Mortality of one-week-old chickens during naturally occurring Marek's disease virus infection.

Marek's disease (MD) is a disease of chickens that occurs worldwide and has serious economic consequences. MD can present as one of several forms, with the most commonly occurring forms being the lymphoproliferative diseases. Under experimental conditions, an early mortality syndrome has been recognized following infection by some but not all strains of MD virus (MDV). This is the first report of a confirmed case of mortality due to naturally occurring MDV infection in 1-week-old, nonvaccinated, chickens. Necrotizing lesions were observed in the bursa of Fabricius, lung, duodenum, jejunum, and proventriculus, and large intranuclear inclusion bodies were a striking feature in tissues with lesions in all birds. Immunohistochemical staining for the pp38 protein of MDV revealed abundant pp38 antigen in the affected tissues, confirming the presence of MDV within the lesions. PCR yielded an amplicon with 97% homology to the meq gene of MDV. No evidence of co-infection by either of the immunosuppressive agents chicken anemia virus and infectious bursal disease virus was detected.

Sequence comparison of pigeon circoviruses.

The genome sequences of eight pigeon circoviruses (PiCV) were determined and compared with four previously published sequences. The viruses compared were from the USA, five European countries, China and Australia and included PiCVs from racing, feral, ornamental and meat pigeons and a Senegal dove (Streptopelia senegalensis). The 12 PiCV genomes, ranging from 2032 to 2040 nucleotides in length, displayed similar organizations. Pairwise comparisons showed that the genome nucleotide sequence identities ranged from 85.1% to 97.8% and that the amino acid identities of the putative replication associated (Rep) and putative capsid (Cap) proteins displayed ranges of 91.5-99.1% and 73.0-99.3%, respectively. Comparative analyses identified conserved nucleotide sequences within the Rep gene and 3' intergenic regions, which would be suitable for diagnostic PCR primers, and variable amino acid sequences within the capsid proteins, which should be considered when selecting virus isolates for vaccine development.

Fatal necrotising fasciitis and myositis in a cat associated with Streptococcus canis.

Necrotising fasciitis and necrotising myositis are rare but serious life threatening conditions reported mainly in human beings and dogs. Most cases to date have been caused by beta-haemolytic streptococci of Lancefield groups A, B, C or G. Necrotising fasciitis has been reported only twice in cats and necrotising myositis has never been described. This paper describes a fatal case of necrotising fasciitis and myositis with pneumonia and septicaemia in a nine-year-old cat. The cat had been undergoing treatment for a suspected tear of the cranial cruciate ligament, but on the seventh day of treatment it suddenly deteriorated and died. On postmortem examination, there was an area of hair loss from its left hindlimb and discoloration of the underlying fascia and biceps femoris muscle. Severe necrotising fasciitis and myositis, with numerous intralesional Gram-positive coccoid bacteria, was diagnosed histologically. Other findings included necrotising pneumonia, pleuritis, focal encephalitis, myocarditis and nephritis. Culture of the affected tissues yielded a pure, heavy growth of Streptococcus canis.

Detection of pigeon circovirus in cloacal swabs: implications for diagnosis, epidemiology and control.

Pigeon circovirus (picv) was detected in cloacal swab samples by means of a newly-developed, sensitive pcr. An initial investigation of 17 Belgian racing pigeons aged up to eight months showed that rates of detection of 88 per cent and above were achieved using samples of cloacal swab, blood and bursa of Fabricius. The sampling of 15 caged pigeons six times when they were from three to 31 weeks of age indicated that picv infections were more readily detected in cloacal swabs than in blood, and that the virus could be detected in cloacal swabs for longer periods after infection than in blood. picv infections were detected in cloacal swabs from 38 of 47 young pigeons aged from two to 31 weeks, from 12 racing lofts, which had clinical signs including diarrhoea and weight loss, regurgitation and respiratory signs. Samples from birds from two infected lofts indicated that picv could be detected in some birds for at least 27 weeks. Although nine of 14 pigeons aged from 32 to 45 weeks were virus-positive, picv was detected in only one of 18 adult pigeons that originated from four infected lofts.

Characterisation of avian pathogenic Escherichia coli (APEC) associated with colisepticaemia compared to faecal isolates from healthy birds.

