Lenn R. Harrison

Proliferative enteropathy in a foal caused by Lawsonia intracellularis-like bacterium

Proliferative enteropathy (proliferative enteritis, proliferative ileitis, intestinal adenomatosis) has been reported in several animal species including the pig,1 dog, foal, blue fox, guinea pig, ferret, hamster, and rabbit. The disease is characterized by adenomatous hyperplasia of crypt epithelial cells in the ileum and colon with intracytoplasmic curved bacteria resembling Campylobacter species. In the single case previously reported in a foal, Campylobacter-like organisms were demonstrated within the cytoplasm of enterocytes by spirochete stains and electron microscopy. In a study using cloned DNA probes to isolated Campylobacterlike organisms, there was failure of the probes to hybridize with common porcine Campylobacter species, suggesting the causative agent to be an unidentified or uncultured species. This finding, along with DNA sequencing and structural characteristics, resulted in this organism being named ileal symbiont intracellularis. Subsequently, the organism was described and classified as a new genus and species, Lawsonia intracellularis. 1 An unweaned 5-month-old mixed-breed female foal with a history of anorexia, lethargy, and profuse watery diarrhea of greater than 1-week duration was presented to the University of Kentucky, Livestock Disease Diagnostic Center, for necropsy. Treatment had not been attempted. At necropsy, the foal was thin with readily apparent skeletal muscle atrophy. Gross lesions were confined to the small intestine. There was irregular thickening of the jejunum and ileum with the ileum being more severely affected. Lesions in the midjejunum were multifocal in nature and consisted of areas of discoid mucosal thickening, whereas the distal jejunum and ileum contained diffuse mucosal thickening resulting in a rugose pattern (Fig. 1). Samples of the small intestine, stomach, cecum, colon, brain, heart, lung, kidney, and spleen were placed in 10% neutral buffered formalin and, following fixation, were embedded in paraffin, sectioned at 5 μm, and stained with hematoxalin and eosin. In addition, selected sections of small intestine were stained by the Warthin-Starry silver impregnation method for bacteria. Pieces of formalinfixed small intestine were postfixed in osmium tetroxide and embedded in epoxy resin. Thin sections were stained with uranyl acetate and lead citrate. Histopathologically, the affected mucosa was thickened and consisted of hyperplastic glandular structures lined by immature epithelial cells (Fig. 2). The hyperplastic epithelium resulted in distortion of the normal villous structure. Some

Prevalence of latent, neuropathogenic equine herpesvirus-1 in the Thoroughbred broodmare population of central Kentucky

REASON FOR PERFORMING STUDY An emerging problem of equine herpesvirus-1 (EHV-1) infection in horses in the USA is a high-mortality myeloencephalopathy that commonly occurs where large numbers of horses are stabled. EHV-1 isolates recovered from recent neurological outbreaks represent a mutant virus strain that possesses enhanced neuropathogenicity. A central question of EHV-1 myeloencephalopathy is the latency carriage rate for these mutants of EHV-1 in USA horse populations. OBJECTIVE To estimate the prevalence of neuropathogenic strains of EHV-1 as latent infections in the Thoroughbred broodmare population of central Kentucky. METHODS Submandibular lymph nodes (SMLN) were collected during post mortem examination of 132 Thoroughbred broodmares. Total DNA purified from SMLN tissue was tested for the presence of latent EHV-1 DNA by an ultrasensitive magnetic bead-based, sequence-capture, nested PCR method. Differentiation of active from latent infections by EHV-1 was achieved by detection of transcripts of EHV-1 glycoprotein B by reverse transcription PCR. RESULTS Latent EHV-1 DNA was detected in the SMLN tissues of 71 (54%) of the 132 mares submitted for necropsy. Thirteen (18%) of the 71 latently infected horses harboured the neuropathogenic biovar of EHV-1. Of the 13 horses latently infected with an ORF30 mutant strain of EHV-1, 11 also carried a latent, wild-type strain of the virus in their SMLN tissues. CONCLUSIONS Neuropathogenic strains of EHV-1 have established a significant presence in the Thoroughbred broodmare population of central Kentucky as latently infected carrier horses. The data also indicate that a highly sensitive DNA detection method is required to identify many instances of EHV-1 latency. POTENTIAL RELEVANCE The presence of a relatively large biological reservoir of latent, neuropathogenic EHV-1 has the potential for posing emerging equine health and economic threats to the future prosperity of the USA horse industry.

Fatal Disease in Nursing Puppies Associated with Minute Virus of Canines

Thirteen cases of a previously undescribed parvoviral infection affecting puppies ranging in age from 5 to 21 days is described. The cases were originally thought to represent an unusual pathologic manifestation of canine parvovirus-2 (CPV-2) infection. However, failure to confirm CPV-2 infection in any of the cases suggested a different parvovirus was involved. Minute virus of canines (MVC) was subsequently isolated from a case by using the Walter Reed Canine Cell Line, the only cell line which will support the growth of MVC. The pathologic and virologic findings for these 13 cases are described in this report.

