Prevalence, disease associations and risk factors for colonization with intestinal spirochaetes (Brachyspira spp.) in flocks of laying hens in north-eastern Italy.

The present study investigated the occurrence of anaerobic intestinal spirochaetes of the genus Brachyspira in laying hen flocks in Treviso province, north-eastern Italy, with respect to prevalence, spirochaete species present, disease associations and risk factors for colonization. A total of 450 faecal samples from 45 sheds on 29 laying hen farms were cultured for intestinal spirochaetes. Nineteen sheds on 12 farms contained chickens with symptoms consistent with avian intestinal spirochaetosis, including reduced egg production, wet litter and/or pasty vents. Spirochaetes were isolated from 157 (34.8%) samples from 21 (72.4%) farms, and from 32 (71.1%) sheds. From these positive samples, 189 spirochaetal isolates were speciated using three polymerase chain reaction assays and a restriction fragment polymorphism analysis of 16S rDNA polymerase chain reaction products. Overall, 52 (27.5%) isolates were identified as pathogenic Brachyspira intermedia, 26 (13.8%) as pathogenic Brachyspira pilosicoli, 93 (49.7%) as non-pathogenic (Brachyspira innocens/Brachyspira murdochii), and 18 (9.6%) were unidentified. Faeces from 14 sheds (31%) on 10 farms (34.5%) contained B. intermedia and/or B. pilosicoli, and disease consistent with avian intestinal spirochaetosis was observed in nine of these sheds on seven farms. There was a significant association (P=0.042) between the presence of spirochaetes and using deep pits rather than conveyor belts for manure disposal. Sheds housing chickens >40 weeks of age were significantly more likely to contain spirochaetes (P=0.048) and pathogenic species (P=007) than sheds housing younger chickens. A significant association (P=0.02) was found between infection with pathogenic spirochaetes and reduced egg production.

Occurrence and antibiotic susceptibility of Helicobacter pullorum from broiler chickens and commercial laying hens in Italy.

In 2005, in order to investigate the occurrence of Helicobacter pullorum in poultry, the caecal contents collected from a total of 60 animals intensively reared in Italy on 15 different farms (9 farms of broiler chicken and 6 of laying hens) were examined at the slaughterhouse. A modified Steele-McDermott membrane filter method was used. Small, greyish-white colonies of Gram-negative, gently curved, slender rod bacteria were preliminarily identified as H. pullorum by a Polymerase Chain Reaction (PCR) assay based on 16S rRNA and were then subjected to an ApaLI digestion assay to distinguish H. pullorum from Helicobacter canadensis. One isolate from each farm was phenotypically characterized by biochemical methods and 1D SDS-PAGE analysis of whole cell proteins; antibiotic susceptibility was also tested. According to the PCR and PCR-RFLP results, all the animals examined were positive for H. pullorum. The 1D SDS-PAGE whole protein profile analysis showed high similarity among the 15 isolates tested. A monomodal distribution for the Minimum Inhibitory Concentrations (MICs) was found for ampicillin, chloramphenicol, gentamicin and tetracycline. For erythromycin and ciprofloxacin, a bimodal trend having a second peak at >128 micro(-1) and 32 micro(-1) was found. The isolation method used in this study seems to be highly suitable for isolating H. pullorum from chicken caecal contents. Moreover, the detection of a high number of colonies phenotypically similar to H. pullorum suggests that this microorganism, when present, colonizes the caecum at high concentration.

Hypertrophy of intestinal smooth muscle in cats.

Pathological findings of four cats with severe and diffuse smooth muscle hypertrophy of the small intestine (MHSI) are reported and compared to those of five cats with segmental MHSI secondary to neoplastic obstruction and four controls. Histology demonstrated a constant association between idiopathic MHSI and submucosal fibrosis and chronic lymphoplasmacytic enteritis. Morphometry (gut diameter, thickness and area of muscular layers, number and density of smooth muscle nuclei) and MIB-1-immunolabelling showed that the thickness increase was mostly due to hypertrophy, but hyperplasia was also evident. Microbiology from ileal content samples was performed in two cats with primary MHSI, and Campylobacter spp. were isolated, which were also demonstrated by immunohistochemistry and ultrastructure. The association of chronic enteritis with idiopathic MHSI suggests that factors released in intestinal inflammation may also act as hypertrophy stimuli for smooth muscle cells.

Occurrence and characterization of gastric Helicobacter spp. in naturally infected dogs.

Helicobacter-like organisms are frequently observed in the stomach of dogs but the relationship between these microorganisms and gastric pathology has not been clearly established. Different species of helicobacters are known to be present in the canine stomach but their specific prevalence in naturally infected dogs is unknown. The aims of this study were to isolate and characterize helicobacters in canine gastric biopsies, to compare the commonly used tests for the identification of Helicobacter spp. and to determine the occurrence of these species in dogs. Twenty-three out of 25 dogs (92%) were positive for Helicobacter-like organisms in cytological screening. Culture was successful from biopsies of 5/25 dogs. The isolates were analyzed by electron microscopy, biochemical and physiological tests, whole protein analysis and 16S rDNA sequencing. Helicobacter felis was identified in four samples and Helicobacter bizzozeronii in one sample. Only the whole protein analysis in combination with electron microscopy was able to clearly discriminate the two species. Compared to the high prevalence of Helicobacter-like organisms, the occurrence of H. felis and H. bizzozeronii, was low (17 and 4%, respectively). No Flexispira rappini-like organisms or H. salomonis were detected. Electron microscopy revealed that H. bizzozeronii-like microorganisms were present in three additional biopsies where we were unable to culture any Helicobacter-like organisms. These observations indicate that in the stomach of dogs not all helicobacters are culturable. The unculturable bacteria appeared to be the prevalent ones and may represent different spiral organisms. The presence of distinct helicobacters with different characteristics can reflect different roles in the pathogenesis of canine gastric disease.