A total of 114 avian pathogenic Escherichia coli (APEC) isolates were collected from cases of colisepticaemia occurring in broilers (77) and layers (37) within Ireland. In addition 45 strains isolated from faeces of healthy birds were included for comparison. All isolates were serogrouped, and examined for known virulence factors, mostly by PCR. The O78 serogroup represented 55 and 27% of broiler and layer colisepticaemic isolates respectively. All isolates were positive for curli fimbriae (crl, csg) and negative for afimbrial adhesin (afa). S-fimbrial (sfa) sequences were present in 8.8% of septicaemic isolates and 8.9% of healthy bird isolates. The majority of E. coli from cases of colisepticaemia (97.4%) and healthy bird (95.6%) isolates were positive for aerobactin (aer), and temperature sensitive haemagglutinin (tsh) was similarly detected in high numbers in 93.9 and 93.3%, respectively. In comparison to E. coli isolates from the faeces of healthy birds, a significantly higher percentage of isolates from septicaemic cases possessed Type 1 fimbriae (fimC) and increased serum survival (iss) gene sequences. Forty-seven (41.2%) isolates from septicaemic birds possessed P-fimbriae (pap) gene sequences, compared with only 15.6% from E. coli isolated from healthy birds. Haemolysin (hlyE) sequences were detected in 46.7% of isolates from healthy birds in comparison with 6.1% of septicaemic isolates. Sequences encoding colicin V (cvaC) were detected in 99.1% of septicaemic isolates and 82.2% of isolates from healthy birds. The K1 capsule was only present in two septicaemic isolates, both taken from layers. Motility was detected in 36.8% of E. coli isolated from cases of septicaemia, compared with 93.3% of isolates from healthy birds. These results demonstrate the presence of 11 virulence genes in E. coli isolated from cases of colisepticaemia within Ireland, and indicate the prevalence of iss and fimC.

Sandwich ELISA detection of Clostridium perfringens cells and alpha-toxin from field cases of necrotic enteritis of poultry.

Sandwich ELISAs (sELISAs) for the detection of Clostridium perfringens cells and alpha-toxin were developed and used to screen intestinal samples from normal broiler chickens and from clinical cases of necrotic enteritis. The assays clearly distinguished between the two sets of samples. The sELISA absorbance values from samples obtained from the majority of healthy birds were low and those from the majority of necrotic enteritis cases were high. Together, the assays provide a suitable test for the rapid screening for the diagnosis of necrotic enteritis in poultry.

New data on the transmission of pigeon circovirus.

Nineteen racing pigeons aged from one to five years were examined postmortem. pcr tests showed that the spleens of 16 of them were positive for pigeon circovirus, the livers of six were positive, and blood from one of them was positive for the virus. Five of 44 embryos in embryonated eggs collected from three lofts were positive by pcr, but swabs taken from the crops of 64 adult birds which were feeding one- to 10-day-old squabs in these three lofts were negative for the viral dna.

Isolation of bovine adenovirus serotype 6 from a calf in the United Kingdom.

Two viruses, designated 99-8130(C) and 99-8130(I), were isolated in calf testis cells from the colon and ileum, respectively, of a suckled beef calf which had developed dysentery and died. Electron microscopy indicated that the mean (sd) size of the viral particles, 83 (2.5) nm, and their morphology were consistent with their being members of the family Adenoviridae. They were confirmed as adenoviruses by PCR when products of the expected size (608 bp) were amplified from both isolates by using a primer pair specific for members of the genus Atadenovirus. A comparison of the sequence of a 567 bp segment of the 99-8130(C) amplicon with that of other prototype bovine adenovirus (BAdV) strains of atadenoviruses identified the isolate as BAdV serotype 6 (BAdV-6), which had 99.3 per cent and 100 per cent identities at the nucleotide and amino acid levels, respectively, with the prototype BAdV-6 strain 671130. A virus neutralisation test was developed and indicated a high prevalence of antibody to BAdV-6 in Northern Irish cattle. There was no evidence of adenoviral inclusions in tissues from the affected calf and no antigen was detected when the tissues were stained by an immunoperoxidase technique, using a homologous antiserum raised in rabbits. The two viruses were the third reported isolation of BAdV-6, and the first from a clinically ill bovine animal.

