Porcine proliferative enteritis: serological, microbiological and pathological studies from three field epizootics.

Three outbreaks of porcine proliferative enteritis were evaluated clinically, pathologically, microbiologically and serologically. The disease was characterized by a chronic intermittent diarrhea. Pathological lesions included a thickened, turbid ileum with the microscopic appearance of proliferating ileal crypt epithelial cells. Comma shaped intracytoplasmic organisms were observed in the apical portions of the proliferating crypt epithelial cells with a Warthin-Starry silver stain. Microbiologically, both Campylobacter sputorum subspecies mucosalis and Campylobacter hyointestinalis, were cultured from ileal specimens of seven pigs with lesions of porcine proliferative enteritis. Microagglutination antibody titers were determined on sera from 12 of 14 pigs with porcine proliferative enteritis and on sera from 91 clinically normal swine. Pigs with porcine proliferative enteritis had a low antibody titer to subspecies mucosalis that ranged from 1-3 with a mean of 2.17. A varied C. hyointestinalis titer from 3-7 with mean of 4.83 was determined. Titers to either subspecies mucosalis and C. hyointestinalis were higher in non-porcine proliferative enteritis pigs. The results indicate that the presence of a positive titer to either C. hyointestinalis or subspecies mucosalis in swine is not indicative of clinical disease. The isolation of C. hyointestinalis from diseased ileal specimens (porcine proliferative enteritis) confirms previous reports implicating this agent in the disease.     

Species-specific DNA probes for Campylobacter species isolated from pigs with proliferative enteritis

Cloned, chromosomal DNA probes from porcine isolates of Campylobacter hyointestinalis and C. mucosalis were developed for the detection and identification of these putative swine enteric pathogens. High molecular weight chromosomal DNA from each species was used to construct genomic libraries in plasmids. Recombinants were selected which hybridized strongly to the homologous organism, but not to any other species of Campylobacter. Species-specific recombinants were labeled with phosphorus-32 and tested for sensitivity by dot blot hybridization to various dilutions of DNA and bacteria from each swine species, including C. hyointestinalis, C. mucosalis, C. coli and C. jejuni. Specificity was tested by hybridizing these probes against various strains of C. hyointestinalis or C. mucosalis, and against reference strains of all other described Campylobacter species. A C. hyointestinalis-specific probe and a C. mucosalis-specific probe were identified which were capable of detecting 1 ng of DNA or 10(4) cfu by bacterial spot blotting on nylon membranes. These probes hybridized to intestinal mucosal scrapings containing C. hyointestinalis and C. mucosalis obtained from pigs with proliferative enteritis, but not to material from normal pigs. Thus, cloned, chromosomal DNA probes may be useful in the detection and identification of bacteria involved in swine proliferative enteritis.     

Experimental transmission of turkey viral hepatitis to day-old poults and identification of associated viral particles resembling picornaviruses.

Turkey viral hepatitis (TVH) was experimentally reproduced in two experiments in 1-day-old poults. In the first experiment, an infectious inoculum was prepared from filtered yolk materials harvested from dead embryonating chicken eggs (ECE) previously inoculated with suspensions of liver and pancreas tissues collected from TVH-affected birds in commercial turkey flocks. One-day-old poults given a yolk-sac inoculation or oral gavage with this preparation developed lesions in the liver and pancreas characteristic of TVH at 20 days postinoculation (PI) in 60% and 14% of the experimentally infected birds, respectively. With the identical inoculum, embryo mortality occurred at 8 and 10 days PI in embryonating turkey eggs (ETE) inoculated into the yolk sac. In the second experiment, an infectious inoculum was prepared from filtered yolk materials from dead ETE harvested in the first experiment. One-day-old poults given a yolk-sac inoculation with this filtered yolk material developed lesions in the liver and pancreas within 5 days PI. At 20 days PI, 67% of the experimentally infected birds had similar lesions. With the inoculum given to these poults, embryo mortality occurred at 6, 8, and 10 days PI in ETE inoculated into the yolk sac. Virus particles 26-28 nm in diameter with icosahedral morphology typical of picornaviruses were identified by EM in the yolk sacs of ETE that died in both experiments, and inoculated ETE that died following passage of filtered suspensions of pancreatic tissues collected from affected birds in the first experiment.     

