Potential Probiotic With Health-Promoting Properties.

Lactobacillus johnsonii ZLJ010 is a probiotic strain isolated from the feces of a healthy sow and has putative health-promoting properties. To determine the molecular basis underlying the probiotic potential of ZLJ010 and the genes involved in the same, complete genome sequencing and comparative genome analysis with L. johnsonii ZLJ010 were performed. The ZLJ010 genome was found to contain a single circular chromosome of 1,999,879 bp with a guanine-cytosine (GC) content of 34.91% and encoded 18 ribosomal RNA (rRNA) genes and 77 transfer RNA (tRNA) genes. From among the 1,959 protein coding sequences (CDSs), genes known to confer probiotic properties were identified, including genes related to stress adaptation, biosynthesis, metabolism, transport of amino acid, secretion, and the defense machinery. ZLJ010 lacked complete or partial biosynthetic pathways for amino acids but was predicted to compensate for this with an enhanced transport system and some unique amino acid permeases and peptidases that allow it to acquire amino acids and other precursors exogenously. The comparative genomic analysis of L. johnsonii ZLP001 and seven other available L. johnsonii strains, including L. johnsonii NCC533, FI9785, DPC6026, N6.2, BS15, UMNLJ22, and PF01, revealed 2,732 pan-genome orthologous gene clusters and 1,324 core-genome orthologous gene clusters. Phylogenomic analysis based on 1,288 single copy genes showed that ZLJ010 had a closer relationship with the BS15 from yogurt and DPC6026 from the porcine intestinal tract but was located on a relatively standalone branch. The number of clusters of unique, strain-specific genes ranged from 42 to 185. A total of 219 unique genes present in the genome of L. johnsonii ZLJ010 primarily encoded proteins that are putatively involved in replication, recombination and repair, defense mechanisms, transcription, amino acid transport and metabolism, and carbohydrate transport and metabolism. Two unique prophages were predicted in the ZLJ010 genome. The present study helps us understand the ability of L. johnsonii ZLJ010 to better adapt to the gut environment and also its probiotic functionalities.

Swine-Derived Probiotic Lactobacillus plantarum Modulates Porcine Intestinal Endogenous Host Defense Peptide Synthesis Through TLR2/MAPK/AP-1 Signaling Pathway.

Host defense peptides (HDPs) have antimicrobial and immunoregulatory activities and are involved in epithelial innate immune defense. Dietary modulation of endogenous HDP synthesis is an effective way to boost the host innate immune system. This study aimed to investigate the role of the probiotic Lactobacillus plantarum strain ZLP001 in porcine HDP induction and the underlying mechanism. To this end, we evaluated the stimulatory effect of L. plantarum ZLP001 on HDP expression in piglet intestinal tissue in vivo and porcine IPEC-J2 cells and 3D4/31 cells in vitro, and we examined the underlying intracellular signaling pathway in IPEC-J2 cells. L. plantarum ZLP001 treatment increased the mRNA expression of jejunal and ileal HDPs in weaned piglets. In IPEC-J2 and 3D4/31 cells, L. plantarum ZLP001 stimulated HDP expression, but different HDP induction patterns were observed, with the various HDPs exhibiting different relative mRNA levels in each cell line. L. plantarum ZLP001 induced porcine HDP expression through toll-like receptor (TLR)2 recognition as evidenced by the fact that HDP expression was suppressed in TLR2-knockdown IPEC-J2 cells. Furthermore, we found that L. plantarum ZLP001 activated the extracellular signal-regulated kinase (ERK)1/2 and c-jun N-terminal kinase (JNK) signaling pathways, as indicated by enhanced phosphorylation of both ERK1/2 and JNK and the fact that HDP expression was suppressed upon inhibition of ERK1/2 and JNK. Furthermore, L. plantarum ZLP001 activated c-fos and c-jun transcription factor phosphorylation and activity. We conclude that L. plantarum ZLP001 induces porcine HDP expression in vivo and in vitro, and the induction seems to be regulated via TLR2 as well as the ERK1/2/JNK and c-jun/c-fos signaling pathways. Modulation of endogenous HDPs mediated by L. plantarum ZLP001 might be a promising approach to improving intestinal health and enhancing diarrhea resistance in weaning piglets.

Effects of oligosaccharides on the growth and stress tolerance of Lactobacillus plantarum ZLP001 in vitro, and the potential synbiotic effects of L. plantarum ZLP001 and fructo-oligosaccharide in post-weaning piglets1.

