Neomycin resistance in clinical Escherichia coli from Danish weaner pigs is associated with recent neomycin use and presence of F4 or F18 fimbriaes.

Neomycin is a first-choice antibiotic for treatment of porcine enteritis caused by enterotoxigenic Escherichia coli (ETEC), but little is known about factors influencing resistance to this drug. The aims of this study were to assess antimicrobial resistance and virulence in 325 E. coli isolates obtained in 2020 from various infections in pigs, and to identify factors associated with neomycin resistance development. Susceptibility to 16 antimicrobial agents was determined by broth microdilution, and occurrence of ETEC-associated virulence factors was screened by PCR and hemolysis on blood agar. Univariate and multivariate logistic regression analyses were performed to determine if age group, virulence factors, or antibiotic use (neomycin and other antibiotics) were associated with neomycin resistance. STa, STb, LT, F4, and F18 were detected in 14%, 37%, 26%, 21% and 23% of the isolates, respectively. Resistance was low for antimicrobials of high public health importance (1.5% for cefotaxime, 1% for colistin and no fluoroquinolone resistance) but high for drugs used for treatment of ETEC enteritis (e.g. 20% for neomycin). Isolates with the ETEC pathotype were significantly associated with the weaner age group and intestinal/fecal origin. Multivariate analysis showed that recent neomycin use and presence of F4 or F18 were significantly associated with neomycin resistance amongst isolates from weaners. These results prove an association between neomycin resistance and use at the farm level. Further research is warranted to determine why neomycin resistance was associated with F4 and F18, and whether neomycin use may co-select for virulent strains.

Risk factors for gastric ulceration in nursery pigs.

Pars oesophageal gastric ulceration is a prevalent condition in swine production and often observed in finisher pigs at slaughter, with reported prevalence ranging from 32 % to 65 %. Although feeding practices in nursery and finisher pigs are similar, little is known about the prevalence and risk factors associated with this condition in nursery pigs. This prospective cohort study aimed to identify risk factors for gastric ulceration in nursery pigs. The objectives were: (1) to estimate the association of inherent pig characteristics (birth weight, sex, parity of sow, litter size, and pen fouling behavior) and management characteristics (antibiotic treatment during suckling and post weaning, piglet rotation during lactation, weaning age and weight) with pars oesophageal lesions; (2) to assess differences in gastric dry matter content, pH, and percentage of solid particles in the sediment in relation to pars oesophageal lesions. A total of 58 piglets tracked from birth to 20 days of age and 210 nursery pigs from birth to 10 weeks of age were euthanized and assessed for pars oesophageal gastric lesions. All nursery pigs were fed finely ground pelleted feed ad libitum. Based on a macroscopic assessment, the pigs' stomachs were categorized as either LPO "lesions in the Pars oesophagea" or NLPO "no lesions in the Pars oesophagea". We observed an overall prevalence of 26.2 % (CI 95 % 0.67 - 0.79) for LPO in nursery pigs at 10 weeks of age, while no lesion were observed in piglets at 20 days of age. A mixed multivariable logistic regression model showed that piglets with a birthweight ≤ 1.11 kg had a 2.11 times higher odds of LPO (CI 95 % 1.02-4.37, p = 0.04) than their heavier counterparts. There was a weak association of LPO in nursery pigs from pens with pen fouling with a 2.14 times higher odds (CI 95 % 0.97-4.72 p = 0.05). Nursery pigs with LPO presented an increased gastric content fluidity with a reduced (p ≤ 0.01) gastric dry matter content (18.7 %), solid phase (74.7 %), and pH (3.22) compared to NLPO nursery pigs (20.5 %, 85.09 %, 3.56, respectively). We have demonstrated that LPO are present in nursery pigs as early as 10 weeks of age. We observed that low birth weight is associated with higher odds for LPO in nursery pigs. These observations might help explain differences between individual pigs in terms of LPO development when groups are exposed to similar environments, management, and feeding conditions.

Association between gastric content fluidity and pars oesophageal ulcers in nursery pigs: a cross-sectional study of high-risk Danish herds using commercial feed.