Reproduction of proliferative enteritis in gnotobiotic pigs

Gnotobiotic pigs dosed orally with filtrates (0.8 and 0.65 micron) of intestinal mucosa from a pig affected by proliferative haemorrhagic enteropathy developed lesions of proliferative enteritis, affecting mainly the ilea. Other piglets dosed with filtrates of affected mucosa from the same source and from other proliferative haemorrhagic enteropathy or intestinal adenomatosis mucosae, did not develop lesions. All inocula contained numerous campylobacter-like organisms evident in stained smears, Campylobacter coli and C mucosalis. C coli colonised the intestines of all the pigs, C hyointestinalis (which was not detected in the inocula) did so in some affected and unaffected pigs while C mucosalis was not recovered from any of the intestines. Although other explanations are possible the number and viability of the intracellular campylobacter-like forms is likely to be the critical factor in infectivity. In affected intestines the crypts were colonised by campylobacter-like organisms, and their attachment and entry into enterocytes was associated with cellular proliferation. Immunofluorescence reactions suggested that the intracellular campylobacter-like organisms were antigenically distinct from the known Campylobacter species. It is possible, therefore, that porcine proliferative enteritis is caused by a further unidentified Campylobacter species, or that there is a marked antigenic change of C hyointestinalis or C coli on entry into porcine enterocytes.     

Possible relationship of proliferative enteritis in pigs and hamsters

Three- to six-week-old hamsters were orally inoculated with broths containing one of the following cultures: Campylobacter mucosalis; C. hyointestinalis; C. coli; C. jejuni, all of porcine proliferative enteritis origin, or else C. jejuni of hamster origin. Hamsters given the last of those organisms were shown to have colonisation of their intestines by C. jejuni and 36 of 40 developed an acute enteritis. Mild hyperplasia of enterocytes in ileal crypts was evident in one hamster 2 days after it was given C. coli. No other lesions were detected. Further 3-week-old hamsters were orally inoculated with homogenised intestinal mucosa collected from 4 pigs (A-D) affected by proliferative enteritis. Lesions of proliferative enteritis were detected in 7 of 41 hamsters necropsied 10-21 days after being dosed with mucosas B or D. Marked hyperplasia of ileal enterocytes, associated with numerous intracellular Campylobacter-like organisms, were invariably detected in experimentally affected hamsters. No particular Campylobacter sp. was consistently isolated. None of the controls had demonstrable lesions. The results suggested that cross-species transmission of proliferative enteritis was possible from pigs to hamsters. Therefore a common initiating or aetiological agent may be present. No specific organism was identified as filling this role by inoculation of hamsters with pure cultures.     

Bacteria in enteric lesions of horses

Thirty-three species of bacteria were isolated from the gastrointestinal mucosa of 23 adult horses and two foals. The bacteria isolated could be related to gross and microscopical lesions in some cases. Clostridium perfringens type A, Actinobacillus equuli, Salmonella typhimurium and Campylobacter coli biotype 1 could all be associated with gastrointestinal lesions. C jejuni biotype 1 and Aeromonas hydrophila were both recovered in this study and have been identified as causes of enteritis in horses or in other species. The case of C coli enteritis appears to be the first such report. The difficulties in examining adult horses with enteritis and relating the lesions seen to the bacteria isolated are discussed.     

Pathogenesis of Brucella abortus in chicken embryos

Chicken embryos inoculated with Brucella abortus at 6, 10, and 12 days of incubation were examined by light and electron microscopy. B. abortus was identified by avidin-biotin immunoperoxidase and immunogold techniques. Death occurred from 2 to 5 days post-inoculation, depending on age of the embryo and route of inoculation. B. abortus was recovered from all infected eggs. Brucellae had spread throughout all tissues and localized preferentially within cells of mesodermal derivation. Organ distribution and degree of bacterial replication varied with age of the embryo at time of inoculation. In 6-day-old embryos, B. abortus localized preferentially in endoderm and mesoderm of yolk sac wall, extra- and intraembryonic serosal epithelia, and glomeruli of the mesonephros. In 10- and 12-day-old embryos, B. abortus spread to all tissues; renal glomeruli, liver, spleen, and heart were most severely infected. Intracellular B. abortus was within the rough endoplasmic reticulum of mesenchymal, mesothelial, yolk endodermal, and hepatic cells. In mononuclear phagocytes, endothelial cells, and granulocytes, bacteria were within membrane-bound vacuoles. Intracellular replication of B. abortus in embryonic tissues, especially yolk endoderm, closely resembled that in experimental infections of trophoblasts.     