REVIEW paper: Mare reproductive loss syndrome

An epidemic of early fetal loss (EFL), late fetal loss (LFL), fibrinous pericarditis, and unilateral uveitis which occurred during the spring of 2001, are together now known as the mare reproductive loss syndrome (MRLS). A similar epidemic with less intensity was reported during the same period of time from southern Ohio, West Virginia, and Tennessee. The same syndrome with lesser intensity recurred in 2002. The estimated economic loss from the syndrome in 2001 and 2002 together was approximately

Microbiologic and pathologic findings in an epidemic of equine pericarditis

500 million. Both EFL and LFL were characterized by the absence of specific clinical signs in aborting mares. Nonhemolytic Streptococcus spp. and Actinobacillus spp. accounted for 65% of the organisms isolated from fetuses submitted for a postmortem during the MRLS period in 2001 and 2002. The pathologic findings in fetoplacental units of LFL included bronchopneumonia and funisitis, and there were no findings in EFL. Epidemiologic studies conducted in 2001 suggested an association between the presences of eastern tent caterpillars (ETC) in pastures with MRLS. Experimental studies in pregnant mares by exposure to ETC, or administration by stomach tube or with feed material, reproduced EFL and LFL. Similar experimental studies in mouse, rats, and goats with ETC were unsuccessful. Currently, 2 hypotheses are proposed for MRLS. One hypothesis proposes that an ETC-related toxin with secondary opportunistic bacterial invasion of the fetus leads to MRLS. The second hypothesis suggests that a breach of gastrointestinal mucosal integrity by hairs of ETC leads to a bacteremia and results in MRLS. In 2004, a similar equine abortion storm was reported from Australia and caterpillar exposure was identified as a risk factor for the abortion. In 2006, the syndrome was observed in Florida and New Jersey.

Transitional cell carcinoma of the urinary bladder in a Thoroughbred, with intra-abdominal dissemination.

During the spring and summer of 2001 and in association with the mare reproductive loss syndrome, 22 terminal and 12 clinical cases of equine pericarditis were diagnosed in central Kentucky. Actinobacillus species were the principal isolates from 8 of 10 nontreated, terminally affected and 3 of 10 clinically affected horses. Enterococcus faecalis and Streptococcus zooepidemicus were cultured from the remaining 2 nontreated terminal cases. No viruses were isolated in tissue culture. Nucleic acid of equine herpesvirus-2 was detected in pericardial and tracheal wash fluids of 3 and 1 individuals, respectively. Microscopic alterations in sections of heart and parietal pericardium were consistent with chronic fibrinous bacterial pericarditis. This report confirms a significant role of Actinobacillus species in equine pericarditis and describes an epidemic of this infrequently observed syndrome in the horse.

Rhodococcus equi and genetic susceptibility: assessing transferrin genotypes from paraffin-embedded tissues

A 14-year-old Thoroughbred gelding with a history of acute onset of hematuria was presented for necropsy. Transitional cell carcinoma of the urinary bladder with intra-abdominal dissemination was diagnosed. Tumor masses were observed on the splenic capsule and surrounding the distal abdominal aorta. Tumor cells showed diffuse cytoplasmic reactivity for cytokeratin but were negative for epithelial membrane antigen, carcinoembryonic antigen, tumor-associated glycoprotein 72, and vimentin.

Septicemia and Peritonitis Due to Actinobacillus equuli Infection in an Adult Horse

Rhodococcus equi is a bacterial pathogen, ubiquitous in the soil, that infects many foals and is lethal to some. Transferrin is an iron-binding protein that has bacteriostatic properties in the blood. Transferrin is also highly polymorphic in most species, with 15 variants identified for horses using biochemical methods, and may be responsible for variation in susceptibility to bacterial pathogens. The objectives of this study were 1) to compare biochemical typing with DNA typing of transferrin, 2) to determine if transferrin DNA from archival paraffin-embedded tissue samples can be recovered and typed with molecular methods, and 3) to determine if there was an association between foal death caused by R. equi and transferrin type. Comparing biochemical methods and DNA sequencing for 41 horses demonstrated correspondence between the typing methods. The allele frequency of archival paraffin-embedded tissue samples from 34 Thoroughbred foals that succumbed to R. equi showed an excess of the F allele and a deficiency of the D allele (P < 0.05). Year of collection (P > 0.8), age of foal (P > 0.3), and sex of foal (P > 0.6) were not statistically associated with transferrin type. The archival material was successfully transferrin typed using DNA sequencing, and there may be an association between foal death caused by R. equi and transferrin type.

Equine abortion and premature birth associated with Cellulosimicrobium cellulans infection

Actinobacillus equuli is a rare cause of peritonitis in adult horses. Septicemia and peritonitis due to A. equuli were diagnosed at necropsy in an 8-year-old Saddlebred mare. The origin of the infection was not known; however, small necrotic colonic mucosal lesions presumed to have been caused by phenylbutazone treatment may have allowed bacterial invasion. A good response to antimicrobial treatment has been documented in the small numbers of previously reported acute cases of peritonitis. Because it is potentially treatable, it is important for pathologists and clinicians to identify horses with A. equuli peritonitis.

Funisitis Associated with Leptospiral Abortion in an Equine Placenta

During the 2002 and 2003 foaling seasons, Cellulosimicrobium (Cellumonas) cellulans (formerly Oerskovia xanthineolytica) was the principal microorganism isolated from fetal tissues or placentas from cases of equine abortion, premature birth, and term pregnancies. Significant pathologic findings included chronic placentitis and pyogranulomatous pneumonia. In addition, microscopic and macroscopic alterations in the allantochorion from 4 of 7 cases of placentitis were similar to those caused by Crossiella equi and other nocardioform bacteria. This report confirms a causative role of C. cellulans infection in equine abortion.