In this study, we evaluated the effects of seven oligosaccharides on the growth rate and stress tolerance of Lactobacillus plantarum ZLP001 in vitro, and the potential synbiotic effects of the most effective oligosaccharide [fructo-oligosaccharide (FOS)] and L. plantarum ZLP001 on the growth performance, apparent nutrient digestibility, fecal microbiota, and serum immune index in weaning piglets. Most oligosaccharides were utilized as carbohydrate sources by L. plantarum ZLP001, but we observed obvious differences in the bacterial growth depending on oligosaccharide type and concentration. Oligosaccharides and glucose significantly alleviated the decrease in L. plantarum ZLP001 viability in artificial gastric fluid, whereas none of the sugars affected viability in artificial intestinal fluid. FOS and galacto-oligosaccharide significantly improved the viability of L. plantarum ZLP001 under heat stress (65 °C for 15 and 30 min). FOS and soybean oligosaccharide significantly increased the viability of L. plantarum ZLP001 in response to cold stress (4 °C for 30 and 60 days). On the basis of the findings of in vitro experiments, we selected FOS for in vivo studies. Eighty-four weaned piglets were randomly assigned to one of the following groups: control (basal diet, no additives), freeze-dried L. plantarum ZLP001 (4.2 × 109 CFU/g, 2 g/kg diet), FOS (5 g/kg diet), and combination (0.2% L. plantarum ZLP001 + 0.5% FOS). Body weight and feed consumption were recorded for determinations of the average daily gain (ADG), average daily feed intake (ADFI), and feed-to-gain ratio (F/G). On day 28, fresh fecal samples were collected to evaluate the apparent digestibility of nutrients and microbiota, and serum samples were collected to determine the immune status. L. plantarum ZLP001 plus FOS significantly increased ADG and decreased the F/G ratio compared with the no-additive control. The combination treatment also increased the apparent nutrient digestibility of dry matter and crude protein. Compared with the control and single supplementation, the combination treatment had a significant regulatory effect on the intestinal microbiota, as evidenced by increases in Lactobacillus spp. and a decrease in Enterobacteriaceae. In addition, the combination treatment increased the concentrations of serum IFN-γ and immunoglobulin G. In conclusion, FOS can be utilized well by L. plantarum ZLP001 and can be combined with it as a potential synbiotic that shows synergistic effects in weaning piglets.

Fecal Microbiota and Its Correlation With Fatty Acids and Free Amino Acids Metabolism in Piglets After a Lactobacillus Strain Oral Administration.

Lactobacillus has a positive effect on the host intestinal microbiota. In piglets, dietary supplementation with Lactobacillus affects general health and plays an important role in nutrient digestion and fermentation. However, this association requires further investigation. Here, we studied newborn piglets from 12 litters. The nursed piglets were given a creep feed beginning on day 10 post-partum and weaned at day 30. Piglets were fed either a control basic diet or a diet including supplementation with Lactobacillus reuteri ZLR003 at 6.0 × 106 CFU/g feed. At day 30 and 60, feces samples were taken and used for sequencing of the V3-V4 hypervariable region of the 16S rRNA gene. At day 60, feces samples and serum samples were also taken and used to measure the short chain fatty acids (SCFAs) and to detect long chain fatty acids (LCFAs) and free amino acids (FAAs), respectively. The results revealed that L. reuteri ZLR003 could improve piglet fecal microbiota composition, especially at the end of weaned period. The concentrations of lactic acid and butyric acid in feces were higher, and acetic acid concentration was lower in the L. reuteri ZLR003 group compared with the control group (P < 0.05). The serum polyunsaturated fatty acids C18:2n6c, C18:3n3, C20:4n6, and C22:6n3 were significantly higher (P < 0.05), as were the serum FAAs Gly, Ala, Val, Iso, Asn, Asp, Glu, Met, Phe, and Leu (P < 0.05), in the L. reuteri group compared with the control group. A correlation analysis revealed that the genera Ruminococcaceae_UCG-010 and Ruminococcaceae_UCG-014 had a negative correlation with the SCFAs content in feces, the genus Prevotella_9 had a higher positive correlation with C18:2n6c, and the genera Megasphaera and Mitsuokella had a more positive significant effect on the serum FAAs content in weaned piglets in the L. reuteri ZLR003 group compared with the control group. In conclusion, L. reuteri ZLR003 influenced the fecal microbiota composition of piglets, and its effects were related to the metabolism of SCFAs, LCFAs, and FAAs. Our findings will help facilitate the application of Lactobacillus strains in pig production.

Swine-Derived Probiotic Lactobacillus plantarum Inhibits Growth and Adhesion of Enterotoxigenic Escherichia coli and Mediates Host Defense.

