Detection of feline calicivirus, feline herpesvirus 1 and Chlamydia psittaci mucosal swabs by multiplex RT-PCR/PCR.

A single tube, multiplex reverse transcription (RT)-polymerase chain reaction (PCR)/PCR assay was developed for detection of feline herpesvirus 1 (FHV1), Chlamydia psittaci and feline calicivirus (FCV) in cats with upper respiratory tract disease (URTD), incorporating a simple, rapid extraction procedure capable of extracting both DNA and RNA. The assay was found to be as sensitive in vitro as simplex assays that have previously been shown to be as sensitive as, or more sensitive than, culture for each pathogen in experimentally infected cats. Conjunctival alone or both conjunctival and oropharyngeal swabs were collected from cats in 104 households with URTD. FHV1 was detected in 18 (17.3%) and C. psittaci was detected in 12 (11.5%) households. The prevalence of C. psittaci was not significantly different to that determined using a duplex PCR assay for C. psittaci and FHV1. The prevalence of FCV was affected by sample storage temperature. Of samples stored at -70 degrees C, 0/31 were positive for FCV but FCV was detected in 10/73 (13.7%) samples stored at 4 degrees C (P=0.006). Of the samples stored at 4 degrees C, 3/19 (15.8%) conjunctival swabs were positive for FCV and 6/32 (18.8%) oropharyngeal/conjunctival swabs were positive for FCV (P=0.79). The potential utility of restriction endonuclease analysis of RT-PCR products resulting from amplification of the hypervariable region of the capsid protein gene of FCV in field samples, without prior cultivation, was also examined. The assay may have considerable importance for diagnosis and epidemiological surveys of feline upper respiratory tract pathogens.

A new disorder of hyaluronan metabolism associated with generalized folding and thickening of the skin.

OBJECTIVE: To describe and characterize a new disorder of hyaluronan metabolism associated with marked abnormalities of cutaneous tissue and to determine whether a relationship with a phenotypically similar disorder in the shar-pei dog exists. METHODS: Biopsy specimens of the skin of a child with extreme cutaneous thickening and folding were examined by light and electron microscopy. The concentration of hyaluronan and the activity of hyaluronidase were measured in the patient's serum and plasma, respectively, and the activity of hyaluronan synthase was examined in cultured dermal fibroblasts. Hyaluronan concentration was also measured in the plasma of 23 shar-pei and 34 control dogs. RESULTS: The patient's skin displayed gross accumulation of hyaluronan, and the serum concentration of hyaluronan was markedly elevated (up to 3100 microg/L) during infancy. Hyaluronan synthase activity of cultured dermal fibroblasts was increased, whereas hyaluronidase activity in plasma was normal (5.5 +/- 0.08 IU/L). Plasma hyaluronan concentration was higher in the shar-pei dogs than in control dogs (median, 378 microg/L vs 73 microg/L, respectively). CONCLUSION: The child we describe has a novel disorder of hyaluronan metabolism, which appears to result from abnormal control of hyaluronan synthesis. An analogous disorder may be present in the shar-pei dog.

Comparison of the polymerase chain reaction and culture for the detection of feline Chlamydia psittaci in untreated and doxycycline-treated experimentally infected cats.

The diagnostic sensitivity of the polymerase chain reaction (PCR) was compared with that of culture on conjunctival swabs over the course of infection in 4 doxycycline-treated and 4 untreated cats that were experimentally infected with feline Chlamydia psittaci. Treated cats were given 25 mg (5 mg/kg) of doxycycline orally twice daily for 3 weeks from day 6 after challenge. Clinical signs improved within 3 days of institution of treatment. Culture remained positive for 1 day and PCR remained positive for up to 5 days after treatment was commenced. No recurrence of clinical signs occurred and the organism could not be detected by either PCR or culture for 2 weeks after cessation of therapy. In the 4 untreated cats, conjunctival swabs were taken daily to day 14 and every 2nd weekday to day 64 after challenge. PCR was significantly more sensitive than culture in untreated cats overall (PCR 85.7%, culture 72.9%, P approximately 0) and for cats with clinical signs (PCR 89.2%, culture 79.2%, P = .008). PCR and culture had equivalent sensitivity (100%) for cats showing clinical signs in the 1st month of infection, whereas PCR was considerably more sensitive than culture for cats showing clinical signs in the 2nd month (PCR 72.9%, culture 47.9%, P = .028). Organisms were not detected by PCR in blood or any tissue collected from treated or untreated cats at postmortem. Thus, effective treatment of chlamydiosis in cats is possible with much shorter treatment regimens than currently recommended, and PCR is the more sensitive diagnostic method in chronically infected cats.

