Parabacteroides faecalis sp. nov. Isolated from Swine Faeces.

A Gram-negative, obligate anaerobic, non-motile, non-spore-forming, rod-shaped bacterial strain designated AGMB00274T was isolated from swine faeces. An 16S rRNA gene analysis indicated that strain AGMB00274T belonged to the genus Parabacteroides, with the highest similarity to Parabacteroides johnsonii (P. johnsonii) DSM 18315T (sequence similarity of 94.9%). The genome size of strain AGMB00274T was 4,308,683 bp, with a DNA G+C content of 42.5 mol%. The biochemical analysis of strain AGMB00274T showed that it was positive for gelatin hydrolysis and α-fucosidase, but negative for the acid production from D-glucose, D-mannitol, D-maltose, salicin, glycerol, D-cellobiose, D-mannose, D-melezitose, D-sorbitol, D-trehalose, and negative for α-arabinosidase, glutamic acid decarboxylase, and pyroglutamic acid arylamidase. The dominant cellular fatty acids (> 10%) of the isolate were anteiso-C15: 0 (23.2%), iso-C15: 0 (16.6%), C18: 1 ω9c (16.4%), summed feature 11 (iso-C17: 0 3-OH and/or C18: 2 DMA) (12.5%), and C16: 0 (11.3%). The major respiratory quinones of strain AGMB00274T were MK-9 (55.4%) and MK-10 (44.6%). The major polar lipid was phosphatidylethanolamine. Based on phylogenetic, genetic, physiological, and chemotaxonomic analyses, as a novel species of the genus Parabacteroides, strain AGMB00274T was proposed with the name Parabacteroides faecalis sp. nov. The type strain used was AGMB00274T (= KCTC 25286T = GDMCC 1.2742T).

Development and application of a multiplex PCR method to differentiate Salmonella enterica serovar Typhimurium from its monophasic variants in pig farms.

Salmonella enterica serovar Typhimurium monophasic variants (Salmonella 4,[5],12:i:-) has increased dramatically, causing human salmonellosis and colonization in pigs. With a difference to S. Typhimurium, the monophasic variants of S. Typhimurium lose the gene cassettes encoding the second phase flagellin. To establish a rapid method to detect and differentiate the two serotypes, we analyzed the published 679 genomes of S. Typhimurium and its monophasic variants and found that no Salmonella 4,[5],12:i:- strains carry both fljB and hin genes. Therefore, we established a novel multiplex PCR method using the fljB-hin region and mdh gene as target sequences to detect and differentiate both serotypes. This method can be used to specifically detect both serotypes with a detection limit for DNA concentration at 10 pg/µL. In addition, the PCR assay successfully differentiated 36 S. Typhimurium isolates from 62 isolates of monophasic variants preserved in our laboratory from 2009 to 2017, which corresponds to the whole-genome-based serotyping results. Application of the multiplex PCR method to 60 fecal samples from a pig farm identified 11.7% (7/60) of S. Typhimurium monophasic variants, which is consistent with the whole-genome-based serotyping results. The multiplex PCR assay is a rapid and precise method for the detection of S. Typhimurium monophasic variants from samples across food production chains.

Modulation of the porcine intestinal microbiota in the course of Ascaris suum infection.

BACKGROUND: The porcine roundworm Ascaris suum impairs feed conversion and weight gain, but its effects on intestinal microbiota remain largely unexplored. METHODS: Modulation of the intestinal microbiota was assessed in pigs that were infected once with 10,000 A. suum eggs and pigs that received a trickle infection (1000 eggs/day over 10 days), compared with a non-infected control group. Six pigs each were sacrificed per group at days 21, 35 and 49 post-infection (p.i.). Faecal samples taken weekly until slaughter and ingesta samples from different intestinal compartments were subjected to next-generation sequencing of the bacterial 16S rRNA gene. RESULTS: The results revealed marked differences between the single- and the trickle-infected group. Single infection caused a remarkable but transient decrease in microbial diversity in the caecum, which was not observed in the trickle-infected group. However, an increase in short-chain fatty acid-producing genera in the caecum on day 21 p.i., which shifted to a decrease on day 35 p.i., was common to both groups, possibly related to changes in excretory-secretory products following the parasite's final moult. Faecal microbial interaction networks were more similar between the single-infected and control group than the trickle-infected group. In addition, a lower degree of similarity over time indicated that A. suum trickle infection prevented microbiota stabilization. CONCLUSIONS: These different patterns may have important implications regarding the comparability of experimental infections with natural scenarios characterized by continuous exposure, and should be confirmed by further studies.

Genomic characterization of Salmonella enterica serovar Choleraesuis from Brazil reveals a swine gallbladder isolate harboring colistin resistance gene mcr-1.1.

