Clinical and pathologic effects of oral administration of transmissible gastroenteritis vaccine to gnotobiotic pigs.

Pigs from 3 litters kept under gnotobiotic conditions were inoculated orally with virulent transmissible gastroenteritis (TGE) virus, a TGE vaccine, or Hank's balanced salt solution at 2 days of age and then euthanatized at intervals ranging from 1 to 7 days after inoculation. Pigs exposed to the vaccine had clinical evidence of diarrhea and weakness. Lesions resembling those of TGE were revealed grossly, microscopically, and by scanning electron microscopy. Viral antigen was seen in intestinal epithelial cells by the direct fluorescent antibody technique. The disease induced by the vaccine virus had a longer incubation period and lesions were less severe than that induced by the virulent virus.

Physiological relationships between microbiological status and dietary copper levels in the pig.

Ten germ-free pigs and 10 conventionally reared pigs were fed one of two nutritionally balanced diets containing either 16 ppm Cu (basal) or 283 ppm Cu (high-Cu) to evaluate the physiological relationships between Cu and microbiological environment. Germ-free pigs tended to have higher ADG and average daily feed intake (ADFI) than conventionally reared pigs. Feeding the high-Cu diet tended to reduce ADG and ADFI in germ-free pigs but it increased ADG and ADFI in conventionally reared pigs. Hemoglobin and hematocrit were higher in germ-free pigs than in conventionally reared pigs (P less than .001), and hematocrit (P less than .01) and erythrocyte count (P less than .06) were reduced by feeding the high-Cu diet. Germ-free pigs had lower total leukocyte count (P less than .01) and the relative percentages of differentiated leukocytes were altered compared with conventionally reared pigs. Feeding the high-Cu diet increased the percentage of band neutrophils and monocytes in germ-free pigs but reduced the percentage of these cells in conventionally reared pigs (P less than .04). Germ-free pigs had higher concentrations of Cu and Zn in liver and plasma (P less than .001) and greater plasma ceruloplasmin oxidase activity (P less than .001) than conventionally reared pigs did. The high-Cu diet increased liver Cu and Zn (P less than .001) and plasma Cu (P less than .001) and reduced liver and plasma Fe (P less than .05). Organ weights (g/kg BW) differed between germ-free and conventionally reared pigs, and feeding the high-Cu diet reduced thymus weights (P less than .002). Intestinal weight and thickness were reduced in germ-free pigs, and feeding the high-Cu diet generally reduced villus height and width and crypt depth in germ-free pigs, whereas it increased these measurements in conventionally reared pigs.

Effects of dietary change and rotavirus infection on small intestinal structure and function in gnotobiotic piglets.

The combined effects of weaning and rotavirus infection on small intestinal structure and function and on growth rate were studied in 28 gnotobiotic piglets. There was little damage by rotavirus to the proximal small intestine, some damage to the mid small intestine and relatively severe damage to the distal small intestine; villi were stunted, crypts lengthened and activities of all brush border enzymes decreased. The damage was short-lived despite the synchronisation of rotavirus infection with simulated weaning. There was no evidence of persistent damage to the small intestine and growth rate was unaffected.

Prevalence of K88, K99, and 987P pili of Escherichia coli in neonatal pigs with enteric colibacillosis.

One hundred nineteen live neonatal pigs with diarrhea less than or equal to 2 weeks old were euthanatized, and frozen sections of their ilea were submitted to an indirect fluorescent antibody technique to identify K88, K99, and 987P pili (also referred to as F-4, F-5, and F-6 pili, respectively) in Escherichia coli. Ten-centimeter ileal sections were used to determine numbers of lactose-fermenting bacteria. Of 52 pigs in which E coli pili were found, 14 had K88 (27%), 23 had K99 (44%), 13 had 987P (25%), and 2 had K88 and K99 simultaneously (4%). Numbers of lactose-fermenting bacteria were significantly (P less than or equal to 0.05) higher in pigs with piliated E coli than in pigs without piliated E coli. Results of this study indicated that piliated E coli are a major cause of enteric disease in neonatal swine in Michigan, and that in pigs less than or equal to 2 weeks of age, K99 was the most frequently encountered pilus antigen.

Adhesion of K99-positive Escherichia coli to intestinal brush borders of pigs.

Ileal samples from 242 pigs, collected at 3 Michigan slaughterhouses, were studied to determine the prevalence of intestinal receptors for K99-positive Escherichia coli. A brush border adhesion test was used to identify the receptors. Of the 242 samples examined, receptors were demonstrated in 230 (95%). After storage of brush border preparations at 4 C, bacterial aggregates lacking identifiable brush border fragments were present in samples tested for adhesion, indicating that K99 receptors may be released from brush border membranes. Seemingly, most, if not all, pigs have intestinal receptors for K99 pili, and an inheritance pattern similar to that observed for K88 receptors probably does not exist for K99 receptors.

