Observation of Helicobacter-like organisms in gastric mucosa of grey foxes (Urocyon cinereoargenteus) and bobcats (Lynx rufus)

Archival specimens of gastric mucosa of 10 raccoons (Procyon lotor), 9 porcupines (Erethizon dorsatum), 6 grey foxes (Urocyon cinereoargenteus), 6 bobcats (Lynx rufus), 4 skunks (Mephitis mephitis), and 3 black bears (Ursus americanus) were microscopically examined for evidence of Helicobacter-like organisms. Such organisms were seen in the specimens from the grey foxes and bobcats only. Histochemical stains (modified Steiner and carbol fuchsin methods) revealed long spiral organisms within lumina of gastric glands; however, neither gross nor microscopic lesions were observed. By electron microscopy (EM), the organisms were found to be free in the glandular lumina and were seen occasionally in the cytoplasm of gastric epithelial cells. Morphologically, 2 different phenotypes of spiral organisms were identified by EM. The organisms associated with bobcats appeared to be more tightly coiled than those seen in grey foxes. The presence of Helicobacter-like organisms in the gastric mucosa of grey foxes has not previously been described.     

Intestinal spirochetosis in wild sika deer (Cervus nippon yesoensis) infected with Brachyspira species

Seven adult free-ranging sika deer (Cervus nippon yesoensis) were examined by histology, immunohistochemistry and electron microscopy for intestinal spirochetal infection. Histologically epithelial and goblet cell hyperplasia and edema of the lamina propria mucosa with macrophage and lymphocyte infiltration were observed in the cecum and colon in 6 of the 7 deer. Numerous argyrophilic spirochetes were present in the crypts and some had invaded epithelial and goblet cells and caused degeneration. Immunohistochemically the organisms stained positively with polyclonal antisera against Brachyspira (Serpulina) hyodysenteriae and B. pilosicoli. Ultrastructurally they were 6-14 microm long, 0.2-0.3 microm wide and had 4-6 coils and 13 axial filaments per cell; such features were closely similar to those in the Brachyspira species. These results showed that the spirochetes were capable of inducing enteritis in deer and this intestinal spirochete infection might already be prevalent among wild sika deer in Japan. There is a possibility that this spirochetal colitis is a new syndrome in sika deer and that the same and/or similar spirochetes have infected ruminants, including sika deer and cattle.     

Brachyspira pilosicoli isolated from pigs in Japan

Two of four weak beta-hemolytic isolates of intestinal spirochetes isolated from pigs in Japan possessed a unique base alignment of TTTTTT on the 16S ribosomal DNA of Brachyspira pilosicoli and were identified as B. pilosicoli. The other two isolates were not identified by this technique. The identified isolates were 4.2 to 11 microm in length and 0.2 to 0.3 microm in diameter, 4 periplasmic flagella at each end were observed dominantly. The isolates were hippurate positive but indole negative. This is the first report on the isolation of B. pilosicoli from pigs in Japan.     

Necrotizing typhlocolitis associated with a spirochete in rheas (Rhea americana).

Necrotizing typhlocolitis was diagnosed in 13 juvenile common rheas (Rhea americana) from three separate of geographically isolated Ohio flocks, with mortality ranging from 25% to 80%. At postmortem examination, a diphtheritic membrane covered ulcerated cecal mucosa. Histologically, cecal sections showed necrosis and granulomatous-to-suppurative inflammation that extended into the submucosa and often surrounded large eosinophilic colonies of bacteria. Warthin-Starry staining showed these colonies to be composed of entangled spirochetes that invaded the submucosa and frequently were present transmurally. Similar organisms were identified by Warthin-Starry staining in the cecum of a juvenile rhea from a fourth flock that histologically had mild lymphocytic typhlitis. Scanning and transmission electron microscopy demonstrated the presence of a spirochete in the ceca. Anaerobic culture yielded a gram-negative, beta-hemolytic spirochete. Coccidia, histomonads, and Salmonella spp. were consistently absent.     

The prevalence of interstitial nephritis and leptospirosis in 283 raccoons (Procyon lotor) from 5 different sites in the United States.

A retrospective histopathological study was carried out on tissues of 283 raccoons from 5 different geographical locations for presence of interstitial nephritis and renal leptospirosis. Results of this study indicate that although interstitial nephritis was common in raccoons from all locations, the presence of renal leptospiral spirochetes was not.     

