Infection of juvenile falcons (Falco spp.) with intestinal Lawsonia intracellularis.

Intestinal infection of many host species with Lawsonia intracellularis are widely reported. Analyses of infections among carnivorous falcons have not previously been reported. Fifty juvenile captive falcons (Falco spp.) with or without Lawsonia infection were investigated in the United Arab Emirates, including clinical laboratory methods. Fresh intestinal biopsy samples were analysed by microbiological techniques for Lawsonia and other bacteria and by standard parasitological and pathological methods. Lawsonia intracellularis infection was diagnosed by microbiological examination and qPCR in 10 of 50 juvenile falcons at case examination. Seven of these 10 falcons were of normal clinical appearance, and the other three had other contributing factors to ill-thrift. A range of other conditions were noted in 40 case control falcons. This first report of Lawsonia infection in falcons suggests that the agent may have a limited contribution to clinical disease in these birds, including ill-thrift syndromes. This lack of clinical disease association mimics that noted among Lawsonia infections recorded in other avian families.

Analysis of acidified feed components containing African swine fever virus.

Mitigation of African swine fever (ASF) virus in contaminated feed materials would assist control activities. Various finely-ground pig feed ingredients (5 cereals, 4 plant proteins, 2 animal proteins, 1 oil, 1 compound) were sprayed and mixed thoroughly with a buffered formic acid formulation (0, 1 or 2% vol/vol) to produce a consistent and durable level of formate (1% or 2%) with consistent acidification of cereal ingredients to less than pH 4. No such acidification was noted in other ingredients. Selected representative feed ingredients were further mixed with infectious ASF virus (106 TCID50) or media alone and incubated for 0, 6, 12, 24, 48, 72 or 168 h. The residual ASF virus at each timepoint was quantified using qPCR and a cell culture based TCID50 assay to determine survivability. Maize, rice bran and compound feed (with or without formate) all reduced infectious ASF virus to levels below the detection threshold of the cell culture assay (101.3 TCID50/mL). A consistent reduction in ASF virus DNA levels was observed by qPCR assay when maize containing ASF virus was mixed with 1% or 2% buffered formic acid. This reduction in viral DNA corresponded to the acidifying pH effect measured. No such reduction in ASF virus DNA levels was noted in non-cereal ingredients containing ASF virus, in which the pH had not been lowered below pH 4 following treatment. Interestingly, residual ASF virus levels in spiked meat/bone meal were greater than control levels, suggesting a buffering effect of that feed ingredient.

Rotavirus group C infections in neonatal and grower pigs in Australia.

BACKGROUND: Rotavirus infections of neonatal and older pigs are widely reported. Analysis of rotavirus group C prevalence and diversity has not previously been reported for Australian pig farms. METHODS: Twenty-seven farms with or without diarrhoea present among neonatal or older pigs were enrolled across eastern Australia. Fresh faecal samples were analysed by ELISA for rotavirus and RNA extractions by polyacrylamide gel electrophoresis and RT-PCR for rotavirus. Rotavirus group C samples were genotyped via sequencing. RESULTS AND CONCLUSIONS: Rotavirus infection was diagnosed in pigs on 10 of 19 farms investigated for neonatal diarrhoea, four with group A and six with group C; also among post-weaned (5- to 11-week-old) diarrhoeic pigs on two farms. Neonatal rotavirus group C infections were exclusively noted in piglets less than 1-week-old, consisting of farm infections with a single VP7 genotype (G5 or G6). Infections in post-weaned pigs were associated with multiple VP7 genotypes (G1, G3). This first report of rotavirus group C infections of Australian pigs suggests they may form a limited population of VP7 genotypes.

Muscle origin of porcine satellite cells affects in vitro differentiation potential.

Post-natal muscle regeneration relies on the activation of tissue stem cells known as satellite cells, to repair damage following exercise trauma and disease. Satellite cells from individual muscles are known to be heterogeneous with regard to proliferation, fusion and transplantation abilities, although the muscle origin has rarely been considered pertinent to their differentiation capabilities. In this study we compared the potential of two functionally distinct skeletal muscle satellite cell populations from porcine diaphragm and hind-limb semi-membranosus muscles. These two muscles were chosen primarily for differences in metabolic and contractile properties: the diaphragm is more continuously active and has a greater oxidative capacity. Cells were induced to differentiate towards myogenic and adipogenic lineages, and here we have shown that cells from diaphragm exhibit a significantly greater degree of myogenesis compared with those from semi-membranosus, while the converse was true for adipogenesis. Unexpectedly, both conditions generated small numbers of cells with neuronal characteristics for both muscle types, although more so in cells derived from the diaphragm. With increased interest in muscle adiposity with age and disease, these findings suggest that muscle origin of satellite cells does affect lineage fate, however whether differences in developmental origin or metabolic activity of the parent tissue govern this, remains to be determined.