BACKGROUND: The objective of this cross-sectional study was to assess the within-herd prevalence of pars oesophageal ulcers (POU) in high-risk Danish herds using commercial diets. Furthermore, we aimed to estimate the association between gastric content fluidity and POU using a generalised additive model (GAM). The study included 200 clinically healthy nursery pigs randomly selected from ten farms (20 pigs from each farm). The 10 farms were selected based on a suspected high prevalence of gastric ulcers. Post-mortem gastric ulcer assessment was based on macroscopic lesions, and gastric content fluidity was assessed based on the solid particle sedimentation percentage (solid phase). RESULTS: We observed an overall prevalence of 35.5% for POU in nursery pigs. Within-herd prevalence varied considerably among farms, with values ranging from 0% in Farm 1 to 84% in Farm 4. Our model showed strong associations between POU and gastric content fluidity (P < 0.001), as well as between POU and farm of origin (P < 0.001). In addition, we observed that the risk of POU decreased non-linearly as the gastric content solid phase percentage increased, i.e. as the gastric content became more solid. CONCLUSION: We have demonstrated that pars oesophageal ulcers are present in Danish herds with nursery pigs fed commercial diets. Furthermore, we have established that gastric content fluidity is strongly associated with POU in nursery pigs. Even so, we cannot conclude that gastric content fluidity is solely responsible for POU. Future research should look into the association between pars oesophageal ulcers and both farm management activities and individual pig factors.

F4- and F18-Positive Enterotoxigenic Escherichia coli Isolates from Diarrhea of Postweaning Pigs: Genomic Characterization.

This study aimed to characterize in silico enterotoxigenic Escherichia coli F4- and F18-positive isolates (n = 90) causing swine postweaning diarrhea, including pathogenic potential, phylogenetic relationship, antimicrobial and biocide resistance, prophage content, and metal tolerance rates. F4 strains belonged mostly to the O149 and O6 serogroups and ST100 and ST48 sequence types (STs). F18 strains were mainly assigned to the O8 and O147 serogroups and ST10, ST23, and ST42. The highest rates of antimicrobial resistance were found against streptomycin, sulfamethoxazole, tetracycline, trimethoprim, and ampicillin. No resistance was found toward ciprofloxacin, cefotaxime, ceftiofur, and colistin. Genes conferring tolerance to copper (showing the highest diversity), cadmium, silver, and zinc were predicted in all genomes. Enterotoxin genes (ltcA, 100% F4, 62% F18; astA, 100% F4, 38.1% F18; sta, 18.8% F4, 38.1% F18; stb, 100% F4, 76.2% F18) and fimbria-encoding genes typed as F4ac and F18ac were detected in all strains, in addition to up to 16 other virulence genes in individual strains. Phage analysis predicted between 7 and 20 different prophage regions in each strain. A highly diverse variety of plasmids was found; IncFII, IncFIB, and IncFIC were prevalent among F4 isolates, while IncI1 and IncX1 were dominant among F18 strains. Interestingly, F4 isolates from the early 1990s belonged to the same clonal group detected for most of the F4 strains from 2018 to 2019 (ONT:H10-A-ST100-CH27-0). The small number of single-nucleotide polymorphism differences between the oldest and recent F4 ST100 isolates suggests a relatively stable genome. Overall, the isolates analyzed in this study showed remarkably different genetic traits depending on the fimbria type.IMPORTANCE Diarrhea in the postweaning period due to enterotoxigenic E. coli (ETEC) is an economically relevant disease in pig production worldwide. In Denmark, prevention is mainly achieved by zinc oxide administration (to be discontinued by 2022). In addition, a breeding program has been implemented that aims to reduce the prevalence of this illness. Treatment with antimicrobials contributes to the problem of antimicrobial resistance (AMR) development. As a novelty, this study aims to deeply understand the genetic population structure and variation among diarrhea-associated isolates by whole-genome sequencing characterization. ST100-F4ac is the dominant clonal group circulating in Danish herds and showed high similarity to ETEC ST100 isolates from China, the United States, and Spain. High rates of AMR and high diversity of virulence genes were detected. The characterization of diarrhea-related ETEC is important for understanding the disease epidemiology and pathogenesis and for implementation of new strategies aiming to reduce the impact of the disease in pig production.

The dose-response relationship between the amount of straw provided on the floor and gastric ulceration of pars oesophagea in growing pigs.