Prevalence of feline Chlamydia psittaci and feline herpesvirus 1 in cats with upper respiratory tract disease.

The epidemiology of feline chlamydiosis and feline herpesvirus 1 (FHV1) infection in cats was determined using a duplex polymerase chain reaction assay. In cats with upper respiratory tract disease (URTD), prevalences of 66 (14.3%) of 462 cats and 98 (21.2%) of 462 cats were found for Chlamydia psittaci and FHV1, respectively. In cats without URTD, prevalences were 1/87 (1.1%) for both pathogens. Younger cats, cats sampled in summer, and cats with conjunctivitis were more likely to be positive for C psittaci than were cats sampled in other seasons and cats without conjunctivitis. Cats with recent contact with cats outside the household, cats with acute disease, and sneezing cats were more likely to be positive for FHV1 than were cats that had not had recent contact with cats outside the household, cats with chronic disease, and cats that were not sneezing. Purebred cats were less likely to be positive for FHV1 than were mixed breed cats and prevalence varied with year of sampling. Coinfection with both pathogens was lower than would be expected from their respective prevalences. Vaccinated cats were equally likely to be positive for FHV1 as unvaccinated cats. In sneezing cats FHV1 was more likely to be detected than C psittaci, particularly in acute cases, and when sneezing was not accompanied by conjunctivitis. Cats with reproductive disease concurrent with URTD were more likely to be infected with FHV1 than with C psittaci. Thus, the factors that should be considered in clinical diagnoses of C psittaci infections are the presence of conjunctivitis, age, and season, whereas contact with other cats, acute disease, and sneezing should be considered in diagnoses of FHV1 infection.

Detection and strain differentiation of feline calicivirus in conjunctival swabs by RT-PCR of the hypervariable region of the capsid protein gene.

Reverse transcription-polymerase chain reaction (RT-PCR) was used to direct the amplification of a 670 to 680 base pair segment that included the hypervariable regions of the capsid protein gene of feline calicivirus (FCV). The segment was amplified from 13/13 cultivated FCV strains including 12 field isolates collected over 18 years. The sensitivities of culture and this RT-PCR for the detection of FCV in conjunctival swabs over the course of infection were then compared and correlated with clinical signs in 5 vaccinated and 3 unvaccinated experimentally-infected cats. Conjunctival swabs were taken daily from days 0 to 14 and on days 16, 19, 21 and 24 after challenge. FCV was detected in 19/144 swabs by RT-PCR and 16/144 swabs by culture. Virus detection correlated poorly with clinical signs regardless of the assay used. The Sau3AI restriction profiles of the RT-PCR products amplified from both cultivated strains and clinical samples were compared. All 13 cultivated isolates, including the 12 field isolates, exhibited different profiles, whereas all profiles from the experimentally-infected cats were identical, and matched the profile of the challenge/vaccine strain. This study has established that the RT-PCR assay described is as sensitive as culture for detection of FCV in conjunctival swabs, that a broad range of field isolates can be detected and rapidly differentiated by restriction endonuclease digestion, and that the assay thus meets the requirements for large scale epidemiological studies of FCV infections in cats. However, it has also shown that even the use of RT-PCR on conjunctival swabs alone is insufficient for accurate diagnosis of FCV infection in cats with conjunctivitis.

Comparison of Chlamydia psittaci from cats with upper respiratory tract disease by polymerase chain reaction analysis of the ompA gene.

Conjunctival swabs were taken from 168 cats with clinical signs of acute or chronic upper respiratory tract disease and tested for Chlamydia psittaci by the polymerase chain reaction (PCR) to amplify the ompA gene coding region. Twenty-two (13 per cent) were positive for C psittaci. The PCR products from positive samples were subjected to restriction endonuclease analysis with the restriction enzymes Alu I and Mse I. The fragments of DNA were detected on silver-stained polyacrylamide gels and the results were compared with the results obtained from chlamydial isolates from cats in Japan, France, the USA and the UK. All the strains had identical restriction patterns. When PCR is used as an epidemiological tool, feline chlamydial strains worldwide appear very similar.

Differential sensitivity of culture and the polymerase chain reaction for detection of feline herpesvirus 1 in vaccinated and unvaccinated cats.