Salmonella enterica serovar Choleraesuis (S. Choleraesuis) is a swine-adapted serovar associated to invasive infections in humans. In Brazil, data of strains of this serovar are scarce. In the present study, six S. Choleraesuis strains of animal (n = 5) and human (n = 1) origin from Brazil were screened for phenotypic antimicrobial resistance using disk-diffusion assay and using whole-genome sequencing data to search for antimicrobial resistance genes, plasmids, prophages, and Salmonella pathogenicity islands (SPIs). Its genetic relatedness was evaluated by MLST and SNP analysis. A single isolate from swine gallbladder harbored the colistin resistance gene mcr-1.1 into a IncX4 plasmid. In the six strains analyzed, resistance was found to tetracycline, nalidixic acid, ciprofloxacin, ampicillin, piperacillin, streptomycin, cefazoline, gentamycin, sulfamethoxazole-trimethoprim, and choloramphenicol, along with resistance genes aac(6')-Iaa, aac(3)-IV, aph(3'')-Ib, aph(6)-Id, aph(4)-Ia, aadA1, aph(3')-IIa, blaTEM-1A, floR, sul1, sul2, tet(B), drfA1, erm(B), mph(B), lnu(G), qacE, and gyrA point mutation Serine83 → Tyrosine and parC Threonine57 → Serine. Furthermore, IncF and IncH plasmids, ten SPIs, and seven prophage types were detected. All strains were assigned to ST145 and five belonged to a common SNP cluster of S. Choleraesuis strains from Brazil. The presence of S. Choleraesuis isolated from animals harboring relevant antimicrobial resistance profiles and virulence determinants reinforced the urge for enhanced surveillance to avoid its transmission to humans through food items.

Molecular characterization of extended-spectrum ß-lactamase producers, carbapenemase producers, polymyxin-resistant, and fosfomycin-resistant Enterobacterales among pigs from Egypt.

OBJECTIVES: To perform the first prospective surveillance evaluating the occurrence of genes encoding colistin resistance, fosfomycin resistance, carbapenemase, or extended-spectrum ß-lactamases (ESBLs) among Enterobacterial isolates recovered from the gut flora of pigs from Egypt. METHODS: Between February and April 2020, 81 rectal swabs were collected from pigs in a slaughterhouse, Cairo, Egypt. Samples were screened for different resistance mechanisms using SuperPolymyxin, ChromID ESBL, SuperFOS, and SuperCarba selective agar plates. Antimicrobial susceptibility testing was performed for all isolates using disk diffusion and broth microdilution techniques. PCR screening was performed for ESBLs, carbapenemases, mcr, and fosA genes. Mating-out assays, multilocus sequence typing analysis, and plasmid typing were also performed. RESULTS: A high prevalence of ESBLs, carbapenemases, fosfomycin, and colistin resistance genes was evidenced among those isolates. The predominant ESBL identified was blaCTX-M-15, followed by blaCTX-M-9. We also identified blaNDM-5 and blaOXA-244. fosA3, fosA4, and fosA6 were identified in E. coli isolates. In addition, 11 MCR-1 producers were recovered. Notably, co-occurrence of ESBL genes and mcr or fosA genes was observed. MLST analysis revealed a high clonal diversity, ruling out the dissemination of one major clone. IncFIB-type was predominantly present among ESBL and FosA producers. The blaNDM-5 gene was carried on an IncX4-type, although the blaOXA-244 gene was chromosomally located. The mcr-1 gene was carried on a diversity of plasmids (IncI2, IncX4, and IncHI2). CONCLUSION: These results raise serious public health concerns as Egyptian pig meat could serve as a reservoir for antimicrobial resistance genes (ARGs), leading to worldwide dissemination.

ABO genotype alters the gut microbiota by regulating GalNAc levels in pigs.

The composition of the intestinal microbiome varies considerably between individuals and is correlated with health1. Understanding the extent to which, and how, host genetics contributes to this variation is essential yet has proved to be difficult, as few associations have been replicated, particularly in humans2. Here we study the effect of host genotype on the composition of the intestinal microbiota in a large mosaic pig population. We show that, under conditions of exacerbated genetic diversity and environmental uniformity, microbiota composition and the abundance of specific taxa are heritable. We map a quantitative trait locus affecting the abundance of Erysipelotrichaceae species and show that it is caused by a 2.3 kb deletion in the gene encoding N-acetyl-galactosaminyl-transferase that underpins the ABO blood group in humans. We show that this deletion is a ≥3.5-million-year-old trans-species polymorphism under balancing selection. We demonstrate that it decreases the concentrations of N-acetyl-galactosamine in the gut, and thereby reduces the abundance of Erysipelotrichaceae that can import and catabolize N-acetyl-galactosamine. Our results provide very strong evidence for an effect of the host genotype on the abundance of specific bacteria in the intestine combined with insights into the molecular mechanisms that underpin this association. Our data pave the way towards identifying the same effect in rural human populations.