Pulmonary function and bronchial hyperreactivity in long-term survivors of bronchopulmonary dysplasia.

Nine children with bronchopulmonary dysplasia were studied at a mean age of 8.4 years. Persistent respiratory symptoms were common. Pulmonary function tests demonstrated airway obstruction and abnormal blood gas levels in most. In six of eight children results of methacholine challenge tests were positive, indicating bronchial hyperreactivity. Abnormal chest radiographs persisted in eight children. The bronchial hyperreactivity is a likely consequence of airway damage in infancy and is the probable reason for episodic wheezing in the early years of life. Improvement in expiratory flow rates following bronchodilator inhalation suggests that such therapy may improve function in these children.

Pulse oximetry advantages in infants with bronchopulmonary dysplasia.

We studied 12 infants with a clinical and radiologic diagnosis of bronchopulmonary dysplasia who were oxygen dependent and older than 30 days. Simultaneous readings of hemoglobin oxygen saturation (SaO2) determined by two pulse oximeters (Nellcor 100, BTI Biox III) and transcutaneous (tc) PO2 (Sensor Medics, Transend) were correlated with SaO2 (Radiometer, OSM 2 Hemoximeter) and PaO2 (Corning 178) measured on blood from an indwelling arterial catheter. For each infant, the fractional inspiratory oxygen (FiO2) was adjusted to obtain three to five sets of data in the range of 70% to 95% SaO2. Fifty-three data points were generated and pooled for analysis. The slope of the regression line generated for the Nellcor 100 was .86; for the BTI Biox III, it was .91; and for the Sensor Medics Transend, it was .55, resulting in average errors of +2.5%, +1.0%, and -29%, respectively, when comparing corresponding transcutaneous and arterial values. When SaO2 was equal to or less than 95%, no infants were hyperoxic. These data confirm reports by others that tcPO2 values do not accurately represent PaO2 values in older infants with bronchopulmonary dysplasia. Pulse oximeters do not require user calibration, and their sensor is unheated so they will not cause skin burns. We conclude that pulse oximetry offers major advantages over tcPO2 measurements in the management of infants with bronchopulmonary dysplasia.

Hyaline membrane disease treated with bovine surfactant.

Six preterm infants with severe hyaline membrane disease requiring ventilation were treated, at a median age of 15.5 hours, with a single intratracheal bolus of a bovine surfactant suspension. Arterial oxygenation increased dramatically, and chest radiograms showed improvement after two to four hours. However, a variable degree of deterioration occurred within 24 hours. All of the infants required oxygen therapy for several weeks, and one developed severe bronchopulmonary dysplasia.

In situ hybridization: an optimised detection protocol for a biotinylated DNA probe renders it more sensitive than a comparable 35S-labelled probe.

The development of a highly sensitive biotinylated in situ hybridization protocol using egg drop syndrome virus infection of domestic fowl as a model is described. This protocol incorporates the use of a monoclonal antibody to biotin as the initial step in a peroxidase-based detection system. The detection of viral nucleic acid in formalin-fixed, paraffin-embedded sections using biotinylated and 35S-labelled probes is compared to the detection of viral antigen by immunocytochemistry. Biotinylated probes detected more positive cells than 35S-labelled probes and were more specific. Biotinylated probes detected more positive cells than immunocytochemistry.

Bacterial chondronecrosis with osteomyelitis ('femoral head necrosis') of broiler chickens: a review.