Fatal Gastrointestinal Infection with ‘Flexispira rappini’‐Like Organisms in a Cat

A 4‐month‐old male British Blue cat with catarrhal to haemorrhagic enteritis showed massive colonization of the stomach, small intestine and caecum with spiral‐shaped bacilli. In the stomach, organisms were located in foveolae and gland lumina and within unaltered and degenerate epithelial cells. Inflammatory infiltration was moderate and T cell dominated. In the intestine, bacilli were found in the gut lumen, between villi, in crypt lumina and within epithelial cells. Degeneration of crypt epithelial cells as well as crypt dilation and moderate to massive macrophage‐dominated infiltration of the mucosa and submucosa were observed. Immunohistochemically, bacilli were positive with an antibody against Helicobacter. Ultrastructurally, the organisms strongly resembled ‘Flexispira rappini’, a spiral‐shaped Helicobacter species known as a normal intestinal colonizer in dogs and mice.     

Microvascular anatomy of the pig eye: scanning electron microscopy of vascular corrosion casts

The microvasculature of the eye of 10 pigs was investigated using scanning electron micrographs of corrosion casts. The ciliary body, iris and bulbar conjunctiva were supplied by the iridociliary ring artery via the long posterior ciliary artery. Capillaries of the ciliary process were of large diameter (23.2-27.5 microm) with an irregular bore, forming a thoroughfare channel draining blood in the ciliary arterioles into the pars plana venous vessels. Arterioles and venules in the iris exhibited a zigzag or spiral features. The third palpebra was supplied by the anterior ciliary artery. The capillary bed of the third palpebra was dense and was formed by many rows of fine hair-pin loops. Capillaries in the bulbar conjunctiva formed a sparse network disposing approximately parallel to the epithelium and formed a well-developed venous plexus, draining into the vortex veins. Retinal arterioles formed a slender and long course to capillaries. Retinal capillaries were extremely thin (3.0-4.0 microm in diameter). The choroid was supplied by the short posterior ciliary arteries. Choroidal arterioles exhibited a thick and short course to the choriocapillaris. The choriocapillaris was flat and sinusoid-like (8.9-13.9 microm in diameter), forming a dense sheet-like network. Blood from the choroid emptied into the episcleral vein via the vortex vein. Blood from the retina was drained by the posterior ciliary veins. The functional significance of this vascular architecture was discussed.     

吸血蓖子硬蜱成虫唾腺莱姆病螺旋体重度感染的发现

为了解莱姆病螺旋体在蜱体内的发育生物学，采用直接免疫荧光试验、银染色法组织学及电子显微镜观察螺旋体，结果显示，感染莱姆病螺旋体的饥饿蓖子硬蜱雌虫未吸血与吸血0．5和1天唾腺中无螺旋体。在吸血开始后2天出现唾腺的轻度感染，3天出现唾腺的中度感染，4天和5天出现重度感染。这些结果表明，莱姆病螺旋体媒介蜱唾腺的感染程度随吸血时间的延长而迅速增高，并可在缓慢吸血期的中后期形成重度感染。莱姆病螺旋体在吸血期媒介唾腺内的大量存在，特别是重度感染的发现，为该病原螺旋体由蟀向脊椎动物宿主传播的唾液分泌途径假设提供了有力证据。本研究对莱姆病螺旋体在吸血蟀唾腺内分裂增殖的可能性进行了讨论。     

Association of cecal spirochetes with pasty vents and dirty eggshells in layers.

Feces-stained eggshells, diarrhea, and typhlitis were identified in two commercial laying flocks in Ohio. Hens with diarrhea had spirochetes in cecal lumina and crypts. On culture, the spirochetes were motile and non-hemolytic, did not produce indole, had 12 to 15 axial filaments, were 9.2 to 11.7 microns in length and 240 to 370 nm in diameter, and had a wavelength of 5.1 to 6.5 microns on transmission electron microscopy.     

