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Abstract. The sensitivity and specificity of an immunoperoxidase monolayer assay (IPMA) was evaluated in a blind serologic study of a group of disease-free pigs and a group of pigs experimentally infected with intestinal homogenate containing Lawsonia intracellularis organisms. Sixty pigs from the control group were kept in the source farm, and another 60 animals were transferred to an isolation unit and challenged intragastrically. All animals were bled before and 21 days after challenge. Fecal samples were collected on the same dates. The IPMA results were tested for sensitivity and specificity in a 2 ϫ 2 table using the challenged and nonchallenged status as gold standard. Sensitivity and specificity were evaluated for different cutoff points (serum dilutions). Specificities of 100% were obtained for all the serum dilutions tested (1:15, 1:30, 1:60, and 1:120). The sensitivity levels were 90.7%, 88.9%, 81.5%, and 75.9% for the serum dilutions 1:15, 1:30, 1:60, and 1:120, respectively. The sensitivity of the dilution 1:15 was slightly, but not significantly, higher than the dilution currently used as the cutoff point (1:30). Cross-reactivity of the IPMA test was evaluated using sera from pigs experimentally inoculated with Brachyspira pilosicoli and various Campylobacter species. All these samples were negative. Sera samples from 3 porcine proliferative enteropathy known negative populations, 40 growing pigs from 2 commercial farms and a group of 6 cesarean-derived and colostrum-deprived pigs, also tested negative by IPMA. The IPMA serologic test with the cutoff point of 1:30 showed specificity of 100% and sensitivity close to 90% and, therefore, is an appropriate diagnostic test for herd screening but not for diagnosing PPE on the individual level.Porcine proliferative enteropathy (PPE) is an intestinal infectious disease caused by the obligate intracellular bacterium, Lawsonia intracellularis. 9,10 The epidemiological status of the swine population worldwide regarding PPE is poorly known. Two of the antemortem methods currently available for diagnosis of PPE are polymerase chain reaction (PCR) of fecal samples 4 and serology. 5 Previous studies 1,5 have shown that the indirect fluorescent antibody (IFA) serology test is much more sensitive than PCR in fecal samples for detecting experimentally infected pigs. Sensitivities were 90%-91% versus 39%-67% for serology and PCR in fecal samples, respectively. The specificity of PCR in fecal samples is virtually 100%. 4 There was no cross-reactivity of hyperimmune sera of pigs experimentally infected with different enteropathogenic bacteria in the IFA test. 5 But there is no information about the specificity of the IFA serology test.Recent data have shown that the immunoperoxidase monolayer assay (IPMA) has similar sensitivity to the currently used serologic test, the indirect immunofluorescence test, for detecting PPE-affected animals. 2 The IPMA test has the advantages of not requiring the use of a fluorescent microscope, being easier to interpret, and having stabl...

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Abstract. Proliferative enteropathy is an intestinal infectious disease caused by the obligate intracellular bacterium Lawsonia intracellularis. Immunohistochemistry staining has superior sensitivity over hematoxylin and eosin and silver staining for detecting L. intracellularis in histological sections. A L. intracellularis-specific monoclonal antibody (MAb) produced in the UK (IG4 MAb) has been described in the literature. However, no monoclonal or polyclonal antibodies are commercially available. Therefore, the objective of this study was to produce and characterize new polyclonal and monoclonal antibodies against L. intracellularis that are suitable for diagnostic use. Proliferative enteropathy (PE) is an intestinal infectious disease characterized by thickening of the distal small and proximal large intestinal mucosa due to enterocyte proliferation associated with the presence of an intracellular bacterium, Lawsonia intracellularis. 23 The 2 major clinical aspects of PE in pigs are acute hemorrhagic diarrhea and sudden death of replacement animals and of finishing pigs close to market age, known as proliferative hemorrhagic enteropathy (PHE), and chronic mild diarrhea and reduced performance in grow-finishing pigs, known as porcine intestinal adenomatosis. 17,24 In addition to these 2 different clinical presentations in pigs, PE has also been reported in several other animal species, including the rat, hamster, guinea pig, rabbit, ferret, ostrich, pig, deer, horse, 3 emu, 21 and monkey. 15 Diagnosis of PE in veterinary diagnostic laboratories is based on detection of histologic lesions using the hematoxylin and eosin stain and Warthin Starry silver stain and by polymerase chain reaction (PCR) in fecal and intestinal samples. An immunohistochemical (IHC) stain using a monoclonal antibody against L. intracellularis 23 detected positive antigen label in tissues from several animal species, including whitetailed deer, 6 a dog, 20 an emu, 21 hamsters, 23 horses, 9,29 a From the Department of Veterinary Pathobiology, University of Minnesota, Saint Paul, MN, 55108 (Guedes, Gebhart). monkey, 15 pigs, 25,26 and rabbits. 28 In addition, the sensitivity and specificity of IHC staining using the monoclonal antibody against L. intracellularis 23 was found to be superior to that of hematoxylin and eosin and Warthin Starry silver stains in pigs with macroscopic lesions suggestive of PE. 12,13 However, this monoclonal antibody 23 is not commercially available to diagnostic laboratories. Therefore, the objective of this study was to produce and to characterize new polyclonal and monoclonal antibodies against L. intracellularis. Materials and methodsBacterial strains and culture conditions. Six different L. intracellularis isolates, Bilophila wadsworthia (L. intracellularis' closest phylogenetic organism), 27 Brachyspira hyodysenteriae, Salmonella choleraesuis, S. typhimurium, and Escherichia coli K88 were used in this study (Table 1). Lawsonia intracellularis isolates were obtained from a collection held at the University o...