Plasmid profiles of six species of Campylobacter from human beings, swine, and sheep

Twenty-four isolates representing 6 species of Campylobacter were screened for plasmids. A large plasmid with an approximate molecular weight of 38 Mdal was detected in 5 C jejuni isolates originally recovered from diarrheic human beings, in one isolate of C coli recovered from diarrheic pigs, and in 1 isolate of C sputorum ssp mucosalis and 2 isolates of C hyointestinalis recovered from pigs with proliferative enteritis. One isolate of C coli and 1 isolate of C hyointestinalis contained an additional smaller plasmid with an approximate molecular weight of 1.6 Mdal; this plasmid was partially mapped by restriction endonuclease digestion. Fifteen Campylobacter isolates contained no detectable plasmids: 2 C coli, 2 C sputorum ssp mucosalis, 2 C fecalis, 1 C fetus ssp fetus, and 8 C hyointestinalis isolates. In summary, 37.5% of the Campylobacter isolates contained a 38-Mdal plasmid, with 8% having both 38 Mdal and 1.6-Mdal plasmids; 62.5% contained no detectable plasmids.     

Campylobacter sputorum subsp mucosalis and Campylobacter hyointestinalis infections in the intestine of gnotobiotic pigs

At 4 days of age, 7 gnotobiotic pigs were orally inoculated with broth cultures of both Campylobacter sputorum subsp mucosalis (CSM) and Campylobacter hyointestinalis (CH). One pig was killed and evaluated each week for 7 weeks. Forty-eight hours after inoculation, CH and CSM were recovered from the feces of the pigs; thereafter, only CH was recovered. Organisms morphologically typical of Campylobacter sp were observed on the mucosal surface and on the crypt epithelial cells of the ileum, cecum, and colon from post-inoculation weeks (PIW) 2 through 7. Bacteria were clustered around the surface opening of goblet cells in pigs at PIW 6 and 7. Crypt epithelial cell proliferation and intracellular bacteria were not seen, except in 1 pig (killed at PIW 7) in which intracellular bacteria were seen only in the cecum. Therefore, CSM and CH did not induce porcine proliferative enteritis in gnotobiotic pigs.     

Comparison of culturing Mycoplasma gallisepticum from fresh eggs and 18-day-old embryos

Twenty-four 70-week-old and sixteen 27-week-old white leghorn hens were challenged with R strain Mycoplasma gallisepticum (MG) by injection into the caudal thoracic air sac and infraorbital sinus. Eggs were collected daily and cultured within 7 days or incubated for 18 days. Vitelline membranes of eggs were cultured directly; in 18-day-old embryos, cultures were taken from the yolk sac, air sacs, and oral cavity. Culture of vitelline membrane of eggs within 2 days was compared with culture of eggs stored 10 days post oviposition. The first MG-positive egg was laid 2 days postinfection (PI). Hens continued to lay positive eggs to the end of the experiments. There was no significant difference in MG recovery between eggs cultured within 2 days and those cultured 10 days post oviposition. MG was isolated at a significantly higher rate from eggs than from 18-day-old embryos. MG was isolated at a higher rate from the yolk sac of 18-day-old embryos than from the air sacs or oral cavity of the same embryos.     

Enterocecocolitis associated with intraepithelial Campylobacter-like bacteria in rabbits (Oryctolagus cuniculus)

