Analysis of breeding and pathology helps refine management practices of a large-scale N'-ethyl-N'-nitrosourea mouse mutagenesis programme.

N'-ethyl-N'-nitrosourea (ENU) is a powerful germline mutagen used in conjunction with phenotype-driven screens to generate novel mouse mutants. ENU also induces genetic lesions in somatic cells and dosage requires optimization between maximum germline mutation rate versus induced sterility and tumourigenesis that compromise the welfare and fecundity of the ENU-treated males. Here, we present our experience with BALB/cAnNCrl and C57BL/6J mice in terms of the pathology induced by ENU and its impact on breeding. In both mouse strains, morbidity and mortality rises with ENU dose. In more than 75% of C57BL/6J males, morbidity and mortality were attributable to the development of malignant T-lymphoblastic lymphoma. Approximately 50% of ENU-treated BALB/cAnNCrl males develop early malignant T-lymphoblastic lymphoma, but the cohort that survives develops late-onset lung carcinoma. Within strains, the latency of these clinically important tumour(s) was not dosage-dependent, but the proportion of mice developing tumours and consequently removed from the breeding programme increased with ENU dosage. The median number of offspring per ENU-treated C57BL/6J male in standard matings with C3H/HeH females decreased with increasing dosage. The two most important underlying causes for lower male fecundity were increased infertility in the highest dosage group and reduced numbers of litters born to the remaining fertile C57BL/6J males due to a higher incidence of morbidity. These findings have allowed us to refine breeding strategy. To maximize the number of offspring from each ENU-treated male, we now rotate productive males between two cages to expose them to more females. This optimizes the number of mutation carrying offspring while reducing the number of ENU-treated males that must be generated.

Upgrading mouse health and welfare: direct benefits of a large-scale rederivation programme.

We report the outcome of a 30-month programme to rederive 310 specific pathogen-free mouse strains to populate a new individually ventilated cage barrier facility at the Mary Lyon Centre (MLC), Medical Research Council (MRC) Harwell. The mice were rederived in a self-contained quarantine suite and embryo-recipient females were health-screened to assess microbiological status, before moving their offspring into the new facility. The MLC currently houses approximately 49,000 mice in about 9750 cages and we have 30 months of follow-up health screen data. Embryo rederivation and hysterectomy have high safety margins; however, the precaution of performing the programme in isolators facilitated the containment and decontamination of two mouse hepatitis virus (MHV) infection outbreaks. Rederivation of the colony has eliminated endemic MHV, mouse adenovirus type 2 (MAV-2), Theiler's murine encephalomyelitis virus, pinworms, intestinal protozoa, Pasteurella pneumotropica, Helicobacter spp. and mites. The improvements in microbiological status have had notable benefits for mouse health and welfare and the science at MRC Harwell. Previously important clinical entities such as sudden death associated with lactation ileus in C3H/HeH mice, early weight loss associated with inflammatory bowel disease in B6-TgN(HDexon1)61Gpb and B6TgN-(HD82Gln)81Dbo (Huntington) mice and early weight loss in male mice mutagenized with N-ethyl-N-nitrosourea have been markedly reduced or eliminated.

Ectopic stratified squamous epithelium in bovine jejunum.

An adult dairy cow developed acute fatal diarrhoea. Necropsy provided no aetiological or morphological cause of the diarrhoea but incidental post-mortem findings included well-circumscribed areas of jejunal squamous epithelium. The cause of this unusual heterotopia is not known, but the lesion may have been of embryological or metaplastic origin.

Preliminary characterisation of esterase and platelet-activating factor (PAF)-acetylhydrolase activities from cat flea (Ctenocephalides felis) salivary glands.