Bacterial chondronecrosis with osteomyelitis (BCO) in chickens was first reported in 1972 and is now recognized as an important cause of lameness in broiler chickens. Recent systematic studies of causes of lameness in birds reared in Northern Ireland have shown that it was the most common cause of lameness, being present in 17.3% of lame birds. Furthermore, it was also detected in birds presented as ''found dead''. Overall losses in male birds due to BCO were estimated to be 0.75% of all birds placed, which, in addition to welfare concerns, represents considerable economic loss. The disease has been seen in birds ranging from 14 to 70 days of age, but most cases occurred around 35 days old. It is most commonly caused by Staphylococcus aureus, but Escherichia coli, coagulase-negative staphylococci and Enterococcus spp. are sometimes involved, as are, rarely, other bacteria. The lesions are most commonly found associated with the growth plates of long bones, particularly the proximal growth plate of the femur and tibiotarsus, but other bones may also be affected. Since lesions were visible to the naked eye in only 40 to 67% of cases, histological examination is recommended where no lesions are visible macroscopically. As the lesion may be present in only one growth plate, and because histological examination is often not carried out, BCO is almost certainly underdiagnosed. The exact pathogenesis of the condition is unknown, but it is thought that adherence of blood-borne bacteria to exposed cartilage at the tips of metaphyseal blood vessels is fundamental. Under controlled experimental conditions, infection of birds with the immunosuppressive viruses chicken anaemia virus and infectious bursal disease virus increased the incidence of the disease, while restricting feed intake reduced the incidence of disease. S. aureus strains identical to, or closely related to, isolates recovered from naturally occurring cases of the disease (as determined by pulsed-field gel electrophoresis) have been recovered from fluff-debris in hatcheries, and also from the environment of breeding flocks, indicating that infection in the breeding farm and in the hatchery could be an important source of infection. It has also been shown that humans can carry poultry strains of S. aureus on their hands. There is a higher incidence of BCO in birds hatched from floor eggs. Thus, hygiene and management practice on breeder farms and in the hatchery may influence the occurrence of the disease. Bacteraemia is a prerequisite for BCO. Indeed, in some flocks suffering losses due to BCO, there are also losses due to staphylococcal septicaemia. Thus, appropriate treatment of affected flocks should reduce losses due to septicaemia. It should also reduce the occurrence of bacteraemia and the development of further cases of BCO. However, birds in which BCO has already developed, are unlikely to respond to treatment. Control of BCO by vaccination seems unlikely in the short term. Simple bacterins have not been effective and much basic research is needed to identify the important virulence factors. Furthermore, more than one type of bacterium is capable of causing the disease. Bacterial interference has been used successfully in humans and turkeys to prevent staphylococcal diseases, and warrants investigation for the prevention of BCO in chickens. This may have an advantage in that the interfering bacterium may also exclude some of the other bacteria that can cause BCO. The recent development of a disease model in which S. aureus is given by a natural route allows the potential for further investigation of the role of predisposing factors, and intervention strategies, including vaccination and bacterial interference, for the prevention of BCO.

Bacterial chondronecrosis with osteomyelitis ('femoral head necrosis') of broiler chickens: a review.

Bacterial chondronecrosis with osteomyelitis (BCO) in chickens was first reported in 1972 and is now recognized as an important cause of lameness in broiler chickens. Recent systematic studies of causes of lameness in birds reared in Northern Ireland have shown that it was the most common cause of lameness, being present in 17.3% of lame birds. Furthermore, it was also detected in birds presented as ''found dead''. Overall losses in male birds due to BCO were estimated to be 0.75% of all birds placed, which, in addition to welfare concerns, represents considerable economic loss. The disease has been seen in birds ranging from 14 to 70 days of age, but most cases occurred around 35 days old. It is most commonly caused by Staphylococcus aureus, but Escherichia coli, coagulase-negative staphylococci and Enterococcus spp. are sometimes involved, as are, rarely, other bacteria. The lesions are most commonly found associated with the growth plates of long bones, particularly the proximal growth plate of the femur and tibiotarsus, but other bones may also be affected. Since lesions were visible to the naked eye in only 40 to 67% of cases, histological examination is recommended where no lesions are visible macroscopically.As the lesion may be present in only one growth plate, and because histological examination is often not carried out, BCO is almost certainly underdiagnosed. The exact pathogenesis of the condition is unknown, but it is thought that adherence of blood-borne bacteria to exposed cartilage at the tips of metaphyseal blood vessels is fundamental. Under controlled experimental conditions, infection of birds with the immunosuppressive viruses chicken anaemia virus and infectious bursal disease virus increased the incidence of the disease, while restricting feed intake reduced the incidence of disease. S. aureus strains identical to, or closely related to, isolates recovered from naturally occurring cases of the disease (as determined by pulsed-field gel electrophoresis) have been recovered from fluff-debris in hatcheries, and also from the environment of breeding flocks, indicating that infection in the breeding farm and in the hatchery could be an important source of infection. It has also been shown that humans can carry poultry strains of S. aureus on their hands. There is a higher incidence of BCO in birds hatched from floor eggs. Thus, hygiene and management practice on breeder farms and in the hatchery may influence the occurrence of the disease. Bacteraemia is a prerequisite for BCO. Indeed, in some flocks suffering losses due to BCO, there are also losses due to staphylococcal septicaemia. Thus, appropriate treatment of affected flocks should reduce losses due to septicaemia. It should also reduce the occurrence of bacteraemia and the development of further cases of BCO. However, birds in which BCO has already developed, are unlikely to respond to treatment. Control of BCO by vaccination seems unlikely in the short term. Simple bacterins have not been effective and much basic research is needed to identify the important virulence factors. Furthermore, more than one type of bacterium is capable of causing the disease. Bacterial interference has been used successfully in humans and turkeys to prevent staphylococcal diseases, and warrants investigation for the prevention of BCO in chickens. This may have an advantage in that the interfering bacterium may also exclude some of the other bacteria that can cause BCO. The recent development of a disease model in which S. aureus is given by a natural route allows the potential for further investigation of the role of predisposing factors, and intervention strategies, including vaccination and bacterial interference, for the prevention of BCO.