We examined 28 suckling, weanling, and young adult rabbits with lethargy, inappetence, and mucinous, semifluid feces. Sixteen of the rabbits had intestinal lesions. In eight of these rabbits, the primary changes were multifocal to diffuse epithelial proliferation and accumulation of lymphocytes, macrophages, or both in the lamina propria of the small intestine, cecum, and sacculated colon. In two of these rabbits, the accumulation of macrophages in the lamina propria was extensive. The other eight rabbits had erosive and suppurative cecocolitis, and four of the rabbits with proliferative lesions also had suppurative cecocolitis. In Warthin-Starry-stained sections of affected intestine, curved or spiral bacteria were visible within degenerated or hyperplastic epithelium, in luminal exudate, or in both. Such organisms were sparse or not found in the other 12 rabbits, which did not have intestinal lesions. The bacteria ultrastructurally resembled intraepithelial Campylobacter-like bacteria previously observed in proliferative enteritis in a variety of species and in acute typhlitis in young rabbits. In immunofluorescence tests, Campylobacter-like bacteria in epithelial cells, crypt lumina, and in luminal exudates in both proliferative and erosive lesions bound monoclonal antibodies and polyclonal antisera prepared against intracellular bacteria found in proliferative enteritis in pigs, hamsters, and ferrets. These observations indicate that a condition similar to proliferative enteritis of swine, hamsters, and other species also occurs in laboratory rabbits.     

The distribution of orally administered 14C-aniline HCl in tissues of dairy cattle, swine and laying pullets

Single or chronic daily (30 d) doses of 14C-aniline (an aromatic amine) were administered to dairy cattle, swine and laying pullets to determine the amount of an ingested synfuel-related chemical that would remain in consumable animal products. The 14C-residues were found in muscle, liver, other edible organs and fat, as well as in milk and eggs. The predominant site of deposition in acute- and chronic-exposed pullets was kidney, followed by internal yolk (immature ovum of ovary) and then liver. Egg yolk was the major site of 14C-radioactivity in whole eggs. Liver, then kidney, were the major sites of deposition in acutely exposed swine; when chronically exposed, only liver showed preferential deposition with other tissues surveyed yielding similar concentration values. Dairy cattle yielded tissue distribution patterns similar to chronic swine. Within 8 h after an acute exposure, 14C-residue was detected in milk which reached its maximum at 24 to 32 h. Our data demonstrate that derived concentration values of 14C-residue in tissues was dependent upon the species studied as well as on the mode of exposure, acute versus chronic. Although the concentration values were variable, sufficient amounts of 14C-aniline and (or) its metabolites were found in consumable products.     

Isolation of Campylobacter from feral swine (Sus scrofa) on the ranch associated with the 2006 Escherichia coli O157:H7 spinach outbreak investigation in California

We report the isolation of Campylobacter species from the same population of feral swine that was investigated in San Benito County, California, during the 2006 spinach-related Escherichia coli O157:H7 outbreak. This is the first survey of Campylobacter in a free-ranging feral swine population in the United States. Campylobacter species were cultured from buccal and rectal-anal swabs, colonic faeces and tonsils using a combination of selective enrichment and antibiotic-free membrane filtration methods. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS, Bruker Daltonics, Inc., Billerica, MA, USA) was used to identify species followed by confirmatory multiplex PCR or 16S rRNA sequencing. Genetic relatedness of Campylobacter jejuni and Campylobacter coli strains was determined by multilocus sequence typing (MLST) and porA allele sequencing. Altogether, 12 (40%) of 30 feral swine gastrointestinal and oral cavity specimens were positive, and six species were isolated: Campylobacter coli, Campylobacter fetus, Campylobacter hyointestinalsis, Campylobacter jejuni, Campylobacter lanienae and Campylobacter sputorum. Campylobacter jejuni subtypes were closely related to MLST sequence type 21 (ST-21) and had identical porA sequences. Campylobacter coli subtypes were unrelated to isolates in the pubMLST/porA database. This feral swine population lived in close association with a 'grassfed' beef cattle herd adjacent to spinach and other leafy green row crop fields. The findings underscore the importance of protecting raw vegetable crops from faecal contamination by wild or feral animals. The study also illustrates a potential risk of Campylobacter exposure for hunters during handling and processing of wild swine meat.     

Comparative evaluation of the effect of two maternal diets on fatty acids, vitamin E and carotenoids in the chick embryo.

