Detection of hepatitis E virus shedding in feces of pigs at different stages of production using reverse transcription-polymerase chain reaction.

The aim of this study was to determine at which production stages hepatitis E virus (HEV) is shed by the highest number of pigs and to estimate the relative risk associated with each stage. For this purpose, 146 fecal samples of pigs from 21 farms were studied. In addition, 1 sample from the manure ditch and another sample of drinking water, collected directly from the trough located in the pen, were taken from 16 farms. HEV RNA was detected in fecal samples from 34 pigs (23.29%). The production stages in which most pigs excreted HEV were weaners (41.7%) and pigs in the first month of feeding (60%). The results of the statistical analysis showed that the principal significant risk stage in HEV shedding was the first month of feeding (odds ratio [OR] 19.5, 95% CI 3.59-106.07, P = 0.001) followed by the weaners stage (OR 9.3, 95% CI .78-48.42, P = 0.008). In 8 out of 16 farms tested (50%) HEV RNA was detected in raw manure and in the water trough of only 1. Detection of HEV in manure ditches raises the concern of how to deal with manure of swine origin, because it is used as soil fertilizer.     

Post-thaw viability of european bison (Bison bonasus) semen frozen with extenders containing egg yolk or lipids of plant origin and examined with a heterologous in vitro fertilization assay.

Basic characteristics of European bison (Bison bonasus) semen were described and the efficacies of two extenders-Triladyl, containing egg yolk, and a synthetic extender, containing soybean lipids-were tested for semen cryopreservation. Seven ejaculates were collected by electroejaculation from a 10-yr-old, European bison bull. Each ejaculate was diluted at 37 degrees C to a final concentration of 200 x 10(6) sperm/ml with Triladyl or the synthetic extender. Extended semen samples were frozen according to a standard bull semen freezing protocol. After 2 wk of storage, one straw from each extender and ejaculate was thawed, and postthaw quality was evaluated by individual sperm motility and movement rate, numbers of sperm morphologic abnormalities and intact acrosomes, functional integrity of the sperm membranes determined by hypoosmotic swelling test (HOST), viability (live-dead, eosin-nigrosin stain), and a heterologous in vitro sperm penetration assay (SPA). A total of 600 in vitro-matured bovine oocytes were inseminated with 1 X 10(6) spermatozoa of Holstein semen frozen-thawed in Triladyl (control) or of European bison semen frozen in Triladyl or the synthetic extender. Nuclear status of the oocytes was determined after 18 h of sperm-oocyte coincubation. Extender had no effect on any evaluated parameters of semen after dilution and cooling (4 hr at 5 degrees C) or in postthaw individual motility, quality of movement, and sperm morphology. However, significantly (P < 0.05) higher numbers of spermatozoa with intact acrosomes, intact membranes (HOST), and viable sperm (P < 0.01) were in semen frozen in Triladyl than in the synthetic extender. Mean values for heterologous SPA for bull (control) and for bison semen frozen in the synthetic extender were very much alike-63.3+/-10.6% and 63.1 +/- 15.9%, respectively; bison semen frozen in Triladyl was lower, 43.0+/-24.2% but not significantly different. Cumulative results from a variety of viability assays of diluted/cooled and frozen-thawed semen, including the heterologous SPA, suggest that European bison semen can be successfully frozen in both extenders tested in this study.     

A retrospective study of pathologic findings in the Amazon and Orinoco river dolphin (Inia geoffrensis) in captivity.

