Objective—To use PCR assays to determine the prevalence of feline herpesvirus 1 (FHV-1), Chlamydophila felis, and Mycoplasma spp DNA in conjunctival cells collected from cats with and without conjunctivitis; to compare results of conventional and real-time fluorogenic PCR assays for amplification of FHV-1 DNA; and to determine whether copy numbers of FHV-1 DNA are correlated with conjunctivitis.
Animals—55 cats with active conjunctivitis, 39 healthy cats that never had conjunctivitis, and 32 cats with a history of conjunctivitis that had been resolved for at least 3 months.
Procedures—Samples were obtained by rolling cotton-tipped applicators on the ventral conjunctiva of awake cats treated topically with proparacaine. The DNA was extracted from the swab specimens and assessed in PCR assays to detect DNA of FHV-1 (fluorogenic PCR assay and conventional PCR assay), Mycoplasma spp (conventional PCR assay), and C felis (conventional PCR assay).
Results—Overall prevalence rates of FHV-1, C felis, and Mycoplasma spp as assessed by the conventional PCR assays were 6.7%, 3.2%, and 9.6%, respectively. Percentage concordance between conventional PCR and fluorogenic PCR assays for FHV-1 was 92.5%. There were no significant differences among the 3 groups of cats for the mean copy number of FHV-1 divided by the copy number of glyceraldehyde-3-phosphate dehydrogenase.
Conclusions and Clinical Relevance—Mycoplasma spp were the most prevalent organism detected and was associated with conjunctivitis. This study could not confirm that there are increased copy numbers of FHV-1 DNA in cats with conjunctivitis, compared with the copy numbers for cats without conjunctivitis.
Objective—To determine whether Ctenocephalides
felis can transmit Mycoplasma haemofelis (Mhf) and
Candidatus Mycoplasma haemominutum (Mhm)
through hematophagous activity between cats.
Procedure—2 cats were carriers of either Mhf or
Mhm. Nine cats had negative results via polymerase
chain reaction (PCR) assay for Mhf and Mhm DNA; 3
of those cats were infected from the chronic carriers
via IV inoculation of blood. At the time of maximum
organism count for each of the Mycoplasma spp, 1
chamber containing 100 C felis was bandaged to the
amplifier cats. Five days later, fleas, feces, larvae, or
eggs from each chamber were analyzed for
Mycoplasma spp DNA. Viable fleas from the chambers
were allocated into new chambers (3 Mhm and
6 Mhf) and attached to naïve cats for 5 days. Cats
were monitored daily for clinical signs and weekly via
CBC and PCR assay for infection with Mhf or Mhm for
a minimum of 8 weeks.
Results—Uptake of Mhf and Mhm DNA into fleas,
feces, and, potentially, eggs and larvae was detected.
Of the naïve cats fed on by Mhf-infected fleas, 1 cat
transiently yielded positive PCR assay results for Mhf
on 1 sampling date without clinical or hematologic
changes consistent with Mhf infection.
Conclusions and Clinical Relevance—Results suggest
that hematophagous transfer of Mhm and Mhf
into fleas occurred and that C felis is a possible vector
for Mhf via hematophagous activity. (Am J Vet Res 2005;66:1008–1012)
To assess whether hyperinoculation of cats with a feline herpesvirus-1, calicivirus, and panleukopenia virus (FVRCP) vaccine could be used as a model to study interstitial nephritis and to assess humoral and cell-mediated immune responses toward vaccinal α-enolase.
6 healthy young adult purpose-bred research cats.
Baseline renal cortical biopsies, whole blood, serum, and urine were collected prior to administration of a commercial FVRCP parenteral vaccine. Vaccine hyperinoculation was defined as a total of 8 vaccinations given at 2-week intervals over a 14-week period. Blood samples were collected immediately prior to each vaccination, and a second renal biopsy was performed 2 weeks after hyperinoculation (week 16). Renal histopathology, renal α-enolase immunohistochemistry, and assays to detect humoral and cell-mediated immune reactions against Crandell-Rees feline kidney (CRFK) cell lysates and α-enolase were performed. An α-enolase immunoreactivity score for renal tubules and glomeruli based on signal intensity was determined by a blinded pathologist.