Prevalence and tissue distribution of Besnoitia darlingi cysts in the Virginia opossum (Didelphis virginiana) in Michigan

Specimens of Virginia opossums (Didelphis virginiana) in Michigan were examined over 1 year to document the presence of Besnoitia darlingi cysts. Cyst morphology, prevalence, seasonal variation, and tissue sites of isolation were studied. Histology and ultrastructural features of the detected cysts and bradyzoites were consistent with B. darlingi. In the opossums, B. darlingi had intracellular tissue cysts. Tissue cysts had a mean diameter of 560 microm and were separated from the host tissue by a thick (5-20 microm) cyst wall. Overall prevalence of B. darlingi cysts in opossums was 10.9% (15/137). Variations in the prevalence were detected during spring (3/17; 17.6%), summer (10/34; 29.4%), and fall (2/60; 3.3%). No cysts were detected in the specimens examined during winter (0/26; 0%). Numerous B. darlingi cysts were detected in ears, conjunctiva, tongue, abdominal muscles, diaphragm, stomach, heart, liver, kidney, lung, and spleen. Cysts were detected mainly in adult female opossums that were debilitated. Ear was the most frequent organ from which the cysts were reported (10/15; 66.7%) when compared individually with other body tissues (P<0.05).     

Neuroaxonal dystrophy in raccoons (Procyon lotor) from Iowa.

During a 12-month period (1998-1999), microscopic evidence of neuroaxonal dystrophy (NAD) in medullae oblongata of raccoons (Procyon lotor) was observed in 17/39 (47% prevalence in adults) from Iowa, USA. Three of the animals were kits (<3 months), 26 were between 1 and 2 years, and 10 were over 7 years. Lesions were not seen in the medullae of the 3 kits. In young adults, the lesions were mild and were seen in 7 animals. More severe lesions were present in the 10 older raccoons. Grossly, the brains were unremarkable. Microscopically, NAD was confined to the dorsal caudal medulla, where certain nuclei (predominantly gracilis and cuneate) were bilaterally affected. Severely affected animals had vacuolar degeneration of neurons or neuronal loss and extensive areas of spongiosis. Tests for the presence of PrP(res) in the brain were negative. Spongiotic areas often contained axonal spheroids. Degenerate neurons and axons occasionally contained amphophilic periodic acid-Schiff-positive granular material. There was a paucity of inflammatory cells in the affected areas. Since lesions were not present in kits, were either absent or mild in young adults, and were severe in older raccoons, the findings may be related to advancing age. Neuroaxonal dystrophy has not been previously reported in raccoons. Retrospective examination of raccoon brains from the eastern and northwestern areas of the country revealed very low prevalence of NAD. Because of the apparently high prevalence of this condition at this geographic location, factors other than age (genetic, nutritional, and/or environmental) may influence this degenerative process in the brains of raccoons in Iowa.     

Gastroduodenal adenocarcinomas and rectal adenoma in a cougar (Felis concolor) infected with Helicobacter-like organisms and spirochetes

A 14-year-old female cougar died from gastroduodenal adenocarcinomas and rectal adenoma. At necropsy, polypoid tumor masses of various sizes were scattered on the mucosal surfaces of the stomach, duodenum, and rectum. Histologically, the gastric tumor was diagnosed as an intestinal type adenocarcinoma and the tumor cells metastasized to the mesenteric lymph nodes, spleen, and lung. Helicobacter-like organisms were detected in the lumina lined by foveolar epithelium. In the duodenum, the carcinoma cells were localized in the limina propria and many of them were intensely positive for proliferating cell nuclear antigen (PCNA). In contrast, the rectal adenoma had a lower number of PCNA-positive cells. In the rectum, chronic inflammation with numerous spirochetes was also noted. These results indicated that the occurrence of the gastrointestinal tumors might be associated with the bacterial infection described above.     

Raccoons (Procyon lotor), but not rodents, are natural and experimental hosts for an ehrlichial organism related to "Candidatus Neoehrlichia mikurensis"

"Candidatus Neoehrlichia mikurensis" has been reported from a variety of rodent and Ixodes tick species in Europe and Asia. Recently, an ehrlichial organism closely related to "Candidatus Neoehrlichia mikurensis" was cultured from a raccoon (Procyon lotor) from Georgia, USA. To determine prevalence and distribution, we conducted a molecular survey of free-ranging raccoons (n=197) from 10 populations in 3 states and found that infections were common in tick-infested populations (50-94%). In an effort to determine the host range of this organism, 10 species of rodents (n=137) trapped in 3 areas where positive raccoons had been detected were tested; all were negative. In addition, captive bred raccoons and several common laboratory animals (mice, rats, and rabbits) were inoculated with the raccoon ehrlichial isolate (strain RAC413). Raccoons became infected with the culture isolate but all other hosts were refractory to infection. The 16S rRNA gene sequence (1379bp) of the RAC413 isolate was most similar (98.4-98.8%) to members of the "Candidatus Neoehrlichia mikurensis" group and phylogenetic analysis confirmed this organism was related to, but distinct from, "Candidatus Neoehrlichia mikurensis". Based on the molecular and natural history uniqueness of this organism from raccoons, we propose that this represents a novel species in the "Candidatus Neoehrlichia" group of ehrlichial organisms.     