River dolphins are especially susceptible to negative human impacts. For their conservation, attempts of relocation or procreation ex situ may become important in the future to avoid their extinction. Additional knowledge and medical experiences of river dolphin management in captivity may aid such conservation efforts. The medical records and necropsy and histopathology reports on 123 captive Amazon River dolphins (Inia geoffrensis) were re-viewed. Of these 123 animals, 105 were necropsied and 70 necropsies were supported with histopathology. Eighteen animals were not necropsied. Among wild-born animals, mortality was highest in the first 2 mo immediately postcapture and transport, accounting for 32 of 123 deaths. Pneumonia and skin lesions (cutaneous and subcutaneous ulcerations and abscesses) were the most common findings, found in 44 of 105 (42%) and 38 of 105 (36%) of gross diagnoses, respectively. At least 10 of 44 cases of pneumonia diagnosed grossly included a verminous component. Cachexia, from a variety of causes, was a major gross finding in 21 animals. Fifteen animals had histologic evidence of significant renal pathology, and this was the primary cause of death in 13 cases. Hepatic pathology was found in 18 cases, and bacterial sepsis was confirmed via histology in 16 cases. Based on these findings, it may be concluded that keys to successful maintenance of this species include 1) prophylactic anthelminthic and antibiotic therapy immediately post-capture; 2) maintenance of animals in larger enclosures than in past attempts, in compatible groups, and in facilities capable of separating aggressive animals; 3) maintenance in microbiologically hygienic water quality at all times; and 4) a proactive program of preventive medicine during the immediate postcapture, quarantine, and maintenance period of captivity.     

Comparison of grazing behaviour, dietary overlap and performance in non-lactating domestic ruminants grazing on marginal heathland areas

During the years 2000–2001, 7 non-lactating beef cows, 40 ewes and 40 does were managed in mixed grazing on a natural heathland vegetation plot (22 ha) with 20% improved pasture (perennial ryegrass) on a hill (1000 m a.s.l.) experimental farm located in the NW of Spain. Samples of faeces and vegetation components were collected monthly to estimate diet selection, using the alkane markers, and diet overlapping level. Animals were weighed monthly to quantify live weight changes and performance of the three livestock species during different periods (spring, summer, autumn, winter) of the grazing season.

The percentage of shrubs in the diet was significantly higher in the small ruminants (ranging between 36% and 85%) than in cows (less than 25%) in any period. Gorse (Ulex gallii) and heather (Erica spp., Calluna) percentages were always significantly higher in does than in ewes, except in autumn for heather. Herbaceous component (namely grasses) was higher in cattle (75–99%) than in small ruminants (15–64%). The lowest mean dietary overlap was found between cattle and goats (50.4%), with large differences during the grazing season, ranging between 20% and 70%.

The three animal species increased their live weight in the first grazing period (spring), when the mean sward height on the improved area was higher than 6.0 cm. However, when the sward height was lower than 3.5 cm (summer–winter), cows lost weight (− 437 g/day) while ewes and goats were still able to increase their weight (29 and 5 g/day, respectively).

Therefore, it seems that small ruminants, mainly sheep, are more suitable than cattle from the vegetation utilization and animal performance points of view, as cows were unable to maintain live weight when the preferred grass availability decreases. Goats were the species that included the highest proportion of heathland vegetation components in the diet, especially gorse, although their performance was significantly lower than in sheep. In consequence, small ruminant production systems could be more sustainable than cattle. The results indicate that mixed grazing of sheep and goats could be appropriate in these vegetation communities, allowing the development of sustainable systems, in which animal performance and the efficiency of resource use are maximized.     

Evaluation of the status of capsicum viruses in the main growing regions of Turkey

The incidence, severity and distribution of six viruses infecting capsicum were determined in the main growing areas of Turkey during the 2004 growing season. The surveys covered 50 randomly selected capsicum fields from four different areas in south-east Anatolia and the eastern Mediterranean region. 515 samples were individually collected and tested by DAS-ELISA for Cucumber mosaic cucumovirus (CMV), Alfalfa mosaic alfamovirus (AMV), Potato X potexvirus (PVX), Potato Y potyvirus (PVY), Pepper mild mottle tobamovirus (PMMoV) and Tobacco etch potyvirus (TEV). 64.8% of ELISA-tested capsicum samples (334 out of 515) were infected by one (41.7%) or more (23.1%) viruses. PVY was the most widespread (26.4%), followed by PVX (25.8%), AMV (25.2%), TEV (23%), PMMoV (9.1%) and CMV (8.3%). Surprisingly high AMV infection was found in three areas (Kahramanmaraş, Şanlıurfa and Gaziantep) where AMV is reported for the first time in this study. However, AMV was not detected in Hatay. PMMoV is another new virus, in all the tested areas.     