The aim of the present study was (1) to determine the dose-response relationship between the amount of straw provided on the floor and oesophageal ulceration in pigs kept under typical Danish production conditions (18 pigs/pen, 0.7m2/pig, partly slatted floor, ad libitum access to feed), (2) to reveal whether straw ingestion explains the effect of straw provision on the stomach health and (3) to elucidate the effect of straw ingestion on the stomach conditions. Data were collected at slaughter (approximately 100kg body weight) on pigs provided with straw amounts in the range 10g to 500g wheat straw/pig/day from 30kg body weight and fed a wheat-based pelleted feed added 15% non-heated and non-pelleted rolled barley. Aims (1) and (2) included 712 pigs kept in 42 pens, whereas (3) was studied on a subset of 37 pigs with either none or obvious amounts of straw in the stomach. The amount of straw provided affected stomach health in a curvilinear manner. Provision of up to approximately 300gstraw/pig/day progressively decreased the risk of oesophageal ulceration and scarring. At larger amounts of straw the ulceroprotective effect of straw was reduced which requires further investigation. Straw ingestion appeared to be an essential intermediary factor for the improvement of stomach health. The number of pigs without straw in the stomach decreased linearly with the logarithm of the amount of straw provided, which explained the effect of straw provision on stomach health. The structure of the stomach contents increased as revealed by the increased weight and dry matter content and decreased sedimentation of the digesta. Provision of approximately 300gstraw/pig/day on the floor may be a potential strategy to reduce, but not inevitably eliminate, oesophageal ulceration in pigs in commercial pig production.

Microbiological, pathological and histological findings in four Danish pig herds affected by a new neonatal diarrhoea syndrome.

BACKGROUND: Neonatal diarrhoea is a frequent clinical condition in commercial swine herds, previously regarded to be uncomplicated to treat. However, since 2008 it seems that a new neonatal diarrhoeic syndrome unresponsive to antibiotics and common management practices has emerged. Routine laboratory examinations have not detected any pathogen related to this syndrome. The primary purpose of this study was to evaluate if well-known enteric pathogens could be associated with outbreaks of neonatal diarrhoea, thus question the hypotheses of a new syndrome. Furthermore, we wanted to evaluate macroscopic and microscopic findings associated with these outbreaks and if possible propose a preliminary piglet-level case-definition on syndrome New Neonatal Porcine Diarrhoea syndrome (NNPDS). RESULTS: Four well-managed herds experiencing neonatal diarrhoea with no previously established laboratory conclusion and suspected to suffer from New Neonatal Porcine Diarrhoea Syndrome, were selected. Within these herds, 51 diarrhoeic and 50 non-diarrhoeic piglets at the age of three to seven days were necropsied and subjected to histological and microbiological examination. Faeces were non-haemorrhagic. Neither enterotoxigenic E. coli, Clostridium perfringens type A or C, Clostridium difficile, rotavirus, coronavirus, Cryptosporidium spp, Giardia spp, Cystoisospora suis nor Strongyloides ransomi were associated with diarrhoea in the investigated outbreaks. Macroscopically, the diarrhoeic piglets were characterized by filled stomachs and flaccid intestines without mucosal changes. The predominant histological lesions were villous atrophy in jejunum and ileum. Epithelial lesions in colon were seen in one third of the case piglets. CONCLUSIONS: The results of the study supported the hypothesis that a new neonatal porcine diarrhoea was present in the investigated herds, since no known pathogen(s) or management factors could explain the diarrhoeal outbreaks. Based on the findings in the four herds the following case-definition of NNPDS was suggested: Non-haemorrhagic diarrhoea during the first week of life, without detection of known infectious pathogens, characterized by milk-filled stomachs and flaccid intestines at necropsy.

Selection of method is crucial for the diagnosis of porcine circovirus type 2 associated reproductive failures.

During a 2-month period a newly repopulated Danish pig herd experienced an increase in numbers of stillborn and mummies, caused by porcine circovirus type 2 (PCV2) associated reproductive failure. Based on recordings of data over time, the progression of the clinical outbreak was studied and the diagnostic value of different techniques was evaluated. Foetal hearts (38 cases and 13 controls) were examined by immunohistochemistry (IHC) and real-time polymerase chain reaction (PCR) for the detection of PCV2; and total immunoglobulin G (IgG) was measured in pleura cavity fluid. PCV2 IHC was positive in 14/38 of the case foetuses, which were delivered during a 9 days period early in the outbreak. On the basis of the results obtained by IHC and PCR, the foetuses were divided into 3 categories: PCV2 negative; moderately positive (10(4) to 10(7) copies per 500 ng DNA); and massively positive for PCV2 (>10(7) copies per 500 ng DNA). All control- and IHC positive foetuses were included in the negative and massively positive groups, respectively. Ten case foetuses had elevated IgG levels, which did not correlate with the IHC or PCR results. Based on the clustering of the IHC positive foetuses, it is suggested that IHC only is suited for diagnosing acute stages of reproductive failure, whereas quantitative PCR can be used as a sensitive diagnostic method within a wider time span. It seems that IgG measurements are unpredictable as indication of intrauterine infection with PCV2.