Microarray analysis of differential expression of cell cycle and cell differentiation genes in cells infected with Lawsonia intracellularis.

Infection of intestinal crypt epithelial cells by the obligate intracellular bacterium Lawsonia intracellularis is directly linked to marked proliferation of the infected enterocytes within 3-5days post-infection. The virulence factor for this unique host cell-proliferative response is not known, but is considered to involve altered crypt cell cycle or differentiation events. McCoy mouse fibroblast cells were infected with L. intracellularis, and then harvested for expressed mRNA at daily time points, with matching non-infected control cell cultures. Mouse DNA microarray (>44,000 transcript targets) analysis of cDNA derived from matching mRNA samples showed over 40 identifiable genes with at least 4-fold changes between days 0 and 3 after infection with L. intracellularis. These included altered transcription of typical host cell 'alarm' response genes, such as interferon-related response genes Isgf3g and Igtp, known to be associated with invading microbial agents. Altered transcription of several genes in these cells known to be active in regulation of the cell cycle or cell differentiation genes, including usp18, Hr, Elavl2 and Slfn2, were also detected. The altered transcription of several of these genes via RT-PCR analysis was confirmed. The microarray-detected altered transcription of cell cycle and cell differentiation genes is of possible interest for links to Lawsonia-related disturbances in epithelial cell differentiation within the intestinal crypt, but this would need to be confirmed in intestinal epithelial cell studies.

Evaluation of porcine ileum models of enterocyte infection by Lawsonia intracellularis.

The early interaction of Lawsonia intracellularis with host cells was examined with the use of porcine ileum models. Two conventional swine were anesthetized, and ligated ileum loops were prepared during abdominal surgery. The loops were inoculated with 108 L. intracellularis or saline. After 60 min, samples of each loop were processed for routine histologic and electron microscopic study. Histologic and ultrathin sections of all the loops appeared normal, with no apposition of bacteria and host cells or bacterial entry events in any loop. Portions of ileum from a single gnotobiotic piglet were introduced as xenografts into the subcutis of each flank of 5 weaned mice with severe combined immunodeficiency disease. After 4 wk, 108 L. intracellularis were inoculated into each of 4 viable xenografts with a sterile needle; the other 3 viable xenografts received saline. Histologic and ultrathin sections of all the xenografts 3 wk after inoculation showed relatively normal porcine intestinal architecture, with normal crypts, crypt cell differentiation, and low villous structures; the xenografts treated with the bacteria also showed intracytoplasmic L. intracellularis within crypt and villous epithelial cells. Thus, entry of L. intracellularis into target epithelial cells and multiplication may not be sufficient alone to directly cause cell proliferation. A proliferative response may require active division of crypt cells and differentiation in conjunction with L. intracellularis growth.

Induction of castration by immunization of male dogs with recombinant gonadotropin-releasing hormone (GnRH)-canine distemper virus (CDV) T helper cell epitope p35.

Immunocastration is a considerable alternative to a surgical castration method especially in male animal species for alleviating unwanted male behaviors and characteristics. Induction of high titer of antibody specific for gonadotropin-releasing hormone (GnRH) correlates with the regression of testes. Fusion proteins composed of canine GnRH and T helper (Th) cell epitope p35 originated from canine distemper virus (CDV) F protein and goat rotavirus VP6 protein were produced in E. coli. When these fusion proteins were injected to male dogs which were previously immunized with CDV vaccine, the fusion protein of GnRH-CDV Th cell epitope p35 induced much higher antibody than that of GnRH-rotavirus VP6 protein or GnRH alone. The degeneration of spermatogenesis was also verified in the male dogs immunized with the fusion protein of GnRH-CDV Th cell epitope p35. These results indicate that canine GnRH conjugated to CDV Th cell epitope p35 acted as a strong immunogen and the antibody to GnRH specifically neutralized GnRH in the testes. This study also implies a potential application of GnRH-based vaccines for immunocastration of male pets.

