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Swine enteric diseases have caused significant economic loss and have been considered as the major threat to the global swine industry. Several coronaviruses, including transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV), have been identified as the causative agents of these diseases. Effective measures to control these diseases are lacking. The major host cells of transmissible gastroenteritis virus and porcine epidemic diarrhea virus have thought to be epithelial cells on small intestine villi. Aminopeptidase-N (APN) has been described as the putative receptor for entry of transmissible gastroenteritis virus and porcine epidemic diarrhea virus into cells in vitro. Recently, Whitworth et al. have reported that APN knockout pigs are resistant to TGEV but not PEDV after weaning. However, it remains unclear if APN-null neonatal pigs are protected from TGEV. Here we report the generation of APN-null pigs by using CRISPR/Cas9 technology followed by somatic cell nuclear transfer. APN-null pigs are produced with normal pregnancy rate and viability, indicating lack of APN is not embryonic lethal. After viral challenge, APN-null neonatal piglets are resistant to highly virulent transmissible gastroenteritis virus. Histopathological analyses indicate APN-null pigs exhibit normal small intestine villi, while wildtype pigs show typical lesions in small intestines. Immunochemistry analyses confirm that no transmissible gastroenteritis virus antigen is detected in target tissues in APN-null piglets. However, upon porcine epidemic diarrhea virus challenge, APN-null pigs are still susceptible with 100% mortality. Collectively, this report provides a viable tool for producing animals with enhanced resistance to TGEV and clarifies that APN is dispensable for the PEDV infection in pigs.

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We measured the effect of pharmacological doses of glucocorticoid on piglet jejunal structure and function during acute viral diarrhea. Weaned piglets, infected experimentally with transmissible gastroenteritis virus, a coronavirus that induces a diarrheal illness similar to human rotavirus infection, received methylprednisolone (30 mg/kg) or saline intramuscularly at 48 and 72 h after infection; noninfected littermate controls were similarly injected with methylprednisolone. Animals were killed at 96 h, at the height of diarrhea, and jejunal epithelium was studied in vitro. Transmissible gastroenteritis, as expected, induced structural, enzyme, and Na transport abnormalities. Methylprednisolone did not affect small intestinal structure or function of noninfected control piglets. In transmissible gastroenteritis-infected piglets, jejunal villi were longer and glucose-facilitated Na absorption was greater after methylprednisolone than after saline treatment. Increased glucose stimulation of Na flux in vitro in the methylprednisolone-treated infected group was not attributable to enhanced Na+-K+-ATPase activity and occurred despite persistence of the virus within mucosal cells, shown by immunofluorescence microscopy. In this piglet model of viral diarrhea, early regeneration of absorptive surface that precedes recovery of disaccharidase function is accelerated by glucocorticoid therapy.

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Intestinal phospholipase B activity in pigs inoculated with transmissible gastroenteritis (TGE) virus was studied. Phospholipase activity was quantified by measuring the hydrolytic release of free fatty acids in homogenized intestinal tissue incubated with lysophosphatidylcholine. An increase in enzyme activity was observed in the cranial and caudal portions of the ileum and jejunum in pigs killed 3, 6, and 8 days after inoculation with TGE virus. Seemingly, phospholipase B may be part of the host immune response against TGE viral infection.

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To investigate the effect of chronic protein-calorie malnutrition on intestinal repair after an enteric infection, we examined small intestinal structure, enzyme activity, and sodium transport in undernourished piglets during the acute and convalescent phases of a viral enteritis, transmissible gastroenteritis (TGE). Gnotobiotic pigs, nutritionally deprived from the age of 7 days, gained less weight than dietary controls from 14 days of age until the end of the study. Animals from malnourished and control diet groups were inoculated with TGE virus at 22-23 days and studied during the acute (40 h) and convalescent (4, 10, and 15 days) stages of this experimental enteritis along with noninfected dietary controls. After TGE infection, we observed a further decrease in weight gain and an increased mortality only in undernourished pigs. In jejunum and ileum of both dietary groups at 40 h after TGE infection, we observed comparable structural lesions, similar decreased activities of mucosal enzymes (sucrase, lactase, sodium-potassium-dependent ATPase), and increased thymidine kinase activities. Also we noted comparable diminution of glucose-stimulated jejunal sodium absorption in both dietary groups at 40 h. In control diet pigs, transport abnormalities recovered by 4 days after TGE infection and normal mucosal structure and enzyme activity returned over 4-15 days. In undernourished piglets, structural repair and enzyme abnormalities were prolonged when compared with the control diet group; glucose-stimulated sodium transport did not recover until 10 days after infection and never regained the enhanced activity seen in noninfected undernourished controls.(ABSTRACT TRUNCATED AT 250 WORDS)

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The aim of this work was to find out the histochemical activity of alkaline and acid phosphatases, and nonspecific esterases in intestinal mucosa in spontaneous swine colibacillosis. The investigated animals were divided into 3 groups: I--oedema disease (12 pigs), II--enetrotoxemia due to E. coli (8 pigs), III--control animals (5 pigs). The above grouping was based on the clinical, bacteriological and post-mortem examination. A slight increase of alkaline phosphatase activity in the jejunum and ileum of sick pigs was observed. The acid phosphatase activity decreased in the duodenum of oedema diseased pigs, while it increased in the cecum and colon--comparing with the control and enterotoxemia groups of animals. The decrease of acid phosphatase activity was observed in the jejunum and ileum of the enterotoxemia group of animals. The difference in the activity of nonspecific esterases concerned only the cecum and colon of diseased pigs, where its decrease was observed.

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Thirty seven piglets with transmissible gastroenteritis (TGE) and 41 with other enteric diseases were examined for evidence of villous atrophy and reduced lactase activity. Widespread villous atrophy appeared indicative of TGE whereas normal lactase activity tended to exclude this possibility. However, both tests are far from specific and neither is recommended to practitioners as an aid to clinical diagnosis.

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