A Bacterial Genome and Culture Collection of Gut Microbial in Weanling Piglet.

The microbiota hosted in the pig gastrointestinal tract are important to health of this biomedical model. However, the individual species and functional repertoires that make up the pig gut microbiome remain largely undefined. Here we comprehensively investigated the genomes and functions of the piglet gut microbiome using culture-based and metagenomics approaches. A collection included 266 cultured genomes and 482 metagenome-assembled genomes (MAGs) that were clustered to 428 species across 10 phyla was established. Among these clustered species, 333 genomes represent potential new species. Less matches between cultured genomes and MAGs revealed a substantial bias for the acquisition of reference genomes by the two strategies. Glycoside hydrolases was the dominant category of carbohydrate-active enzymes. Four-hundred forty-five secondary metabolite biosynthetic genes were predicted from 292 genomes with bacteriocin being the most. Pan genome analysis of Limosilactobacillus reuteri uncover the biosynthesis of reuterin was strain-specific and the production was experimentally determined. This study provides a comprehensive view of the microbiome composition and the function landscape of the gut of weanling piglets and a valuable bacterial resource for further experimentations. IMPORTANCE The microorganism communities resided in mammalian gastrointestinal tract impacted the health and disease of the host. Our study complements metagenomic analysis with culture-based approach to establish a bacteria and genome collection and comprehensively investigate the microbiome composition and function of the gut of weanling piglets. We provide a valuable resource for further study of gut microbiota of weanling piglet and development of probiotics for prevention of disease.

Salmonella Infection Causes Hyperglycemia for Decreased GLP-1 Content by Enteroendocrine L Cells Pyroptosis in Pigs.

Inflammatory responses have been shown to induce hyperglycemia, yet the underlying mechanism is still largely unclear. GLP-1 is an important intestinal hormone for regulating glucose homeostasis; however, few studies have investigated the influence of digestive tract Salmonella infection on enteroendocrine L cell secretions. In this study, we established a model of Salmonella-infected piglets by oral gavage in order to analyze the effects of Salmonella infection on enteroendocrine L cell function. Furthermore, in vitro lipopolysaccharide (LPS) was administered to STC-1 cells to clarify its direct effect on GLP-1 secretion. The results showed that significantly increased blood glucose in the group of Salmonella-infected piglets was observed, and Salmonella infection decreased blood GLP-1 content. Then, ileal epithelium damage was observed by histological detection, and this was further verified by TUNEL staining. We identified activation of TLR signaling demonstrating up-regulated expressions of TLR4 and nuclear factor-kappa B (NF-ΚB). Furthermore, it was shown that Salmonella induced pyroptosis of enteroendocrine L cells and enhanced the secretion of IL-1ß through augmenting gene and protein expressions of NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a carboxyl-terminal CARD (ASC), Caspase 1, and gasdermin D (GSDMD). Meanwhile, in vitro LPS treatment induced the pyroptosis of STC-1 cells and reduced the secretion of GLP-1. Altogether, the results demonstrated that Salmonella infection can reduce secretion of GLP-1 by inducing pyroptosis of intestinal L cells, which may eventually result in hyperglycemia. The results provided evidence for the cause of hyperglycemia induced by inflammation and shed new light on glucose homeostasis regulation.

Escherichia fergusonii, an Underrated Repository for Antimicrobial Resistance in Food Animals.

A total of 1,400 samples of food animals (pigs, chickens, and ducks) were collected between July and September 2019 in China to uncover the prevalence of E. fergusonii and its potential role in the evolution of antimicrobial resistance (AMR). An isolation of E. fergusonii was performed and pulsed-field gel electrophoresis (PFGE) was used to uncover the genetic relationship. The AMR of E. fergusonii isolates was comprehensively characterized using broth microdilution-based antimicrobial susceptibility testing, S1-PFGE, southern hybridization, whole-genome sequencing, and in-depth bioinformatics analysis. As a result, a total of 133 E. fergusonii isolates were obtained. These isolates could be grouped into 41 PFGE subclades, suggesting a diverse genetic relationship. The resistance phenotypes of sulfafurazole (97.74%) and tetracycline (94.74%) were the most frequently found. Of the E. fergusonii isolates, 51.88% were extended spectrum beta-lactamase (ESBL)-positive. Forty-three different AMR genes were revealed based on 25 genome sequences harboring mcr-1. Briefly, aph(6)-Id, aph(3'')-Ib and tet(A) genes were the most frequently observed, with the highest rate being 76.00% (19/25). Three mcr-1-harboring plasmids were identified after Nanopore sequencing, including pTB31P1 (IncHI2-IncHI2A, 184,652 bp), pTB44P3 (IncI2, 62,882 bp), and pTB91P1 (IncHI2-IncHI2A, 255,882 bp). Additionally, 25 E. fergusonii isolates harboring mcr-1 were clustered together with other E. fergusonii isolates from different regions and sources available in GenBank, suggesting a possible random process of mcr-1 transmission in E. fergusonii. In conclusion, E. fergusonii is widespread in food animals in China and might be an important reservoir of AMR genes, especially mcr-1, and facilitate the evolution of AMR. IMPORTANCEE. fergusonii, a member of the genus Escherichia, has been reported to transmit via the food chain and cause diseases in humans. However, the prevalence of multidrug-resistant E. fergusonii, especially mcr-1-positive E. fergusonii isolates, has rarely been reported. Here, we collected 1,400 samples from food animals in three provinces of China and obtained 133 E. fergusonii isolates (9.5%). We found that the prevalence of E. fergusonii isolates was diverse, with high levels of antimicrobial resistance. Among them, 18.8% E. fergusonii isolates carried the colistin resistance gene mcr-1. Thus, E. fergusonii may facilitate the evolution of colistin resistance as a reservoir of mcr-1. As far as we know, the prevalence and AMR of E. fergusonii in the food animals in this study was first reported in China. These findings increase our understanding of the role of E. fergusonii in public health and the evolution of antibiotic resistance.