Weaning stress renders piglets susceptible to pathogen infection, which leads to post-weaning diarrhea, a severe condition characterized by heavy diarrhea and mortality in piglets. Enterotoxigenic Escherichia coli (ETEC) is one of typical strains associated with post-weaning diarrhea. Thus, prevention and inhibition of ETEC infection are of great concern. Probiotics possess anti-pathogenic activity and can counteract ETEC infection; however, their underlying mechanisms and modes of action have not yet been clarified. In the present study, the direct and indirect protective effects of Lactobacillus plantarum ZLP001 against ETEC infection were investigated by different methods. We found that bacterial culture and culture supernatant of L. plantarum ZLP001 prevented ETEC growth by the Oxford cup method, and ETEC growth inhibition was observed in a co-culture assay as well. This effect was suggested to be caused mainly by antimicrobial metabolites produced by L. plantarum ZLP001. In addition, adhesion capacity of L. plantarum ZLP001 to IPEC-J2 cells were observed using microscopy and counting. L. plantarum ZLP001 also exhibited a concentration-dependent ability to inhibit ETEC adhesion to IPEC-J2 cells, which mainly occurred via exclusion and competition mode. Furthermore, quantitative real time polymerase chain reaction (qPCR) analysis showed that L. plantarum ZLP001 upregulated the expression of host defense peptides (HDPs) but did not trigger an inflammatory response. In addition, L. plantarum ZLP001 induced HDP secretion, which enhanced the potential antimicrobial activity of IPEC-J2 cell-culture supernatant after incubation with L. plantarum ZLP001. Our findings demonstrate that L. plantarum ZLP001, an intestinal Lactobacillus species associated with piglet health, possesses anti-ETEC activity. L. plantarum ZLP001 might prevent ETEC growth, inhibit ETEC adhesion to the intestinal mucosa, and activate the innate immune response to secret antimicrobial peptides. L. plantarum ZLP001 is worth investigation as a potential probiotics.

Probiotic Lactobacillus plantarum Promotes Intestinal Barrier Function by Strengthening the Epithelium and Modulating Gut Microbiota.

Weaning disturbs the intestinal barrier function and increases the risk of infection in piglets. Probiotics exert beneficial health effects, mainly by reinforcing the intestinal epithelium and modulating the gut microbiota. However, the mechanisms of action, and especially, the specific regulatory effects of modulated microbiota by probiotics on the intestinal epithelium have not yet been elucidated. The present study aimed to decipher the protective effects of the probiotic Lactobacillus plantarum strain ZLP001 on the intestinal epithelium and microbiota as well as the effects of modulated microbiota on epithelial function. Paracellular permeability was measured with fluorescein isothiocyanate-dextran (FD-4). Gene and protein expression levels of tight junction (TJ) proteins, proinflammatory cytokines, and host defense peptides were determined by RT-qPCR, ELISA, and western blot analysis. Short-chain fatty acid (SCFA) concentrations were measured by ion chromatography. Fecal microbiota composition was assessed by high-throughput sequencing. The results showed that pretreatment with 108 colony forming units (CFU) mL-1 of L. plantarum ZLP001 significantly counteracted the increase in gut permeability to FD-4 induced by 106 CFU mL-1 enterotoxigenic Escherichia coli (ETEC). In addition, L. plantarum ZLP001 pretreatment alleviated the reduction in TJ proteins (claudin-1, occludin, and ZO-1) and downregulated proinflammatory cytokines IL-6 and IL-8, and TNFα expression and secretion caused by ETEC. L. plantarum ZLP001 also significantly increased the expression of the host defense peptides pBD2 and PG1-5 and pBD2 secretion relative to the control. Furthermore, L. plantarum ZLP001 treatment affected piglet fecal microbiota. The abundance of butyrate-producing bacteria Anaerotruncus and Faecalibacterium was significantly increased in L. plantarum ZLP001-treated piglets, and showed a positive correlation with fecal butyric and acetic acid concentrations. In addition, the cell density of Clostridium sensu stricto 1, which may cause epithelial inflammation, was decreased after L. plantarum ZLP001 administration, while the beneficial Lactobacillus was significantly increased. Our findings suggest that L. plantarum ZLP001 fortifies the intestinal barrier by strengthening epithelial defense functions and modulating gut microbiota.

Gene expression profile changes in the jejunum of weaned piglets after oral administration of Lactobacillus or an antibiotic.

The small intestine plays an essential role in the health and well-being of animals. Previous studies have shown that Lactobacillus has a protective effect on intestinal morphology, intestinal epithelium integrity and appropriate maturation of gut-associated tissues. Here, gene expression in jejunum tissue of weaned piglets was investigated by RNA-seq analysis after administration of sterile saline, Lactobacillus reuteri, or an antibiotic (chlortetracycline). In total, 401 and 293 genes were significantly regulated by chlortetracycline and L. reuteri, respectively, compared with control treatment. Notably, the HP, NOX1 and GPX2 genes were significantly up-regulated in the L. reuteri group compared with control, which is related to the antioxidant ability of this strain. In addition, the expression of genes related to arachidonic acid metabolism and linoleic acid metabolism enriched after treatment with L. reuteri. The fatty acid composition in the jejunum and colon was examined by GC-MS analysis and suggested that the MUFA C18:1n9c, and PUFAs C18:2n6c and C20:4n6 were increased in the L. reuteri group, verifying the GO enrichment and KEGG pathway analyses of the RNA-seq results. The results contribute to our understanding of the probiotic activity of this strain and its application in pig production.

Complete genome sequence of probiotic Lactobacillus reuteri ZLR003 isolated from healthy weaned pig.

Lactobacillus reuteri ZLR003 was isolated from the caecum mucosa of healthy weaned pigs with displaying probiotic properties in our laboratory. Here, we present the complete genome sequence of L. reuteri ZLR003, which consists of a circular 2,234,097bp chromosome (G+C content of 38.66%). Such information will provide insights into the molecular mechanism of its probiotic activity and facilitate its application in animal production.