Iron requirements of baby pigs reared in germ-free or conventional environments on a condensed milk diet.

Baby pigs were maintained in either a germ-free or a conventional environment and fed a sterile condensed milk diet (10 micrograms Fe/g milk solids) supplemented with 0, 50 or 100 micrograms Fe/g solids from FeSO4.7H2O or given an im injection of 100 mg Fe from Fe dextran. Pigs were reared on these diets for 4 wk, and weekly measures of gain, food consumption and hematology were taken. The pigs were then killed, and organ weights were taken and tissues analyzed for Fe concentration. All conventional pigs not given supplemental Fe died. None of the unsupplemented pigs raised in the germ-free environmental died. Conventional pigs gained faster and more efficiently and became anemic more rapidly. However, conventional and germ-free pigs were similar in the efficiency with which they incorporated supplemental Fe into hemoglobin or stored Fe in liver and spleen. We concluded from this study that both germ-free and conventional pigs fed a condensed milk diet require 50 to 100 micrograms Fe/g milk solids or more than 100 mg of Fe from Fe dextran once parenterally to maintain desirable hematological traits for 4 wk.

Immune responses of the bovine fetus and neonate to Escherichia coli: plaque-forming and intestinal immune responses.

The immune responses of 26 Angus-Hereford fetuses and neonates to Escherichia coli O26:K60:NM were studied after bacterin or saline solution was injected (in utero) into the amniotic fluid. Calves were euthanatized at birth or were orally revaccinated; some were challenge exposed with live organisms. The hemolytic plaque assay was used to determine the presence of cells producing immunoglobulins M, G1, and G2 (IgM, IgG1, and IgG2) in 4 segments of the small intestine, mesenteric lymph nodes, and spleen. The passive hemagglutinin activity of intestinal washings was also determined. Anti-O26 passive hemagglutinin activity in the intestinal washings of principal calves was greater than in that of control calves, but in a given segment of the small intestine, usually this activity was relatively small and less consistent than the plaque-forming response. Greater numbers of plaque-forming cells were observed in the small intestine of 14 of the 15 principal calves when compared with the control calves tested.

Lesions of experimentally induced colibacillosis in neonatal gnotobiotic pigs: a scanning electron microscopic study.

Scanning electron and light microscopy were used in studies of stomach, duodenum, cranial, and caudal portions of the jejunum, ileum, cecum, and spiral colon from 18 gnotobiotic pigs. Five pigs were raised as controls and 13 were exposed at 6 days of age by oral administration of 1.6 X 10(6) colony-forming units of Escherichia coli O138:K81:NM. Infiltration of leukocytes into the mucosa of the stomach seen with the light microscope has not been previously reported. The irregular pattern of the mucosal surface of the stomach formed by the gastric pits and the mucosal extensions on the individual rugae revealed with scanning electron microscopy was different than anticipated. Sections of the ileum from control and infected pigs contained collapsed cells around the extrusion zone at the tips of the villi. These collapsed cells were more numerous in infected pigs and appeared to have sloughed from the area of the extrusion zone resulting in exposure of the lamina propria. Cecum and spinal colon were free of changes. Alterations of the mucosa of the intestinal tract of gnotobiotic pigs infected with E coli as visualized by scanning electron microscopy were considered too inconsistent to be of diagnostic significance.

Immune responses of the bovine fetus and neonate to Escherichia coli: quantitation and qualitation of the humoral immune response.

The humoral immune responses of fetuses and neonates of Escherichia coli O:26:K60:NM were studied in 26 Angus-Hereford crossbred calves. Bacterin (5.0 X 10(10) organism) was injected in utero directly into the amniotic fluid of seventeen 7- to 8.5-month-old fetuses (principals). Saline solution was injected in the same manner into 9 control fetuses. Colostrum-deprived neonates were allotted to 10 groups and either were euthanatized at birth or were subjected to oral revaccination, challenge inoculation with the homologous organism, or both. The resistance to challenge exposure was a function of previous in utero injection of bacterin, age when challenged, and dose of challenge organisms used. Control calves were susceptible to only a large challenge dose, whereas almost all of the prinicipal calves were resistant. Revaccination of principal calves with bacterin at birth, exposure to the large challenge dose, or both, caused a marked increase in anti-O26 passive hemagglutination titers. Results of quantitative and qualitative radioimunossay indicated that the immune response to the O26 antigen was mainly of the immunoglobulin M (IgM) class, although there were also demonstrable changes in immunoglobulins (Ig) G1 and G2. The actively acquired immune responses were serotype specific, and there was no cross reactivity with 4 other E coli serotypes. An unidentified immunoprecipitate band was observed in immunoelectrophoretograms of whole bovine serum which may represent another class of Ig or which may be a subclass of IgG1 or IgG2.

Enteric colibacillosis: evaluation of strains of Escherichia coli utilizing the ligated loop technique in gnotobiotic swine.