Macrococcus armenti sp. nov., a novel bacterium isolated from the skin and nasal cavities of healthy pigs and calves.

Gram-positive coccoid bacteria were isolated from the nasal cavities of pigs and calves as well as from axillar and inguinal skin regions of pigs. Phylogenetic analysis of seven strains based on complete genome, 16S rRNA, hsp60, dnaJ, rpoB and sodA gene sequences and MALDI-TOF MS profiles revealed that they belonged to the genus Macrococcus with the closest relatedness to Macrococcus canis, Macrococcus caseolyticus subsp. caseolyticus and Macrococcus caseolyticus subsp. hominis. DNA relatedness of the type strain JEK37T with the type strains of M. canis, M. caseolyticus subsp. caseolyticus and M. caseolyticus subsp. hominis was 23.4, 23.1 and 23.0 % by digital DNA-DNA hybridization and 80.39, 80.45 and 80.87 % by average nucleotide identity (ANI) calculations, confirming that they do not belong to the same species. The DNA G+C content of JEK37T was 35.65 mol%. The novel strains can be differentiated from M. canis KM 45013T by the ability to fermentate d-ribose and by the absence of DNAase production and haemolysis, from M. caseolyticus subsp. caseolyticus CCUG 15606T by the ability to fermentate sucrose and from both species by the inability to grow in 9 and 12% NaCl. They differ from M. caseolyiticus subsp. hominis by the presence of α-glucosidase. The most common fatty acids of JEK37T were C14 : 0, C18 : 1 ω9c and C18 : 0. Known polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, aminolipid, aminoglycolipid, aminophospholipid, glycolipid and phospholipid. Cell-wall peptidoglycan of JEK37T was of type A3α l-Lys-Gly2-L-Ser-Gly (similar to A11.3) and the respiratory quinolone was menaquinone 6. Based on their genotypic and chemotaxonomic characteristics, these strains represent a novel species of the genus Macrococcus, for which we propose the name Macrococcus armenti sp. nov. The type strain is JEK37T (=DSM 112712T=CCOS 1982T).

16S rRNA gene sequencing analysis of gut microbiome in a mini-pig diabetes model.

BACKGROUND: Currently, increasing attention is being paid to the important role of intestinal microbiome in diabetes. However, few studies have evaluated the characteristics of gut microbiome in diabetic miniature pigs, despite it being a good model animal for assessing diabetes. METHODS: In this study, a mini-pig diabetes model (DM) was established by 9-month high-fat diet (HFD) combined with low-dose streptozotocin, while the animals fed standard chow diet constituted the control group. 16S ribosomal RNA (rRNA) gene sequencing was performed to assess the characteristics of the intestinal microbiome in diabetic mini-pigs. RESULTS: The results showed that microbial structure in diabetic mini-pigs was altered, reflected by increases in levels of Coprococcus_3 and Clostridium_sensu_stricto_1, which were positively correlated with diabetes, and decreases in levels of the bacteria Rikenellaceae, Clostridiales_vadinBB60_group, and Bacteroidales_RF16_group, which were inversely correlated with blood glucose and insulin resistance. Moreover, PICRUSt-predicted pathways related to the glycolysis and Entner-Doudoroff superpathway, enterobactin biosynthesis, and the l-tryptophan biosynthesis were significantly elevated in the DM group. CONCLUSION: These results reveal the composition and predictive functions of the intestinal microbiome in the mini-pig diabetes model, further verifying the relationship between HFD, gut microbiome, and diabetes, and providing novel insights into the application of the mini-pig diabetes model in gut microbiome research.

Exploring the role of gut microbiota in host feeding behavior among breeds in swine.