Evaluation of the API 50CH and API ZYM systems for rapid characterization of Clavibacter michiganensis subsp. sepedonicus, causal agent of potato ring rot

API 50CH and API ZYM systems were used to characterize fifty-three strains of Clavibacter michiganensis subsp. sepedonicus from different geographic locations and several reference strains of the same and different species, including other potato pathogens. Clavibacter michiganensis subsp. sepedonicus strains displayed a high level of homogeneity, both in carbohydrate utilization and in enzymatic activity. Using API 50CH and API ZYM it was possible to differentiate C. michiganensis subsp. sepedonicus strains from the remaining taxa analysed in this study, which included representative strains of the other subspecies of C. michiganensis as well as other bacterial pathogens affecting potatoes. Therefore, these systems could be used as an effective method to characterize C. michiganensis subsp. sepedonicus. Such a procedure would constitute an alternative system to the conventional nutritional and physiological identification tests currently included in the official methods employed in the European Union to detect and identify this bacterium. The results obtained with the API systems agreed with the current taxonomic classification of C. michiganensis, clearly separating sepedonicus from the other subspecies belonging to this species.     

<Emphasis Type="Italic">Amaranthus leaf mottle virus</Emphasis>: 3′-end RNA sequence proves classification as distinct virus and reveals affinities within the genus <Emphasis Type="Italic">Potyvirus</Emphasis>

Amaranthus leaf mottle virus (AmLMV) was classified as a member of the genus Potyvirus on the basis of its particle morphology, serology, and biological properties (Casetta et al., 1986). Based on these properties, an Amaranthus viridis-infecting virus isolated in Spain, causing mottle and leaf blistering as well as reduced growth has been identified as AmLMV. The 3′ terminal genomic region of this and a reference isolate from Italy has been sequenced and reveals a 95% nucleotide identity between the two isolates. The sequenced part comprises the coat protein with 281 amino acids and 315 nucleotides of the 3′ untranslated region (UTR) preceding a polyadenylated tail. Pairwise comparisons and phylogenetic analysis of the nucleotide and deduced amino acid sequences of the CP and 3′ UTR of the cloned cDNAs with those of other potyviruses shows that AmLMV is a distinct potyvirus closely related to Potato virus Y.     

A comparison of the adrenal cortical response to ACTH stimulation in Angora and non-Angora goats

The adrenal cortex is believed to be implicated in the high incidence of abortion in the Angora goat. Stimulation testing with adrenocorticotrophic hormone (ACTH) was used to assess the adrenal cortical function in 5 Angora does from herds with a history of abortion and 5 non-Angora does. An acute test involving a single intramuscular (i.m.) injection of 0.25 mg of synthetic ACTH was given during anoestrus, at mid-oestrus, on day 90 and on day 120 of gestation. Blood samples were collected from the jugular vein at 30 min intervals for 1 h before and 5 h after injection. Cortisol concentrations rose within 30 min and returned to baseline values within 3.5 h. Cortisol production was lower (p<0.01) in the pregnant state compared to the non-pregnant state in both groups. Production of cortisol was consistently lower (p<0.05) in the Angora does compared to the non-Angora does during anoestrus and pregnancy and marginally so at mid-oestrus. A chronic stimulation test involving once daily injections of 0.5 mg of a depot form of ACTH i.m. for 7 days commencing on day 90 of pregnancy was also conducted. Cortisol concentrations rose to reach a peak on the third day of treatment in both groups. The values then declined in the Angora does despite continued ACTH treatment, while those for the non-Angora does exhibited a second peak. During and following this treatment, two non-Angora does delivered live kids (day 95, day 120). Out of 7 Angora pregnancies, one Angora doe aborted two dead fetuses at day 116. No significant difference in the cortisol response in the acute test was detected between the animals that aborted and their respective cohorts, but the two non-Angora does that aborted had significantly lower cortisol concentrations during depot ACTH administration. Progesterone and oestradiol levels did not differ between Angora and non-Angora animals during pregnancy or on the test days. The results suggest that the steroidogenic response of the adrenal cortex to ACTH stimulation is significantly less in Angora does with a history of abortion than it is in non-Angora does and support the view that the Angora goat would make a more limited adrenal cortical response to a stressful occurrence during pregnancy.     