The enteropathogenicity of 6 strains of Escherichia coli was evaluated via the ligated loop technique in 15 gnotobiotic swine 3 to 4 weeks old. Strain enteropathogenicity in gnotobiotic swine, as determined on the basis of visual loop distention and light microscopic evaluation of intestinal sections, compared favorably with that reported in conventional swine, except that jejunal loops of gnotobiotic swine may be somewhat less sensitive to certain strains. In addition, postoperative mortality rate among gnotobiotic pigs was greater than that reported in conventional pigs. Two types of aberrant loop reactions were observed during the experiment. The 1st, and the more commonly encountered type, was due to the leakage of enteropathogenic organisms or enterotoxin from one loop to another; this was considered to be a true false-positive reaction. The 2nd type of unexpected positive reaction occurred in 2 sacs inoculated with the nonenteropathogenic strain 115. This latter reaction may have represented either a reversion of the organism to its previous enteropathogenic state or a unique reaction of the gnotobiotic intestine to this strain. False-positive loops were not identified in any other sacs, whether inoculated or not. Results of these experiments support the hypothesis that false-positive loops originate from naturally occurring infection.

Experimental colibacillosis in gnotobiotic baby pigs. I. Microbiological and clinical aspects.

Sixty-two gnotobiotic pigs were used in three experiments to determine the means whereby two related strains of Escherichia coli colonized the intestinal tract. Pigs were exposed by a method simulating neonatal contamination of the umbilical stump. Bacteremia was produced within one and one half hours, and by 24 hours the infection was generally well established in the gastrointestinal tract. By 48 hours after exposure, the bacteremia had subsided so that only an occasional isolation from organs other than the gastrointestinal tract was made. Oral exposure of one litter of germfree pigs produced heavy colonization of the entire gastrointestinal tract within four hours. Evidence of intermittent bacteremia was present in pigs of this litter. Diarrhea appeared earlier when pigs were exposed orally than when they were exposed by way of the umbilical stump.

Experimental colibacillosis in gnotobiotic body pigs. II. Pathology.

Sixty-two neonatal gnotobiotic pigs were used in three experiments to determine the lesions produced by two closely related strains of Escherichia coli O138:K81:NM (of Michigan origin) and O138:K81 (of Minnesota origin). Exposure was by subcutaneous injection of bacterial culture into the umbilical stump or by oral inoculation. Gross signs common to monocontaminated pigs included distention of the flaccid small and large intestines with fluid contents. Edema was prominent in various tissues of most pigs exposed via the umbilical stump but not in those exposed orally. Histological lesions were predominantly in the gastrointestinal tract and were variable. At one extreme acute hemorrhagic enteritis was present in two pigs, while at the other extreme in a few pigs it was difficult to distinguish tissues of infected pigs from those of noninfected germfree pigs. Significant histological lesions common to monocontaminates included mild inflammatory reaction, hydropic degeneration of the intestinal epithelium, evidence of interference with normal function of the villus-draining mechanisms, and vascular changes generally indicated by edema. The findings suggest that interference with normal absorption of nutrients plays at least some role in the pathogenesis of colibacillosis in young gnotobiotic pigs.

Polyserositis and arthritis due to Escherichia coli in gnotobiotic pigs.

Forty gnotobiotic pigs from six litters were exposed orally to Escherichia coli 083:K.:NM at 69 to 148 hours of age, while 17 pigs from the same litters served as unexposed controls. Clinical signs of infection included fever, anorexia, diarrhea, lameness, and reluctance to move.Eighty-four percent of the exposed pigs in four litters died, while only 13% in two litters died. Gross and microscopic lesions included serofibrinous to fibrinopurulent polyserositis in 96% of the exposed pigs in four litters and 33% of the exposed pigs in two litters. A few pigs had gross and/or microscopic lesions of arthritis. Escherichia coli was routinely isolated from the serous and synovial cavities of infected pigs.Anti-hog cholera serum administered orally as a colostrum substitute gave partial protection against E. coli infection.

Comparison of body weights, organ weights and histological features of selected organs of gnotobiotic, conventional and isolator-reared contaminated pigs.

Twenty-seven pigs from three litters were used in a comparison of body weights, organ weights, and selected histological features of germfree, conventional and isolator-reared contaminated pigs. At three weeks of age conventional pigs were heavier than pigs of the other two groups. The mandibular lymphnodes, stomachs, and small intestines of contaminated pigs were significantly heavier than the same organs of germfree pigs. This difference was not found in superficial inguinal or prefemoral lymph nodes. Other statistically significant organ weight differences were found.Histologically, the lymph nodes of conventional and contaminated pigs were much more active than those of germfree pigs, although secondary nodules were occasionally found in lymph nodes of germfree pigs. Greater quantities of iron-containing pigment were found in the spleens of germfree pigs than in spleens of the other two groups. Hepatic interlobular septa were somewhat more developed in conventional pigs than in germfree or contaminated pigs.