BACKGROUND: The interplay between the gut microbiota and feeding behavior has consequences for host metabolism and health. The present study aimed to explore gut microbiota overall influence on feeding behavior traits and to identify specific microbes associated with the traits in three commercial swine breeds at three growth stages. Feeding behavior measures were obtained from 651 pigs of three breeds (Duroc, Landrace, and Large White) from an average 73 to 163 days of age. Seven feeding behavior traits covered the information of feed intake, feeder occupation time, feeding rate, and the number of visits to the feeder. Rectal swabs were collected from each pig at 73 ± 3, 123 ± 4, and 158 ± 4 days of age. DNA was extracted and subjected to 16 S rRNA gene sequencing. RESULTS: Differences in feeding behavior traits among breeds during each period were found. The proportion of phenotypic variances of feeding behavior explained by the gut microbial composition was small to moderate (ranged from 0.09 to 0.31). A total of 21, 10, and 35 amplicon sequence variants were found to be significantly (q-value < 0.05) associated with feeding behavior traits for Duroc, Landrace, and Large White across the three sampling time points. The identified amplicon sequence variants were annotated to five phyla, with Firmicutes being the most abundant. Those amplicon sequence variants were assigned to 28 genera, mainly including Christensenellaceae_R-7_group, Ruminococcaceae_UCG-004, Dorea, Ruminococcaceae_UCG-014, and Marvinbryantia. CONCLUSIONS: This study demonstrated the importance of the gut microbial composition in interacting with the host feeding behavior and identified multiple archaea and bacteria associated with feeding behavior measures in pigs from either Duroc, Landrace, or Large White breeds at three growth stages. Our study provides insight into the interaction between gut microbiota and feeding behavior and highlights the genetic background and age effects in swine microbial studies.

Turicibacter bilis sp. nov., a novel bacterium isolated from the chicken eggshell and swine ileum.

Three novel, anaerobic, Gram-positive bacteria were isolated from the eggshell of two separate white leghorn chicken flocks and the ileum of a healthy pig, and designated MMM721T, ISU324 and PIG517 respectively. Cells were pleomorphic and capable of forming long chains of rods or coccoid clusters. Phylogenetic analysis of the 16S rRNA gene sequences identified these strains to be within the genus Turicibacter, of which only one species, Turicibacter sanguinis, has been formally described. However, whole genome sequencing of novel isolates returned a digital DNA-DNA hybridization value of 22.5 % and average nucleotide identity (ANI) values of 76.4 % (ANIb) and 86.0 % (ANIm), indicating divergence between the type strain MMM721T and T. sanguinis, suggesting the strains represented a novel species. The major fatty acid methyl esters of strain MMM721T were C16 : 0, C18 : 1 ω7c and C18 : 0. The strains mainly produced the volatile fatty acid lactate, along with smaller amounts of acetate and butyrate. Together, these data indicate that MMM721T, along with ISU324 and PIG517, represent a novel species within the genus Turicibacter. We propose the name Turicibacter bilis sp. nov. for the species. The type strain is MMM721T (=ATCC TSD-238T=CCUG 74757T).

Leptospirosis is an emerging infectious disease of pig-hunting dogs and humans in North Queensland.

BACKGROUND: Leptospirosis is a zoonotic disease with a worldwide distribution, caused by pathogenic serovars in the genus Leptospira. Feral pigs are known carriers of Leptospira species and pig hunting using dogs is a common recreational activity in Queensland, Australia. METHODOLOGY AND PRINCIPAL FINDINGS: This study aimed to determine the seroprevalence of Leptospira spp. serovars in pig-hunting dogs above the Tropic of Capricorn in Queensland and by establishing the geographic distribution, serovars and incidence of human cases of leptospirosis in Queensland, identify potential overlap between human and canine exposure. We also explored the knowledge and risk-taking behaviours of pig-hunting dog owners towards zoonotic diseases. Ninety-eight pig-hunting dogs deemed healthy by physical examination and owned by 41 people from Queensland had serum submitted for Microscopic Agglutination Testing (MAT) to determine antibody titres against Leptospira serovars, while 40/41 dog owners completed a survey on their knowledge of diseases relating to pig hunting. Human leptospirosis cases (n = 330) notified to Queensland Health between 2015-2018 were analysed. Approximately one quarter (23/87; 26%) of unvaccinated pig-hunting dogs were seropositive to Leptospira spp. Although harder to interpret, 8/11 (73%) vaccinated dogs were seropositive to Leptospira spp. Pig hunters may be more likely to contract leptospirosis compared with the general Queensland population, based on responses from surveyed hunters. The highest concentration of human leptospirosis was in the wet tropics region of Far North Queensland. There was little overlap between the serovars dogs were exposed to and those infecting humans. The dominant serovar identified in unvaccinated dogs was Australis (13/23; 57%), with serovar Arborea (36/330; 10.9%) responsible for the highest number of human leptospirosis cases. Topaz was the second most common serovar in both humans and dogs and was previously unrecorded in Australian dogs. Most hunters surveyed used hand washing as a zoonotic disease risk reduction technique. CONCLUSIONS: Leptospirosis is an emerging disease of growing significance. The infection requires a 'one health' approach to understand its epidemiology. With shifting climatic patterns influencing human-animal-environment interactions, ongoing monitoring of diseases like leptospirosis is critical to helping prevent infection of individuals and disease outbreaks.