The diagnostic sensitivities of the polymerase chain reaction (PCR) and culture were compared and correlated with clinical signs in 5 vaccinated cats and 3 unvaccinated cats that were experimentally infected with feline herpesvirus 1. Conjunctival swabs were taken each day from 0 to 14 days and on 21, 28 and 30 days after challenge. PCR (49.3%) was significantly more sensitive than culture (30.1%) as assessed by an adjusted McNemar's test to account for non-independence of results between days within each cat (P = 0.02). PCR was considerably more sensitive (34.1%) than culture (8.2%) in vaccinated cats (P = 0.001), whereas there was no significant difference in sensitivities in the unvaccinated cats, where the sensitivity of PCR was 74.5% and that of culture was 66.7% (P = 0.17). In vaccinated cats showing clinical signs, the sensitivities of culture and PCR were 14.8% and 55.6% respectively (P = 0.03), whereas in unvaccinated cats the sensitivities were 80.6% and 96.8% respectively (P = 0.07). This study suggests that disease due to feline herpesvirus 1 has been significantly underdiagnosed, particularly in vaccinated cats.

Treatment of opportunistic mycobacterial infections with enrofloxacin in cats.

Marked improvement was observed in the condition of 6 cats with opportunistic mycobacterial infections during treatment with enrofloxacin, a fluoroquinolone antibiotic. Complete remission was achieved in 3 cats after 3 to 7 weeks of treatment. The other 3 cats were euthanatized after 1 to 2 weeks of treatment for reasons not related to the treatment. Lesions did not recur within the follow-up period, which ranged from 9 to 16 months. Treatment of opportunistic mycobacterial infection in cats is complicated because many mycobacteria are resistant to antituberculosis drugs, which also can be toxic to cats, and because results of susceptibility testing with other antimicrobials do not always correlate with clinical response. Often, neither satisfactory nor long-term response is observed in cats treated surgically or with the antibiotics currently recommended. These findings suggested that enrofloxacin is effective in the treatment of infections caused by Mycobacterium smegmatis and M fortuitum var fortuitum in cats.

A clinical trial of a topical preparation of miconazole, polymyxin and prednisolone in the treatment of otitis externa in dogs.

A topical preparation containing miconazole, polymyxin and prednisolone was shown to be more effective in the treatment of otitis externa in 167 dogs than 2 other ear preparations containing antibiotics, an antimycotic and a corticosteroid. With miconazole, polymyxin and prednisolone, the recurrence rate was 26.7% compared with 72.6% and 54.3% when the other products were used. The mean duration of treatment required to achieve resolution of clinical signs was 9.6 days, compared with 12.2 days and 13.0 days and no cases failed to respond to treatment, compared with 17.7% and 14.3%. Malassezia canis alone (71%) or in association with bacteria (18%) was recovered from 44 of 49 ears cultured.

Clinical features of ceroid lipofuscinosis in border collie dogs.

Ceroid lipofuscinosis was diagnosed by histopathological and histochemical findings in 17 related border collie dogs and by clinical signs in 6 of their litter mates. Behavioural changes, first hyperactivity and later aggression, commenced at 16 to 23 (mean 19.5) months of age. Motor abnormalities and blindness were observed at the mean ages of 20.8 and 21.2 months, respectively. All dogs were euthanased 1 to 6 months after the onset of clinical signs, mean age 23.1 months. Pedigree data supported an autosomal recessive mode of inheritance.

Thiamin deficiency in cats and dogs associated with feeding meat preserved with sulphur dioxide.

Thiamin deficiency was diagnosed in cats and dogs being fed fresh minced meat, which contained sulphur dioxide as a preservative and less than 0.5 mg/kg thiamin. Thiamin in the meat and in added dietary ingredients, including a supplementary vitamin mixture, was destroyed by the sulphur dioxide.

Epidemiological features of snail and slug bait poisoning in dogs and cats.

Snail and slug baits were the most common cause of poisoning in dogs and cats in a survey of 34 veterinary practices. During a 7-month period, 280 dogs and 12 cats were poisoned by metaldehyde (57%) or methiocarb (43%) containing baits with case fatality rates of 8.1% and 9.1% in dogs and 16.7% and 50% in cats, respectively. The ages of poisoned dogs ranged from 2 months to 17 years and cats from 3 months to 5 years. There was no decrease in frequency of poisoning or case fatality rate demonstrated with increasing body size in dogs. Poisoning was most frequent in the month of October. Baits were eaten off the ground or obtained from storage areas, often despite a variety of preventive measures. It is suggested that these products should be reformulated to make them less attractive to domestic pets.

Inherited copper toxicosis in Bedlington terriers.

Chronic hepatitis and increased hepatic copper concentrations, from 1,600 to 6,361 micrograms/g dry tissue were found in 4 related, Australian-bred Bedlington terriers. Two dogs were asymptomatic and 2 were clinically ill with signs referable to liver dysfunction. Two dogs were treated with d-penicillamine. After one year there was no improvement in the histopathological liver changes in either dog or significant lowering of hepatic copper level in one dog.