Evaluation of protective immunity in pigs following oral administration of an avirulent live vaccine of Lawsonia intracellularis.

OBJECTIVE: To evaluate the efficacy of an orally administered avirulent live vaccine to protect pigs against challenge exposure with virulent Lawsonia intracellularis. ANIMALS: 108 weaned 3-week-old pigs (35 in experiment 1 and 73 in experiment 2). PROCEDURE: 2 experiments were conducted. On day 0, vaccinates were orally administered vaccine via drench or in drinking water, whereas challenge-control pigs were administered cultured medium. On day 21, pigs were challenge exposed with a virulent heterologous isolate of L. intracellularis. Clinical observations, weights, seroconversion, and fecal excretion of L. intracellularis were measured until day 42. At study termination, pigs were euthanatized and examined for L. intracellularis-specific lesion development of the ileum and colon. RESULTS: Pigs receiving a single dose of vaccine were protected when challenge exposed with virulent L. intracellularis (at least 10(77) TCID50/dose). In experiment 1, vaccinates had significantly less fecal excretion (47% and 40% for days 35 and 42, respectively), compared with challenge-control pigs. In experiment 2, vaccinates had significantly less fecal excretion (50% and 58% for days 35 and 42, respectively), compared with challenge-control pigs. Significant reductions in lesion development were evident in the ileum of vaccinated pigs (70% and 56% at day 42 for experiments 1 and 2, respectively), compared with challenge-control pigs. CONCLUSIONS AND CLINICAL RELEVANCE: Oral administration by drench or via drinking water of an avirulent live vaccine against L. intracellularis resulted in substantial protection against proliferative enteropathy among vaccinates and offers a better way to reduce stress of pigs during vaccine administration.

Search for Lawsonia intracellularis and Bilophila wadsworthia in malabsorption-diseased chickens.

Proliferative enteropathy is an important enteric disease caused by Lawsonia intracellularis. A wide range of host species can be infected by the same bacterium, yet the clinico-pathologic features among these hosts remains almost identical. The disease has been recognized regularly among ratites, but not in other avian families, such as galliforms, even though these suffer uncharacterized enteric conditions. Fresh ileum-colon contents were obtained from 228, 3- to 8-week-old chickens with enteric disease, kept at 14 large commercial farms in the southern USA. DNA was extracted from each sample and subjected to polymerase chain reactions (PCR) with primers specific to eubacterial DNA, L. intracellularis, and Bilophila wadsworthia. All chicken samples were positive for eubacterial DNA, 29 chickens (13%) were positive for B. wadsworthia DNA, and none were positive for L. intracellularis DNA. Given the ubiquitous nature of L. intracellularis, we consider it likely that some avian families do not carry the necessary mechanism for L. intracellularis viability. Bilophila wadsworthia appears to be a consistent member of the colonic flora of some host animals. Neither bacterium appears to be associated with malabsorption syndromes in chickens.

Molecular characterization confirms the presence of a divergent strain of canine coronavirus (UWSMN-1) in Australia.

Canine coronavirus (CCV) UWSMN-1 was originally identified from an outbreak of fatal gastroenteritis in breeding colonies. In this report, we examined whether UWSMN-1 represents a novel divergent strain or is the result of recombination events between canine and feline coronavirus strains. Sequencing of various regions of the spike and polymerase genes confirms that UWSMN-1 is widely divergent from other CCV and feline coronavirus strains. These data raise the possibility that this strain is the first member of a novel third subtype of CCV.

Cerebral arteriosclerosis in an aged captive polar bear (Ursus maritimus).

Cerebral arteriosclerosis was observed upon necropsy of a 36-yr-old female captive polar bear (Ursus maritimus) that developed a sudden onset of seizure-like activity and died. The medium and large cerebral arteries of the meninges had moderate to severe diffuse discoloration and mineralization of the matrix of the tunica media, with little or no associated cellular reaction. Scanning electron microscopy of the affected arteries showed discrete crystalline calcified deposits in the media and sclerosis of the arterial wall. There were no lesions in the brainstem. The findings suggested a sudden and rapidly fatal loss of blood flow to the brain caused by long-standing arterial lesions. Incidental findings included numerous 0.1- to 10-cm-diameter, hepatic cysts lined with hyperplastic biliary epithelium, a unilateral, unipolar, 3-cm-diameter renal tubular adenoma, and approximately 250 active Baylisascaris sp. nematodes in the intestines.