Hyperinoculation with the vaccine was not associated with clinicopathologic evidence of renal dysfunction, and interstitial nephritis was not recognized by light microscopy in the time studied. The mean serum absorbance values for antibodies against CRFK antigen and α-enolase were significantly (P < 0.001) higher at weeks 4, 8, and 16 versus week 0. Renal tubular and glomerular α-enolase immunoreactivity scores were higher at week 16 compared to baseline.
Findings suggested that systemic immunological reactions occurred and renal tissues were affected by vaccine hyperinoculation; however, short-term FVRCP vaccine hyperinoculation cannot be used to study interstitial nephritis in cats.
Objective—To develop a broad-range 28S ribosomal DNA quantitative PCR (qPCR) assay for detection of fungal DNA in equine endometrial samples.
Sample—12 fungal samples from a clinical diagnostic laboratory and 29 samples obtained from 17 mares.
Procedures—The qPCR assay was optimized with commercially acquired fungal organisms and validated with samples obtained from the clinical diagnostic laboratory. Subsequently, 29 samples from 17 mares suspected of having fungal endometritis were evaluated via the qPCR assay and via traditional fungal culture and endometrial cytology. Amplicons from the qPCR assay were subjected to genetic sequencing to identify the organisms.
Results—The qPCR assay theoretically had a detection threshold of 2 organisms of Candida albicans. Fungal DNA was amplified from all 12 fungal samples from the commercial diagnostic laboratory. Fungal identification by use of genetic sequencing was successful for 34 of 36 amplicons from the 12 samples assayed. A fungal agent was identified via qPCR assay and genetic sequencing in all 12 samples; in contrast, a fungal agent was identified in only 8 of 12 samples via standard fungal culture and biochemical analysis. The qPCR assay detected fungal DNA in samples from 12 of 17 mares suspected of having fungal endometritis.
Conclusions and Clinical Relevance—A rapid, sensitive, and repeatable qPCR assay was developed for detection of fungal DNA from equine endometrial samples. The qPCR may prove to be clinically useful as an adjunct to microbial culture and cytologic examination to provide identification of fungal organisms in a timely manner.
Objective—To evaluate the efficacy of twice-daily ophthalmic application of 0.5% cidofovir solution in cats with experimentally induced primary ocular feline herpesvirus-1 (FHV-1) infection.
Animals—Twelve 6-month-old sexually intact male cats.
Procedures—Cats were randomly assigned to either a treatment or control group. Ocular infection with FHV-1 was induced (day 0) in all cats via inoculation of both eyes with 104 plaque-forming units of a plaque-purified FHV-1 field strain. Twice daily for 10 days beginning on day 4 after virus inoculation, the treatment group received 1 drop of 0.5% cidofovir in 1% carboxymethylcellulose in both eyes, and the control group received 1 drop of 1% carboxymethylcellulose in both eyes. A standardized scoring method was used to evaluate clinical signs of FHV-1 infection in each cat once daily for 24 days. The amount of ocular viral shedding was assessed by use of a quantitative real-time PCR procedure every 3 days during the study period. Clinical scores and viral quantification were averaged over the pretreatment (days 0 to 3), treatment (days 4 to 14), and posttreatment (days 15 to 24) periods for each cat.
Results—During the treatment period, clinical scores and amount of viral ocular shedding were significantly lower in the treatment group, compared with findings in the control group.
Conclusions and Clinical Relevance—Twice-daily application of 0.5% cidofovir solution in both eyes significantly decreased the amount of viral shedding and the severity of clinical disease in cats with experimentally induced ocular FHV-1 infection.
To determine hepatic copper concentrations and zonal distribution in ferrets with and without hepatobiliary disease, validate rhodanine-based qualitative copper scoring and digital copper quantification in ferret hepatic samples, and ascertain whether clinical features predicted copper accumulation.
34 ferrets, including 7 with necroinflammatory disease, 5 with hepatocellular carcinoma, 13 with non-necroinflammatory disease, and 9 with no hepatobiliary disease.
Rhodanine-based digital copper quantification was validated by use of liver dually measured by atomic absorption spectroscopy and digital scanning (R2 = 0.98). Clinical features and hepatic copper scores and concentrations (dry weight liver) were compared between groups. Zonal copper distribution was determined.