Nucleotide sequence-based identification of a novel circovirus of canaries.

Circovirus-like, spherical particles measuring 16 to 18 nm in diameter were detected in organ homogenates from adult canaries that had died after a short illness characterized by dullness, anorexia, lethargy and feather disorder. A polymerase chain reaction method, based on degenerate primers specific to conserved amino acid sequences in the circovirus replication-associated protein, was used to amplify DNA specific to a novel circovirus, tentatively named canary circovirus (CCV). Sequence analysis of a 510 nucleotide genomic fragment indicated that CCV exhibited 67.4, 64.9, 53.7 and 53.9% nucleotide identities and 70.0, 61.8, 40.4 and 40.1% amino acid identities with columbid (pigeon) circovirus (CoCV), beak and feather disease virus (BFDV), porcine circovirus type 1 and porcine circovirus type 2, respectively. CCV therefore represents a new avian virus of the genus Circovirus of the family Circoviridae, and is more closely related to CoCV than BFDV.The availability of nucleotide sequence data will facilitate the development of DNA-detecting diagnostics with which the prevalence of CCV infections can be assessed.

Comparison of Staphylococcus aureus recovered from personnel in a poultry hatchery and in broiler parent farms with those isolated from skeletal disease in broilers.

Personnel from one broiler hatchery, and workers on 18 separate broiler parent farms which supply the hatchery, were tested for hand and nasal carriage of Staphylococcus aureus. In both locations, nasal carriage of S. aureus was more common than hand carriage. A total of 63 S. aureus strains were characterised by biotyping, protein A analysis and pulsed field gel electrophoresis (PFGE) typing. Of these, 36 were recovered from broiler hatchery personnel, 14 from broiler parent farm personnel and 13 from cases of skeletal disease in commercial broilers. Biotyping and protein A analysis indicated that none of the strains recovered from hatchery personnel were of the poultry biotype, but that two strains recovered from the hands of two broiler parent farm personnel could be grouped together with 12/13 of strains recovered from skeletal disease in broilers, as poultry biotypes. PFGE-typing could not distinguish 9/13 strains recovered from skeletal disease in broilers and one of the strains from the broiler parent farm personnel from isolate 24 (I. 24), which is the predominant S. aureus strain type associated with clinical disease in N. Ireland broiler flocks. The present study found no evidence of nasal carriage of S. aureus strains of poultry biotype by humans. The finding of hand carriage by broiler parent farm personnel, suggests that handling by personnel may contribute to the dissemination of I. 24 or other S. aureus strains associated with skeletal disease in broilers.

The use of pulsed field gel electrophoresis to investigate the epidemiology of Staphylococcus aureus infection in commercial broiler flocks.

Pulsed field gel electrophoresis (PFGE) analysis of SmaI restriction patterns was used to type 109 isolates of Staphylococcus aureus collected from broiler farms and hatcheries in Northern Ireland. Forty-seven isolates from clinical conditions in broilers and 62 strains from hatcheries, were examined. The PFGE patterns demonstrated a similarity between 85% of strains from clinical sources and 71% of the hatchery isolates. The association of disease with the predominant strain type and presence of these same strains in the hatchery, indicates that the hatchery is a potential source of the infection for clinical broiler disease.