Development of an indirect competitive enzyme linked immunosorbent assay for the quantitative detection of Mycoplasma hyopneumoniae during the vaccine production process.

Inactivated Mycoplasma hyopneumoniae vaccine is used extensively to control M. hyopneumoniae infection worldwide. Quantification techniques are essential in the process of standardizing and validating vaccines. In this study, we developed and optimized an indirect competitive enzyme linked immunosorbent assay (ic-ELISA) for the rapid quantification of M. hyopneumoniae antigen during vaccine production. Briefly, whole M. hyopneumoniae antigen was coated onto microtiter plates, and a polyclonal antibody against M. hyopneumoniae recombinant elongation factor thermo unstable (EF-Tu) protein was prepared and added with the samples to be tested. The methods were optimized and showed significant reproducibility, with coefficients of variation of 4.01% and 6.14% for the intra-and inter-assays, respectively. Quantification of M. hyopneumoniae cultures at different growth stages using the ic-ELISA test showed a similar curve to that of the traditional color changing units (CCU) assay, with a delay in the time when the amount reached the peak and started to fall. In the inactivated vaccine production process, the cultures could be harvested later than that for the live vaccine, at about 12 h after the end of the logarithmic growth phase. Different batches of cultures were measured for their relative potency value compared with the in-house reference vaccine, which was used to determine whether the cultures met the antigen amount requirements for vaccine preparation. The curves of the CCU titer and ic-ELISA titer in the logarithmic phase correlated strongly and a linear regression equation was established to calculate the CCU values rapidly using the ic-ELISA results. In conclusion, an ic-ELISA method was established to rapidly assess the amount of antigen in an M. hyopneumoniae culture during the vaccine production process.

Effects of 25-hydroxyvitamin D3 on growth performance, serum parameters, fecal microbiota, and metabolites in weaned piglets fed diets with low calcium and phosphorus.

BACKGROUND: With the development of intensive farming, long-term exposure of pigs to poor light conditions is not conducive to the production of vitamin D3 , and vitamin D3 deficiency could affect absorption and metabolism of calcium (Ca) and phosphorus (P). 25-Hydroxyvitamin D3 (25OHD3 ) has higher bioactivity than regular vitamin D3 . This study investigated the effects of 25OHD3 on performance, serum parameters, fecal microbiota, and metabolites in weaned piglets fed with low Ca-P diet. RESULTS: It was found that a low Ca-P diet supplemented with 50 µg/kg 25OHD3 (NC + 25-D) improved (P < 0.05) average daily gain (ADG) in phase 2 and in the overall period of the experiment, and increased (P < 0.05) the immunoglobulin G (IgG), immunoglobulin A (IgA), catalase (CAT), bone-specific alkaline phosphatase (BALP), and osteocalcin (OC) serum content on day 28 compared with a low Ca-P diet (NC), but no differences were observed between a normal Ca-P diet (PC) and the NC + 25-D diet. Compared with NC, the abundance of Firmicutes was higher (P < 0.05) in PC and NC + 25-D. NC + 25-D decreased (P < 0.05) the abundance of Streptococcaceae compared with PC and NC, and increased (P < 0.05) the abundance of Lachnospiraceae compared with NC. Serum 25OHD3 was negatively correlated with the abundance of fecal Streptococcaceae (P < 0.05), and positively correlated with the abundance of fecal Lachnospiraceae (P < 0.05). CONCLUSION: Supplementation of 25OHD3 in a low Ca-P diet improved serum immunity, bone biochemical parameters, and fecal microbiota such as decreased Streptococcaceae abundance and increased Lachnospiraceae abundance, which could subsequently promote growth of piglets. The effects were similar to that of a normal Ca-P diet. © 2021 Society of Chemical Industry.