An ultrastructural study of adenocarcinoma of the small intestine in sheep

The ultrastructural appearance at primary and metastatic sites of ten ovine small intestinal adenocarcinomas was that of scirrhous tubular adenocarcinoma. Polygonal undifferentiated tumor cells had desmosomes, folded nuclei, and moderate numbers of mitochondria but few other organelles. More differentiated cells were columnar with apical microvilli and basal nuclei. They contained granular endoplasmic reticulum, Golgi and secretory granules. Microvillus-lined intracytoplasmic lumina (5-10 micron diameter), fibrous filaments (10 nm diameter, up to 1.4 micron length) and tubular paracrystalline arrays (hexagonal symmetry, 37-nm periodicity) in lumina and secretory granules were seen in some tumor cells in all ten sheep.     

Control of terrestrial animal rabies in Anne Arundel County, Maryland, after oral vaccination of raccoons (1998-2007)

Objective-To evaluate the effectiveness of an oral rabies vaccination (ORV) project conducted from 1998 through 2007 in Anne Arundel County, Md, for the control of rabies in terrestrial animals. Design-Retrospective analysis of surveillance data (1997 through 2007). Animals-Free-ranging raccoons (Procyon lotor) and other terrestrial mammals. Procedures-Vaccinia-rabies glycoprotein recombinant virus oral rabies vaccine-bait units were distributed annually by aircraft and ground teams targeting free-ranging raccoons. Approximately 2 to 4 weeks following the vaccine-bait placement, raccoons were live trapped, sedated, processed, and then released. Serologic samples were tested for the presence of rabies virus-neutralizing antibodies (RVNAs). Bait acceptance was estimated by analysis of tetracycline biomarking of sampled teeth. Rabies incidence was determined by the passive identification of rabid terrestrial animals. Results-The incidence of rabies in terrestrial animals decreased 92% between 1997 (the year prior to the start of the ORV project) and 2007. The mean RVNA prevalence across all years was 33% among trapped raccoons in areas baited with a fish meal polymer bait type, whereas the mean bait acceptance was 30%. Adult raccoons had a seropositivity rate twice that of juvenile raccoons, whereas the bait acceptance rate between adults and juveniles did not differ significantly. For areas baited with a coated sachet bait, adults and juveniles had the same seroprevalence. Juveniles had better seroprevalence when the annual campaign started in September and October, compared with August. Conclusions and Clinical Relevance-The ORV project contributed to a significant decrease in annual incidence of terrestrial animal rabies in Anne Arundel County, Md, during the 10-year project period. For fish meal polymer baits, juvenile raccoons accessed bait at the same rate as adult raccoons but had a significantly lower prevalence of RVNAs. For coated sachet baits, seroprevalence was the same in both age groups. The time of year the bait distribution occurred and the bait type used may be partial explanations for the difference in RVNA seroprevalence between adults and juvenile raccoons.     

Characterization of the attachment of Treponema hyodysenteriae to Henle intestinal epithelial cells in vitro

Properties of the attachment of Treponema hyodysenteriae to Henle intestinal epithelial (HIE 407) cells were examined. The frequency of attachment depended on the motility and viability of the spirochetes. Rabbit hyperimmune and swine convalescent antisera inhibited attachment. Treatment of HIE cells with neuraminidase had no effect on attachment; however, treatment of spirochetes with the enzyme decreased adherence significantly (P = 0.01). Attachment was inhibited by N-acetylneuraminic acid, D-glucuronic acid, and fetuin. Adherence was increased following coincubation with N-acetylglucosamine or yeast mannan. Surface antigens of T hyodysenteriae, isolated by chemical extraction, competitively inhibited adherence. Concentrated T hyodysenteriae culture supernatant fractions inhibited adherence, but concentrated phosphate buffered-saline washings of the spirochete and concentrated uninoculated media did not inhibit adherence. Sialic acid was detected in unwashed T hyodysenteriae and spent culture supernatant fractions in higher concentrations than from washed spirochetes and uninoculated media. It was concluded that the binding adhesins on T hyodysenteriae for cultured HIE cells may contain sialic acid residues.     