An investigation into the efficacy of hatchery disinfectants against strains of Staphylococcus aureus associated with the poultry industry.

The efficacy of 18 commercial disinfectants was investigated using the type strain, isolate 24 (I. 24), of pulsed-field gel electrophoresis pattern related Staphylococcus aureus that have shown to be associated with clinical disease in Northern Ireland broilers. Eight quartenary ammonium compound (QAC), four peroxygen, three amphoteric (AMP), one phenolic along with two chlorine-based disinfectants were tested at their manufacturer's recommended concentration (MRC) and at three 10-fold dilutions of the MRC. The efficacy of disinfectants against I. 24 was assessed in conditions with no hatchery organic matter (HOM) and in conditions with no HOM present. In addition, 17 S. aureus strains, related and non-related to I. 24 and obtained from the poultry industry were screened for any increase in resistance relative to I. 24. All disinfectants were effective against all test strains when tested in the absence of HOM. Products from the QAC and peroxygen groups were the most potent. The performance of all disinfectants was reduced in the presence of HOM. Under these conditions all chlorine-based, two out of three AMP, and one out of eight QAC disinfectants were not effective against I. 24 when tested at the MRC. The results emphasise the importance of proper application on appropriate areas, using the correct concentration and exposure time for the disinfectant.

Examination of adenovirus-types in intestinal vascular endothelial inclusions in fatal cases of enteric disease in cattle, by in situ hybridisation.

There have been occasional reports in the literature of a severe naturally occurring enteric disease of cattle in which adenoviral inclusions were found in intestinal vascular endothelium. Bovine adenovirus type 10 (BAV-10) was identified by in situ hybridisation (ISH) in the inclusions of all 13 such cattle detected in Northern Ireland [Smyth, J.A., Benkö, M., Moffett, D.A., Harrach, B., 1996. J. Clin. Microbiol. 34, 1270-1274]. The present paper describes ISH examination of the vascular endothelial inclusion bodies in a further seventeen cattle with enteric disease, from Canada, Great Britain, Italy and The Netherlands. BAV-10 was identified in the inclusions of ten cases, and Subgroup 2 BAVs in six cases. BAV-10 was identified in cattle from Canada, The Netherlands and Great Britain. This is the first recognition of BAV-10 infection outside Northern Ireland and New Zealand. The results also show that at least two adenovirus serotypes may induce inclusion bodies in intestinal vascular endothelium of cattle. It will, therefore, be difficult in the short term to develop a simple test to allow diagnosis of this form of adenoviral infection in living animals, and thus to determine the relationship between it and clinical disease.

Evaluation of polymerase chain reaction and dot blot hybridisation tests in the diagnosis of pigeon circovirus infections.

Polymerase chain reaction (PCR) and dot blot hybridisation (DBH) tests for detecting pigeon circovirus (PiCV) DNA were developed and evaluated using tissue samples obtained from diseased and clinically normal pigeons, which originated in Belgium and Northern Ireland. When PCR product was visually detected, the limit of detection of the PCR test was 31 fg, while that of the DBH was 1.6p g. For evaluation purposes, the results of the PCR and DBH tests, performed with DNAs extracted from samples of bursa of Fabricius (BF), were compared with those of in situ hybridisation (ISH) and histology. In 32 samples tested by all four tests, 27 (84%) were positive by PCR, 24 (75%) were positive by ISH, 20 (63%) were positive by DBH, and 13 (41%) were positive by histology. Additional PCR testing showed that in some disease-affected birds, PiCV DNA could be detected in a range of tissues including thymus, spleen, liver, kidney and brain. The PCR detection of PiCV DNA in BF samples from clinically normal birds indicated that PCR can detect infections in the absence of disease, a finding that mitigates against its use as a disease diagnostic. In addition, nucleotide sequence determinations indicated that PCR test performance was adversely affected by the sequence diversity exhibited by selected PiCVs. The application of the DBH test to dilutions of test samples indicated that the BF from some diseased pigeons contained very large amounts of virus DNA, as much as 10(13)genome copies/g tissue, and suggested that this test may be a convenient method of providing a semi-quantitative estimate of virus load.