Effect of parity segregation on Mycoplasma hyopneumoniae infection dynamics and pneumonic lesions in pigs / Efeito da ordem de parto segregada sobre a dinâmica de infecção de Mycoplasma hyopneumoniae e lesões pulmonares ao abate

Gilts represent a group risk for Mycoplasma hyopneumoniae vertical transmission in swine herds. Therefore, parity segregation can be an alternative to control M. hyopneumoniae infections. The study evaluated the effect of parity segregation on M. hyopneumoniae infection dynamics and occurrence and severity of lung lesions at slaughter. For that, three multiple site herds were included in the study. Herd A consisted of the farm where gilts would have their first farrowing (parity order (PO) 1). After the first farrowing PO 1 sows were transferred to herd B (PO2-6). Herd C was a conventional herd with gilt replacement (PO1-6). Piglets born in each herd were raised in separated nursery and finishing units. Sows (n = 33 (A), 37 (B), 34 (C)) in all herds were sampled prior to farrowing and piglets (n = 54 (A), 71 (B), 66 (C)) were sampled longitudinally at 21, 63, 100, 140 days of age and at slaughter for M. hyopneumoniae detection by PCR and lung lesions scoring. M. hyopneumoniae prevalence in sows did not differ among herds. Prevalence of positive piglets was higher at weaning in the PO1 herd (A) (P < 0.05). However, prevalence of positive pigs from 100 days of age to slaughter age was higher in the PO2-6 herd (B) (P < 0.05). Lung lesion occurrence and severity were higher in herd B. The authors suggested that the lack of a proper gilt acclimation might have influenced the results, leading to sows being detected positive at farrowing, regardless of the parity.

As leitoas consistem em um grupo de risco na transmissão vertical de Mycoplasma hyopneumoniae dentro do sistema de produção de suínos. Dessa forma, a segregação de partos poderia ser utilizada como alternativa para controlar as infecções por M. hyopneumoniae. O objetivo deste estudo foi avaliar o efeito da segregação de partos sobre a dinâmica de infecção de M. hyopneumoniae e a ocorrência e severidade das lesões pulmonares ao abate. Para isso três sistemas de produção de suínos com três sítios cada foram incluídos no estudo. A granja A consistia da unidade onde as leitoas tem o primeiro parto, ou seja, alojava somente de fêmeas de ordem de parto 1 (Granja OP1). Após o primeiro parto as fêmeas OP1 foram transferidas para a granja B (Granja OP2-6), ou seja, consistia de fêmeas de ordem de parto 2 a 6, e a granja C consistiu em uma granja convencional com reposição de leitoas (Granja OP1-6), com fêmeas de ordem de parto 1 a 6. Os leitões nascidos de cada granja foram transferidos e criados em creches e terminações segregadas. As matrizes (n = 33 (A), 37 (B), 34 (C)) de todas as granjas do estudo foram amostradas previamente ao parto e os leitões (n = 54 (A), 71 (B), 66 (C)) foram amostrados longitudinalmente aos 21, 63, 100 e 140 dias de idade e ao abate. Em todos os momentos de coleta, as amostras foram avaliadas por PCR para detecção de M. hyopneumoniae. As lesões pulmonares foram avaliadas e escores de lesão foram atribuídos ao abate. A prevalência de matrizes positivas para M. hyopneumoniae não diferiu entre as granjas (P > 0,05). A prevalência ao desmame foi maior na granja A (OP1) (P < 0,05). No entanto, dos 100 dias de idade até o abate a prevalência de leitões positivos para M. hyopneumoniae foi maior na granja B (OP2-6) (P < 0,05). A ocorrência e severidade de lesões pulmonares foram maiores na granja B. Os autores sugerem que a falta de uma aclimatação adequada das leitoas pode ter influenciado nos resultados, levando à detecção de matrizes positivas ao parto, independente da ordem de parto.

Intestinal microbiota modulation and improved growth in pigs with post-weaning antibiotic and ZnO supplementation but only subtle microbiota effects with Bacillus altitudinis.

The objective was to evaluate the effect of dietary Bacillus altitudinis spore supplementation during day (D)0-28 post-weaning (PW) and/or D29-56 PW compared with antibiotic and zinc oxide (AB + ZnO) supplementation on pig growth and gut microbiota. Eighty piglets were selected at weaning and randomly assigned to one of five dietary treatments: (1) negative control (Con/Con); (2) probiotic spores from D29-56 PW (Con/Pro); (3) probiotic spores from D0-28 PW (Pro/Con); (4) probiotic spores from D0-56 PW (Pro/Pro) and (5) AB + ZnO from D0-28 PW. Overall, compared with the AB + ZnO group, the Pro/Con group had lower body weight, average daily gain and feed intake and the Pro/Pro group tended to have lower daily gain and feed intake. However, none of these parameters differed between any of the probiotic-treated groups and the Con/Con group. Overall, AB + ZnO-supplemented pigs had higher Bacteroidaceae and Prevotellaceae and lower Lactobacillaceae and Spirochaetaceae abundance compared to the Con/Con group, which may help to explain improvements in growth between D15-28 PW. The butyrate-producing genera Agathobacter, Faecalibacterium and Roseburia were more abundant in the Pro/Con group compared with the Con/Con group on D35 PW. Thus, whilst supplementation with B. altitudinis did not enhance pig growth performance, it did have a subtle, albeit potentially beneficial, impact on the intestinal microbiota.