Purification and characterization of a protease from Actinobacillus pleuropneumoniae serotype 1, an antigen common to all the serotypes.

A high molecular-mass proteolytic enzyme of Actinobacillus pleuropneumoniae serotype 1, was purified from culture supernatants (CSN) by using DEAE-cellulose and sepharose-4B-gelatin chromatography. In 10% SDS-polyacrylamide gels copolymerized with porcine gelatin, the protease showed a single band of activity of > 200 kDa. However, minor molecular-mass proteolytic bands were observed when the protease was electrophoresed in the presence of either 5% beta-mercaptoethanol, 50 mM dithiothreitol, or 0.25 M urea. Furthermore, when the > 200-kDa purified protein was passed through a sucrose gradient, several bands with proteolytic activity were found: 62, 90, 190, and 540 kDa. The proteolytic activity was increased in the presence of calcium or zinc and was not affected after being heated at 90 degrees C for 5 min. Proteolytic activities were also observed in CSN from all A. pleuropneumoniae serotypes and biotypes. The purified protease hydrolyzed porcine IgA and IgG in vitro. In addition, by immunoblot the protease was recognized by serum of naturally infected pigs with serotypes 1 and 5, and by serum of pigs experimentally infected with serotypes 1, 2, 8, or 9. Serum of a pig vaccinated with CSN of a serotype 3 strain also recognized the protease, but not sera of pigs vaccinated with a bacterin (serotype 1). Proteins from CSN of all the serotypes, which were precipitated with 70% (NH4)2SO4, were recognized by a polyclonal antibody raised against the purified protease. Taken together these results indicate that an antigenic protease is produced in vivo by all the serotypes of A. pleuropneumoniae. The results indicate that proteases could have a role in the disease and in the immune response of pigs infected with A. pleuropneumoniae.     

Pulmonary pharmacokinetics of tulathromycin in swine. Part I: Lung homogenate in healthy pigs and pigs challenged intratracheally with lipopolysaccharide of Escherichia coli

The objective of the study was to assess the pharmacokinetics of tulathromycin in lung tissue homogenate (LT) and plasma from healthy and lipopolysaccharide (LPS)‐challenged pigs. Clinically healthy pigs were allocated to two dosing groups of 36 animals each (group 1 and 2). All animals were treated with tulathromycin (2.5 mg/kg). Animals in group 2 were also challenged intratracheally with LPS from Escherichia coli (LPS‐Ec) 3 h prior to tulathromycin administration. Blood and LT samples were collected from all animals during 17‐day post‐tulathromycin administration. For LT, one sample from the middle (ML) and caudal lobes (CL) was taken. The concentration of tulathromycin was significantly lower in the ML after the intratracheal administration of LPS‐E. coli (P < 0.02). In healthy pigs and LPS‐challenged animals, the distribution of the drug into the lungs was rapid and persisted at high levels for 17‐day postadministration. The distribution of the drug within the lung seems to be homogenous, at least between the middle and caudal lobes within dosing groups. The concentration versus time profile of the drug and pharmacokinetic parameters in two different lung areas (middle and caudal lobe) were consistent within the groups. The clinical significance of these findings is unknown.