1. The fatty acid profile of egg yolk and vitamin E and carotenoid accumulation in the egg yolk and embryonic tissues were investigated in relation to the maternal diet. 2. Two hundred fertile eggs (Ross 308 Broiler Breeder), obtained from hens fed on a maize-based (M-diet) or a wheat-based diet (W-diet), were incubated using standard conditions. 3. The egg yolk and embryo tissues (residual yolk, yolk sac membrane, liver, kidney, lung, muscles, adipose tissue and plasma) were collected on d 18 of incubation and on d 21 (newly-hatched chicks) and analysed for fatty acids, vitamin E and carotenoids. 4. The diets did not differ in terms of fatty acid or alpha-tocopherol concentrations. The concentration of carotenoids in the M-diet was 11.8 mg/kg and in the W-diet was 5.6 mg/kg with lutein and zeaxanthin being major carotenoids. 5. Eggs from the M-group contained higher (P<0.01) concentrations of beta+gamma-tocopherols, total carotenoids, lutein and zeaxanthin. Chickens hatched from those eggs were characterised by the increased concentrations of total carotenoids and zeaxanthin in all the tissues studied. The concentration of beta+gamma-tocopherol was enhanced only in the liver and yolk sac membrane. 6. It is concluded that the maternal diet plays an important role in antioxidant systems formation during chick embryonic development; the M-diet can increase the antioxidant potential of the egg yolk and embryonic tissues compared to the antioxidant potential provided by parent birds fed the W-diet.     

Isolation from turkey breeder hens of a reassortant H1N2 influenza virus with swine, human, and avian lineage genes

Type A influenza viruses can infect a wide range of birds and mammals, but influenza in a particular species is usually considered to be species specific. However, infection of turkeys with swine H1N1 viruses has been documented on several occasions. This report documents the isolation of an H1N2 influenza virus from a turkey breeder flock with a sudden drop in egg production. Sequence analysis of the virus showed that it was a complex reassortant virus with a mix of swine-, human-, and avian-origin influenza genes. A swine influenza virus with a similar gene complement was recently reported from pigs in Indiana. Isolation and identification of the virus required the use of nonconventional diagnostic procedures. The virus was isolated in embryonated chicken eggs by the yolk sac route of inoculation rather than by the typical chorioallantoic sac route. Interpretation of hemagglutination-inhibition test results required the use of turkey rather than chicken red blood cells, and identification of the neuraminidase subtype required the use of alternative reference sera in the neuraminidase-inhibition test. This report provides additional evidence that influenza viruses can cross species and cause a disease outbreak, and diagnosticians must be aware that the variability of influenza viruses can complicate the isolation and characterization of new isolates.     

A descriptive study of the frequency and characteristics of proliferative enteropathy in swine in Ontario by analyzing routine animal health surveillance data

Routine surveillance data, collected on pathology submissions at the Animal Health Laboratory in Guelph between 1992 and 1997, were analyzed to determine demographic, clinical, and pathologic characteristics of cases of proliferative enteropathy and the frequency of this condition relative to other infectious enteric diseases in swine in Ontario. The most commonly reported disease was Escherichia coli enteritis (average cases/year = 70.0). Among infectious enteropathies that occur typically in neonatal pigs, coccidiosis (28.4 cases/year) and rotaviral enteritis (5.6 cases/year) were reported. Among infectious enteropathies generally associated with diarrhea in weaner and grower/finisher pigs, the most frequently reported was proliferative enteropathy (27.6 cases/year), followed by swine dysentery (23.3 cases/year), transmissible gastroenteritis (19.6 cases/year), and salmonellosis (8.4 cases/year). Diarrhea and bloody diarrhea were reported in 29% and 31%, respectively, of herds diagnosed with proliferative enteropathy. Important gross intestinal lesions included mucosal hypertrophy (62% of cases), hemorrhage (47%), and mucosal necrosis (34%). Histologic intestinal lesions included epithelial hyperplasia (90% of cases), mucosal necrosis (59%), and inflammation (49%). Our results suggest that proliferative enteropathy is a major infectious enteric disease in grower/finisher pigs in Ontario.     