Sporadic Dissemination of tet(X3) and tet(X6) Mediated by Highly Diverse Plasmidomes among Livestock-Associated Acinetobacter.

The emergence of high-level tigecycline resistance mediated by plasmid-borne tet(X) genes greatly threatens the clinical effectiveness of tigecycline. However, the dissemination pattern of plasmid-borne tet(X) genes remains unclear. We here recovered tet(X)-positive Acinetobacter isolates from 684 fecal and environmental samples collected at six livestock farms. Fifteen tet(X)-positive Acinetobacter isolates were identified, mainly including 9 tet(X3)- and 5 tet(X6)-positive Acinetobacter towneri isolates. A clonal dissemination of tet(X3)-positive A. towneri was detected in a swine farm, while the tet(X6)-positive A. towneri isolates mainly disseminated sporadically in the same farm. A tet(X3)-carrying plasmid (pAT181) was self-transmissible from a tigecycline-susceptible A. towneri strain to Acinetobacter baumannii strain ATCC 17978, causing 64- to 512-fold increases in the MIC values of tetracyclines (including tigecycline). Worrisomely, pAT181 was stably maintained and increased the growth rate of strain ATCC 17978. Further identification of tet(X) genes in 10,680 Acinetobacter genomes retrieved from GenBank revealed that tet(X3) (n = 249), tet(X5)-like (n = 61), and tet(X6) (n = 53) were the prevalent alleles mainly carried by four species, and most of them were livestock associated. Phylogenetic analysis showed that most of the tet(X3)- and tet(X6)-positive isolates disseminated sporadically. The structures of the tet(X3), and tet(X6) plasmidomes were highly diverse, and no epidemic plasmids were detected. However, cross-species and cross-region transmissions of tet(X3) might have been mediated by several plasmids in a small proportion of strains. Our study implies that horizontal plasmid transfer may be insignificant for the current dissemination of tet(X3) and tet(X6) in Acinetobacter strains. Continuous surveillance for tet(X) genes in the context of One Health is necessary to prevent them from transmitting to humans. IMPORTANCE Recently identified plasmid-borne tet(X) genes have greatly challenged the efficiency of tigecycline, a last-resort antibiotic for severe infection, while the dissemination pattern of the plasmid-borne tet(X) genes remains unclear. In this study, we identified a clonal dissemination of tet(X3)-positive A. towneri isolates on a swine farm, while the tet(X6)-positive A. towneri strains mainly disseminated sporadically on the same farm. Of more concern, a tet(X3)-carrying plasmid was found to be self-transmissible, resulting in enhanced tigecycline resistance and growth rate of the recipient. Further exploration of a global data set of tet(X)-positive Acinetobacter genomes retrieved from GenBank revealed that most of the tet(X3)- and tet(X6)-positive isolates shared a highly distant relationship, and the structures of tet(X3) and tet(X6) plasmidomes exhibited high mosaicism. Notably, some of the isolates belong to Acinetobacter species that are opportunistic pathogens and have been identified as sources of nosocomial infections, raising concerns about transmission to humans in the future. Our study evidenced the sporadic dissemination of tet(X3) and tet(X6) in Acinetobacter strains and the necessity of continuous surveillance for tet(X) genes in the context of One Health.

Fruit and Vegetable Supplemented Diet Modulates the Pig Transcriptome and Microbiome after a Two-Week Feeding Intervention.

A study was conducted to determine the effects of a diet supplemented with fruits and vegetables (FV) on the host whole blood cell (WBC) transcriptome and the composition and function of the intestinal microbiome. Nine six-week-old pigs were fed a pig grower diet alone or supplemented with lyophilized FV equivalent to half the daily recommended amount prescribed for humans by the Dietary Guideline for Americans (DGA) for two weeks. Host transcriptome changes in the WBC were evaluated by RNA sequencing. Isolated DNA from the fecal microbiome was used for 16S rDNA taxonomic analysis and prediction of metabolomic function. Feeding an FV-supplemented diet to pigs induced differential expression of several genes associated with an increase in B-cell development and differentiation and the regulation of cellular movement, inflammatory response, and cell-to-cell signaling. Linear discriminant analysis effect size (LEfSe) in fecal microbiome samples showed differential increases in genera from Lachnospiraceae and Ruminococcaceae families within the order Clostridiales and Erysipelotrichaceae family with a predicted reduction in rgpE-glucosyltransferase protein associated with lipopolysaccharide biosynthesis in pigs fed the FV-supplemented diet. These results suggest that feeding an FV-supplemented diet for two weeks modulated markers of cellular inflammatory and immune function in the WBC transcriptome and the composition of the intestinal microbiome by increasing the abundance of bacterial taxa that have been associated with improved intestinal health.