Hepatic copper concentration was strongly correlated with copper scores (ρ = 0.88). Ferrets with hepatobiliary disease were significantly older and had significantly higher serum alkaline phosphatase and γ-glutamyltransferase activities and creatinine concentrations. Centrilobular copper accumulated in 23 of 34 (64%) ferrets with (n = 15) and without (8) hepatobiliary disease. Median copper concentrations were not significantly different between ferrets with and without hepatobiliary disease but were significantly higher within neoplastic hepatic tissue in ferrets with hepatocellular carcinoma. Hepatic copper concentrations exceeded feline (> 180 µg/g) and canine (> 400 µg/g) reference limits in 19 and 9 ferrets, respectively. Hepatic copper > 1,000 µg/g occurred in 5 ferrets with and 2 without hepatobiliary disease. Clinical features did not predict copper accumulation.
Rhodanine-based digital copper quantification and qualitative copper scoring discerned liver copper accumulation in ferrets. Ferrets with and without hepatobiliary disease displayed a propensity for centrilobular hepatic copper accumulation of uncertain clinical importance. Clinical and clinicopathologic features could not exclusively implicate pathologic copper accumulation.
Objective—To determine whether administration of
Crandell-Rees feline kidney (CRFK) cell lysates or vaccines
against feline viral rhinotracheitis, calicivirus,
and panleukopenia (FVRCP vaccines) that likely contain
CRFK cell proteins induces antibodies against
CRFK cell or feline renal cell (FRC) lysates in cats.
Animals—14 eight-week-old cats.
Procedure—Before and after the study, renal biopsy
specimens were obtained from each cat for histologic
evaluation. Each of 4 FVRCP vaccines was administered
to 2 cats at weeks 0, 3, 6, and 50. Between
weeks 0 and 50, another 3 pairs of cats received 11
CRFK cell lysate inoculations SC (10, 50, or 50 µg
mixed with alum). Clinicopathologic evaluations and
ELISAs to detect serum antibodies against CRFK cell
or FRC lysates were performed at intervals.
Results—Cats had no antibodies against CRFK cell
or FRC lysates initially. All cats administered CRFK
cell lysate had detectable antibodies against CRFK
cell or FRC lysates on multiple occasions. Of 6 cats
vaccinated parenterally, 5 had detectable antibodies
against CRFK cell lysate at least once, but all 6 had
detectable antibodies against FRC lysate on multiple
occasions. Cats administered an intranasal-intraocular
vaccine did not develop detectable antibodies
against either lysate. Important clinicopathologic or
histologic abnormalities were not detected during
Conclusions and Clinical Relevance—Parenteral
administration of vaccines containing viruses likely
grown on CRFK cells induced antibodies against
CRFK cell and FRC lysates in cats. Hypersensitization
with CRFK cell proteins did not result in renal disease
in cats during the 56-week study. (Am J Vet Res 2005;66:506–511)
OBJECTIVE To evaluate the pharmacokinetics of zonisamide following rectal administration of 20 or 30 mg/kg suspended in sterile water or polyethylene glycol (PEG) to healthy dogs and determine whether either dose resulted in plasma zonisamide concentrations within the recommended therapeutic target range (10 to 40 μg/mL).
ANIMALS 8 healthy mixed-breed dogs.
PROCEDURES Each dog received each of 2 doses (20 or 30 mg/kg) of zonisamide suspended in each of 2 delivery substrates (sterile water or PEG) in a randomized crossover study with a 7-day washout period between phases. A blood sample was collected from each dog immediately before and at predetermined times for 48 hours after zonisamide administration. Plasma zonisamide concentrations were determined by high-performance liquid chromatography, and data were analyzed with a noncompartmental model.
RESULTS Mean maximum plasma concentration, time to maximum plasma concentration, mean residence time, and elimination half-life did not differ significantly among the 4 treatments. The mean maximum plasma concentration for all 4 treatments was less than the therapeutic target range. The mean ± SD area under the concentration-time curve for the 30 mg/kg-in-water treatment (391.94 ± 237.00 h•μg/mL) was significantly greater than that for the 20 mg/kg-in-water (146.19 ± 66.27 h•μg/mL) and 20 mg/kg-in-PEG (87.09 ± 96.87 h•μg/mL) treatments.
CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that rectal administration of zonisamide at doses of 20 and 30 mg/kg failed to achieve plasma zonisamide concentrations within the recommended therapeutic target range. Therefore, rectal administration of zonisamide cannot be recommended as a suitable alternative to oral administration.