Monitoring of transmission of tuberculosis between wild boars and cattle: genotypical analysis of strains by molecular epidemiology techniques.

An epidemiological survey for the monitoring of bovine tuberculosis transmission was carried out in western Liguria, a region in northern Italy. Fifteen Mycobacterium bovis strains were isolated from 63 wild boar samples (62 from mandibular lymph nodes and 1 from a liver specimen). Sixteen mediastinal lymph nodes of 16 head of cattle were collected, and 15 Mycobacterium bovis strains were subsequently cultured. All M. bovis strains isolated from cattle and wild boars were genotyped by spoligotyping and by restriction fragment length polymorphism (RFLP) analysis with the IS6110 and IS1081 probes. All M. bovis strains showed the typical spoligotype characterized by the absence of the 39 to 43 spacers in comparison with the number in M. tuberculosis. A total of nine different clusters were identified by spoligotyping. The largest cluster included 9 strains isolated from wild boars and 11 strains isolated from cattle, thus confirming the possibility of transmission between the two animal species. Fingerprinting by RFLP analysis with the IS6110 probe showed an identical single-band pattern for 29 of 30 strains analyzed, and only 1 strain presented a five-band pattern. The use of IS1081 as a second probe was useful for differentiation of M. bovis from M. bovis BCG but not for differentiation among M. bovis strains, which presented the same undifferentiated genomic profile. In relation to the epidemiological investigation, we hypothesized that the feeding in pastures contaminated by cattle discharges could represent the most probable route of transmission of M. bovis between the two animal species. In conclusion, our results confirmed the higher discriminatory power of spoligotyping in relation to that of RFLP analysis for the differentiation of M. bovis genomic profiles. Our data showed the presence of a common M. bovis genotype in both cattle and wild boars, confirming the possible interspecies transmission of M. bovis.

Interaction between vascular prostheses and rifampicin in the prevention of the grafts infection. An experimental study.

Infections caused by synthetic prostheses are relatively rare (1.5-6%) but serious complication in vascular surgery. There is no doubt that during and immediately after surgery bacterial contamination may occur. An in vitro study was carried out in the Vascutek laboratories, which revealed a high affinity between prostheses in Dacron gel and Rifampicin. This affinity, the result of an ionic bond, was demonstrated by the fact that after 5 days Rifampicin was still present on the prostheses. Encouraged by this result, an experimental study was carried out in sheep. Five sheep were operated on making a prosthetic graft in both of the common carotid arteries: on one side a Gelseal Dacron prosthesis was implanted after being soaked for 15 minutes in a solution containing 1 mg/ml Rifampicin. A Knitted Dacron prosthesis was implanted in the contralateral carotid artery, again after pretreatment with Rifampicin. Explants were made after 2, 24, 48, 72 and 96 hours, and the concentration of Rifampicin on the prostheses was assessed on the basis of the diameter of the inhibition area on Staphylococcus aureus cultures. The results showed that the Gelseal Dacron prostheses maintained Rifampicin concentrations with an antibacterial activity up to 72 hours; this property disappears with the Knitted Dacron prostheses after only 24 hours. These results confirm the high affinity of Gelseal Dacron and Rifampicin also in in vivo experimental models.

Faecal bacteria of wild ruminants and the alpine marmot.

Faecal samples from 60 red deer (Cervus elaphus), 13 roe deer (Capreolus capreolus), 7 chamois (Rupicapra rupicapra), 41 alpine marmot (Marmota marmota) and soils mixed with deer faeces from the Stelvio National Park were examined for Campylobacter sp. and Salmonella sp. with negative results. The same material, especially deer faeces, was a habitat highly suitable for Yersinia sp.: Y. enterocolitica (two biotypes) was isolated twice, Y. kristensenii (two serotypes) was isolated 19 times, Y. frederiksenii and Y. intermedia were isolated once. Antibiotic-resistant Escherichia coli were isolated from 16 specimens from wild ruminants, one from marmot and two from feeding places.

Vaccination of cattle with live and inactivated rabies vaccines: a study of antibody response.

The authors vaccinated 152 cattle divided into three groups against rabies. The first group received the ERA strain and the second group an inactivated vaccine. The third group received the inactivated vaccine on two occasions with an interval of 60 days between the two doses. Their antibody response was surveyed with the fluorescent foci-inhibition test carried out on blood samples collected during a 10-month period. All animals developed an almost identical antibody response. However, at the sixth and tenth months, there was a higher number of seropositive animals in the groups vaccinated with the killed vaccine.

A survey of 120 isolates of Malassezia (Pityrosporum) pachydermatis. Preliminary study.

The morphological, cultural and biochemical characteristics of 120 isolates of Malassezia (Pityrosporum) pachydermatis, isolated from chronic otitis externa in the dog, are discussed.

Mycoplasma mycoides subsp mycoides, LC type, from goats in Italy.

A mycoplasma identified as Mycoplasma mycoides subsp mycoides LC type was isolated from an outbreak of caprine pleuropneumonia and serofibrinous peritonitis. This isolate was the first to be reported in Italy. Experimentally, a rapidly fatal condition followed its inoculation into goats and sheep but inoculation of calves did not lead to clinically apparent infection. Although the organism was recovered for up to 45 days following experimental inoculation of calves, pathological changes were limited to the drainage lymph nodes except in one case where there were histological changes in the lungs.