水貂肠炎病毒高免卵黄抗体的制备及应用

用水貂肠炎病毒免疫蛋鸡制备卵黄抗体,结果表明,经５次免疫制备的卵黄抗体,于最后一次免疫第１０天ＡＧＰ效价达１：３２,第３０天效价为１：８。该抗体较稳定。４℃保存１２０天,－２０℃保存１８０天效价不变。用该抗体治疗细小病毒肠炎病犬,治愈率达９３．９％,证实用水貂肠炎病毒制备卵黄抗体是成功的,可用于国小病毒肠炎的治疗。     

In ovo injection of branched‐chain amino acids: Embryonic development,hatchability and hatching quality of turkey poults

In this study, the influence of a branched‐chain amino acid blend (BCAA composed of 3 l ‐leucine:1 l ‐valine:2 l ‐isoleucine) injected into the amniotic fluid was evaluated for embryonic growth, yolk‐sac (YS) utilization and development of gastrointestinal tract (GIT) and skeletal muscles of turkey embryos from day 24 of incubation (24E) to hatching, together with hatchability, poult quality and liver L* (lightness), a* (redness) and b* (yellowness) values at hatch. At day 22 of incubation, embryonated eggs (n = 240) were assigned to three treatments, that is, eggs were not injected (control, NC) or injected with 1.5 ml sterile solution with 0.9% salt (SA) or 0.2% BCAA blend (BCAAb). These solutions were injected manually into the amniotic fluid of the embryonated eggs. To determine weights and lengths (where appropriate) of the studied organs and tissues, four embryonated eggs and poults per treatment were selected at 24E and at hatch. While the BCAAb decreased the YS and embryo weight, hatchability and the liver L* value, it increased the weight and quality of poults and the weights of breast and thigh muscles at hatch. In conclusion, the in ovo feeding of the BCAA blend negatively affected hatchability but positively affected hatching weight and poult quality by improving development of skeletal muscles and by regulating energy metabolism.     

Hatchability rate and embryonic growth of broiler chicks following in ovo injection royal jelly

1. The objectives of this study were to compare the hatchability, chick body and internal organs weights and plasma testosterone concentration of hatchlings after in ovo administration of royal jelly (RJ) on d 7 of incubation.

2. Fertile eggs (n = 150) were injected into the air sac or yolk sac with 0.5 ml normal saline solution or normal saline and pure RJ. The eggs were randomly divided into 5 groups of 30 eggs each: NC, control eggs receiving no injection; ASA, air sac–injected eggs given normal saline solution; ARJ, air sac–injected eggs injected with pure RJ; YSA, yolk sac–injected eggs receiving normal saline solution and YRJ, yolk sac–injected eggs given pure RJ.

3. Injection of RJ significantly decreased hatchability (46.7%) compared with injection of saline solution (68.3%). Hatchability was lower in ARJ (33.3 %) and YRJ (60.0%) groups than in the NC group (90.0%). Hatchability in ASA (70.0%) and YSA (66.66%) groups were comparable to the NC group.

4. In ovo injection of RJ into both sacs increased chicks’ absolute and relative body, heart, liver and testes weights compared to the control group whereas plasma testosterone concentration was similar among the different groups.

5. It was concluded that in ovo injection of RJ may be an effective method to increase the body weight of hatched chicks as an absolute value (CWT) and chicks' internal organ weights.     

Dynamics of yolk sac resorption and post-hatching development of the gastrointestinal tract in chickens, ducks and geese

Under similar environmental conditions, 100 male Shaver-Starbro hybrid chickens, 100 male Mulard cross-line ducklings and 100 male Landaise goslings kept in 30 metabolic cages from the first day of life (10 birds of each species separately per cage) were chosen for the study. The animals were fed semi ad libitum diets with crude protein content and energy density similar to that used for poultry species, avoiding feed residues. During the first 21 days of life they received starter diets and from day 22 to 42 grower diets. Maize accounted for 23-40% and barley constituted 10-18%. Wheat accounted for 20.0% in starter diets and 10-18% in grower diets. Resorption of yolk sac residues, performance, development of the intestinal tract in young chickens, ducklings and goslings were assessed. During the first 5 days of life intensive absorption of yolk sac ingredients was observed. On day 7, residues of the yolk sac were found in more than 30% of chickens compared with approximately 10% in geese. In ducklings residues of the yolk sac were not found. On day 16 unabsorbed yolk sacs were found in approximately 10% of chickens. Liver and pancreas weighed the highest in geese and in ducks; in chickens the weight of these organs was significantly lower (p < 0.01). The relative values calculated to 100 g of metabolic body weight show that the average small intestine in chickens was significantly longer (p < 0.01) than in ducks or geese. The intestinal tract developed earlier in the chicken than in the water fowl.