Naphthyl esterase and platelet-activating factor (PAF)-acetylhydrolase activities were detected in the salivary glands of the cat flea, Ctenocephalides felis. Salivary naphthyl esterase activity is disgorged during exploratory probing. Whole extracts of salivary glands contain esterase activity against the short-chain naphthyl esters alpha-naphthyl acetate (approximately 210pmol/min/gland pair; 10.0micromol/min/mg specific activity; K(m) approximately 59microM) and beta-naphthyl acetate (approximately 110pmol/min/gland pair; 5.2micromol/min/mg specific activity; K(m) approximately 132microM). Salivary gland extracts have PAF-acetylhydrolase activity (approximately 5pmol/min/gland pair; 0.24micromol/min/mg specific activity) but do not have detectable acetylcholinesterase activity. Native-PAGE and IEF resolve three and six salivary gland naphthyl esterase bands, respectively, and both patterns are different from carcass esterases. Salivary gland naphthyl esterase activity binds reversibly to Concanavalin A, and enzymatic deglycosylation with glycopeptidase F produced a new, fast-migrating salivary gland naphthyl esterase band on Native-PAGE. Renaturation of esterase activity after SDS-PAGE gave approximately 56kDa, approximately 57kDa and approximately 58kDa naphthyl-esterase-positive bands. On gel filtration naphthyl esterase and PAF-acetylhydrolase activities co-elute as a single peak with an apparent molecular weight of approximately 59kDa. This partially purified pool of enzyme had esterase activity against a series of short-chain alpha- and beta-naphthyl esters. The heterogeneity of salivary gland esterases, their relationship to PAF-acetylhydrolase, and the possible physiological functions of salivary gland PAF-acetylhydrolase activity are discussed.

Anaemia, diarrhoea and opportunistic infections in Fell ponies.

This report summarises clinical and pathological observations on Fell pony foals with a range of signs that included ill thrift, anaemia, respiratory infection, glossal hyperkeratosis and diarrhoea. Some of the foals had normochromic, normocytic anaemia and some had low levels of plasma proteins, including immunoglobulin G. Antibiotic and supportive treatment was ineffective and all affected foals died or were killed on humane grounds. Postmortem examination of 12 foals and tissues from 2 other foals revealed a range of lesions that included glossal hyperkeratosis, typhlocolitis, intestinal cryptosporidiosis, granulomatous enteritis, proliferative and necrotising bronchiolitis consistent with adenovirus infection; lesions similar to those in the respiratory tract were present in the salivary gland and pancreas of individual foals. Lymphoid tissue was judged to be smaller than expected. These observations suggest the possibility of opportunistic infections secondary to some form of undefined immunocompromised state.

Resection of the common digital extensor tendon in a gelding.

A four-year-old gelding was lame owing to a chronic septic common digital extensor tendon and sheath. The horse had been treated by open surgical lavage but the sepsis had recurred after three months. Physical, ultrasonographic, cytological and histological examinations confirmed chronic septic tenosynovitis and tendonitis. The entire intrathecal component of the common digital extensor tendon was resected under general anaesthesia and the synovial lining of the sheath was ablated. Postoperatively the horse regained good limb function and became sound.

Genitourinary dysplasia in a cat.

A six-month-old kitten had congenital urethral sphincter mechanism incompetence due to urethral hypoplasia and associated uterine hypoplasia and vaginal aplasia. Diagnosis was based on radiographic examination, surgical exploration and histological examination of the lower urinary tract. Surgical correction resulted in a marked clinical improvement. The cat became fully continent following treatment with phenylpropanolamine.

Heterotopic pancreas at a site of colon stricture and ulceration in a guineapig.

A one-month-old female Dunkin Hartley guineapig presented with a thin hair coat and distended abdomen. Post-mortem findings after euthanasia were a protein-rich modified peritoneal transudate and heterotopic pancreas at a site of colon stricture and ulceration. Complications of pancreatic heterotopia are well recognized in man but not in animals. The finding in this individual is probably unrelated to sudden deaths in other animals in the group.

Characterization of apyrase activity from the salivary glands of the cat flea Ctenocephalides felis.