The currently used indirect fluorescent antibody test (IFAT) for the detection of antibodies against porcine proliferative enteropathy (PPE) was compared to an immunoperoxidase monolayer assay (IPMA). Serum samples used in this comparison were collected from 5-week-old pigs on day 0 (pre-experimental challenge) and on days 7, 14, 21, and 28 after oral inoculation with intestinal homogenate from pigs affected by PPE (28 challenged pigs) and sucrose phosphate glutamate solution (2 control pigs). All animals were euthanized 4 weeks after inoculation. Immunohistochemistry staining was applied to formalin-fixed, paraffin-embedded sections of ileum for the detection of Lawsonia intracellularis antigen. The serology results with each method agreed in all samples, except on days 0 and 7 in 1 control animal, which was positive by IPMA, but negative by IFAT. The percentage of agreement between IFAT and IPMA was 98.6%.

Clostridium difficile has emerged as a major cause of neonatal colitis in piglets, displacing classic bacterial pathogens. However, there is no information regarding the distribution of this microorganism in pig farms in

BackgroundInfluenza A viruses circulating in pigs in Brazil are still not characterized, and only limited data are available about swine influenza epidemiology in the country. Therefore, we characterized the hemagglutinin (HA) and neuraminidase (NA) genes of influenza viruses isolated from Brazilian pigs. We also evaluated one case of probable swine‐to‐human transmission.MethodsTwenty influenza viruses isolated from pigs during 2009–2010 in five Brazilian states (Minas Gerais, Sao Paulo, Parana, Rio Grande do Sul, and Mato Grosso) were used. One human isolate, from a technician who became ill after visiting a swineherd going through a respiratory disease outbreak, was also used in the study. Phylogenetic analysis for the HA and NA genes and hemagglutinin amino acid sequence alignment were performed.ResultsAll isolates clustered with pandemic H1N1 2009 (pH1N1) viruses and appeared to have a common ancestor. Genetic diversity was higher in the HA than in the NA gene, and the amino acid substitution S203T in one of HA's antigenic sites was found in most of the samples. The human isolate was more related to swine isolates from the same herd visited by the technician than to other human isolates, suggesting swine‐to‐human transmission.ConclusionOur results show that pH1N1 was disseminated and the predominant subtype in Brazilian pigs in 2009–2010.

The effect of an oral treatment with the tartrate salt of tylvalosin on the development of proliferative enteropathy in 60 experimentally challenged pigs was studied. Thirty of the pigs were fed a diet medicated with 50 ppm tylvalosin and 30 were fed the unmedicated diet. The treated animals started to receive the medicated feed the day before they were inoculated, and continued to receive it for 14 days. The pigs' bodyweight, feed consumption and clinical signs were evaluated, and they were examined postmortem 20 days after inoculation, and samples of ileum were collected for immunohistochemistry (IHC) for Lawsonia intracellularis. Clinical signs of the disease were more evident in the untreated group than in the treated group. The average daily weight gain, average daily feed consumption and feed conversion efficiency were better in the treated group. The combined length of intestine with lesions was 2847 cm in the untreated group and 183 cm in the treated group. The tylvalosin treatment significantly reduced the level of L intracellularis infection; almost half of the treated pigs were IHC-negative compared with 3.3 per cent of the untreated pigs.