Role of the raccoon in the transmission of pseudorabies: a field and laboratory investigation

Serum samples were collected from 42 raccoons trapped in a pseudorabies enzootic area of Missouri. All samples were negative for neutralizing antibodies to pseudorabies virus (PRV). Raccoons were orally exposed with one of four dose levels of PRV. The raccoons were found to be susceptible to moderately large doses of PRV tissue culture infective dose (10(4) - 10(5) TCID50). All raccoons given 10(3) TCID50 of PRV survived and did not develop clinical signs. The PRV was consistently isolated from tonsillar swabs collected from raccoons that died after exposure. The PRV was also isolated from the brain, tonsils, lungs, and salivary glands of raccoons that died. It was isolated from samples of the brain and tonsils collected from a raccoon 5 days after death. Neutralizing antibodies to PRV could not be detected in the serum of raccoons that survived PRV exposure. Tonsillar swabs and tissue samples collected from these raccoons were free of PRV. The results indicated that the raccoon may serve as a short-term reservoir for PRV, but it is unlikely to have an ipizootiologic role as a long-term, subclinical carrier of the virus.     

Morphological and quantitative study of spirochetes in the feces of normal and infected SPF pigs during the incubation period of swine dysentery.

Spirochetes found in feces of normal and infected pigs during the eight days preceding the onset of the clinical signs of swine dysentery have been studied using electron and phase contrast microscopy. According to their dimensions, diameter, length and pitch, three morphological groups have been described: small, intermediate and large spirochetes. On the basis of their axial filaments arrangement, eight types have been observed. Compared to the control pigs, there was no increase in the total number of spirochetes in infected pigs, except at day 6 before the onset of clinical signs. However, infected pigs have shown a wide spectrum of different morphological types of spirochetes in comparison to control, uninfected pigs, which showed only the type "1-2-1" spirochete of axial filaments arrangement. This last type was predominant in infected swine during the entire incubation period as compared to other spirochete types described.     

A field investigation of subclinical mastitis in sheep in southern England

The prevalence, aetiology and epidemiological features of subclinical mastitis were investigated in 358 lowland ewes in seven flocks in southern England. Milk samples (2092) were collected at 3-weekly intervals; those which were both bacteriologically and Whiteside test positive were deemed to have originated from glands with subclinical mastitis. The period prevalence of subclinical mastitis was 11.7% and the prevalence remained relatively constant over the course of lactation (5.5-7.0%). The predominant bacterial isolates from 48 glands with subclinical mastitis were streptococci (42%), coagulase-negative staphylococci (33%), Pasteurella haemolytica (17%) and Staphylococcus aureus (8%). Coagulase-negative staphylococci were the predominant isolates (53%) from samples which did not show a positive Whiteside test result. The prevalence of subclinical mastitis increased with age of ewe but was not influenced by the presence of teat lesions. There was a significant association between the development of clinical mastitis (26 glands) and antecedent subclinical mastitis caused by the same organism (10 glands).     

Prevalence and Spatial Distribution of Salmonella Infections in the Pennsylvania Raccoon (Procyon lotor)

A study was conducted to determine the prevalence and spatial distribution of Salmonella infection in Pennsylvania raccoons (Procyon lotor), common wildlife mammals known to occupy overlapping habitats with humans and domestic food animals. The Pennsylvania Game Commission provided a total of 371 raccoon intestinal samples from trapped and road‐killed raccoons collected between May and November 2011. Salmonella was isolated from the faeces of 56 (15.1%) of 371 raccoons in 35 (54%) of 65 counties across Pennsylvania. The five most frequently isolated serotypes were Newport (28.6%), Enteritidis (19.6%), Typhimurium (10.7%), Braenderup (8.9%) and Bareilly (7.1%). Pulsed‐field gel electrophoresis (PFGE) analysis of the Salmonella isolates and subsequent comparison to the Pennsylvania Department of Health human Salmonella PFGE database revealed 16 different pulsetypes in Salmonella isolates recovered from raccoons that were indistinguishable from pulsetypes of Salmonella collected from clinically ill humans during the study period. The pulsetypes of seven raccoon Salmonella isolates matched those of 56 human Salmonella isolates by month and geographical region of sample collection. Results from Clustered Regularly Interspaced Short Palindromic Repeats and Multi‐Virulence Locus Sequence Typing (CRISPR‐MVLST) analysis corroborated the PFGE and serotyping data. The findings of this study show that several PFGE pulsetypes of Salmonella were shared between humans and raccoons in Pennsylvania, indicating that raccoons and humans might share the same source of Salmonella.