Apyrase activity (ATP diphosphohydrolase, EC 3.6.1.5) was detected in salivary glands of the cat flea Ctenocephalides felis. Whole extracts of salivary glands contain approximately 21 ng of protein, 145 U/mg ADP'ase and 158 U/mg ATP'ase activity; AMP is not hydrolysed by salivary gland extracts. DEAE-Sepharose CL-6B anion exchange chromatography, and Cibacron Blue affinity chromatography each give a single coincident peak of ADP/ATP'ase activity. Biogel P-100 gel filtration of salivary gland homogenates made in buffer containing Triton and protease inhibitors, separated enzymatic activity into 57 kD and 44 kD peaks of ADP/ATP'ase activity. Partially purified ADP/ATP'ases are dependent on divalent cations and activation increases between 0.125 mM and 5.0 mM calcium. At 5 mM, magnesium is almost equally effective as calcium in activating ADP/ATP'ase but manganese and zinc are less so, and EDTA abolishes activity. ADP/ATP'ases have a pH optima of 7-9. The Km for ADP hydrolysis by whole extracts and partially purified enzyme is approximately 66 microM ADP. The co-purification of ADP'ase and ATP'ase activity by three physiochemical techniques and parallelism between ADP and ATP hydrolysis under varying conditions of pH and activating cation indicates enzymatic activity is attributable to true apyrase(s).

Heterotopic neural tissue in the pharynx of a 7-week-old kitten.

A 7-week-old male kitten had a pharyngeal mass (1 x 2 cm) causing displacement of the tongue. The surgically resected tissue was seen to be a moderately discrete subepithelial mass comprising islands of neuroglia and neurons separated by dense collagenous connective tissue. It is not known whether this mass retained any connection with the brain. Histochemical and immunohistochemical examination confirmed the presence of neurons and a pleocellular glial population, supporting a diagnosis of heterotopic neural tissue. The cat remained well 20 months after surgical treatment. Heterotopic neural tissue is well-recognized in man but has not been described in animals.

Esophagitis due to a herpesvirus associated with mortality in a psittacine aviary.

In a disease outbreak at a psittacine aviary, eight Pionus sp. and Amazon parrots became sick. Three of four Amazon parrots died with severe esophagitis, and epithelial intranuclear inclusion bodies containing herpesvirus were prominent. Focal liver necrosis with rare hepatocyte intranuclear inclusion bodies were found in two of the dead Amazon parrots.

Multisystemic inflammatory disease in a borzoi dog.

A six-week-old female borzoi puppy from a brother-sister mating developed a generalised illness characterised by anorexia, temporary intention tremor, episodic pyrexia, tachypnoea, conjunctivitis, otitis and neck pain. Haematological abnormalities included an inflammatory leukogram and regenerative anaemia. Blood cultures remained sterile; clinical chemistry values were unremarkable. The puppy had recurrent seizures and was euthanased when 18 weeks old. Post mortem examination revealed a multisystemic inflammatory disease involving thyroids, lymph nodes, spleen, pancreas, bladder and lung, but no lesions to account for the neurological signs. The cause of this generalised disease was not recognised. The histological features are unusual and resemble those described in other dogs of this breed.

Corpus allatum and corpus cardiacum regulation of long hyaline gland protein synthesis in the male grasshopper, Melanoplus sanguinipes.

Rates of accumulation of, and radiolabel incorporation into, the major secretory product of the long hyaline gland, a glycoprotein LHPI, were used to examine the role of the corpus allatum (CA) and corpus cardiacum (CC) in the regulation of protein synthesis in the accessory reproductive gland of the male grasshopper, Melanoplus sanguinipes. Incorporation of [35S]methionine into LHPI in vitro indicated that LHPI was an endogenous long hyaline gland protein, whose in vivo synthesis in allatectomized animals was stimulated by topical application of juvenile hormone (JH) III. This response was not dependent on the presence of the extrinsic lobe of the CC, the CC/brain complex, or on continued feeding and suggests a direct effect of JH on the gland. Severance of the nervi corporis allati I and II reduced accumulation of LHPI but unilateral CA removal did not, perhaps because of a compensatory activation of the remaining CA. Normal postcopulatory stimulation of LHPI accumulation was abolished by allatectomy. These results suggest that the physiological level of JH may regulate LHPI synthesis. Corpus cardiacum effects were small in comparison to those of the CA; they were evident only in cardiacectomized animals that were allatectomized also and exerted over prolonged periods when the animals continued to feed. Endocrine manipulations produced no qualitative changes in long hyaline gland LHPI immunoreactive protein, only quantitative ones.