Evaluation of feline heartworm disease based on gross necropsy, serology, pulmonary histopathology, and radiographic evidence in adult shelter cats in northeastern Alabama.

BACKGROUND: Veterinary knowledge regarding feline heartworm has been increasing significantly over the past two decades. Necropsy surveys of shelter cats have shown feline adult heartworm infection prevalence to be 5-20% of the rate in unprotected dogs; however, other studies have shown feline heartworm antibody prevalence up to 33%, reflecting higher exposure rates and potential immature adult infections. Thus, the true prevalence of feline heartworm infection is likely underestimated due to the limitations of current diagnostic techniques, inadequate testing protocols, and the high likelihood of cats exhibiting transient clinical signs or dying without confirmation of infection. Diagnosing Feline Heartworm Disease (FHWD), also referred to as Heartworm Associated Respiratory Disease (HARD), is one of the conundrums of veterinary medicine. The purpose of this study was to evaluate and characterize the occurrence of Heartworm Associated Respiratory Disease [HARD] in shelter cats, naturally-infected with D.immitis. METHODS: Fifty shelter cats slated for euthanasia between December 2009 and June 2010 were investigated by gross necropsy, radiography, serology, and lung histopathology using techniques that have been established in experimental models of cat heartworm infection. The relationship between pulmonary vascular disease and serological markers for heartworm was also examined using correlations and statistical modeling. Serology included standard heartworm antigen test and a commonly used heartworm antibody test. Also included were heat-treated heartworm antigen test and two additional heartworm antibody tests previously evaluated on experimentally-infected cats. RESULTS: None of the cats were heartworm antibody (HW Ab) positive on a commonly used HW Ab test used by many reference laboratories even though 20% of the study cats were heartworm antigen (HW Ag) positive on heat-treated samples. Two additional HW Ab test were positive on 26% and 22% of the study cats. The combination of heat-treated HW Ag, HW Ab tests, and histopathology indicated 34% of the study cats had HARD. CONCLUSIONS: Utilizing both, the above tests, and thoracic radiographs, enhanced the ability to predict vascular disease, possibly caused by infection with immature and adult heartworms and supported the premise that cats develop heartworm disease at the same rate as dogs.

Evaluation of the contribution of gyrA mutation and efflux pumps to fluoroquinolone and multidrug resistance in pathogenic Escherichia coli isolates from dogs and cats.

OBJECTIVE: To investigate the contribution of gyrA mutation and efflux pumps to fluoroquinolone resistance and multidrug resistance among Escherichia coli isolates from dogs and cats. SAMPLE POPULATION: 536 clinical isolates of E coli. PROCEDURES: Minimum inhibitory concentrations (MICs) were determined for enrofloxacin and 6 other drug classes by use of broth microdilution techniques. Real-time PCR assay was used to determine the mutation in gyrA; Phe-Arg-ß-naphthylamide, an efflux pump inhibitor, was used to examine the contribution of efflux pump overexpression. RESULTS: The MIC for fluoroquinolones increased in a stepwise fashion and was lowest in the absence of mutations, higher with a single point mutation, and highest with 2 point mutations. Level of resistance in the latter category was high (8 times the breakpoint), but this was associated with expression of the AcrAB efflux pump. Inhibition of the efflux pump resulted in a reduction in the MIC to less than the susceptible breakpoint for isolates with an MIC ≤ 4 mg/L, regardless of the presence of a mutation. The greatest magnitude in MIC decrease (MIC was decreased by a factor of > 67 fold) was for isolates with a single mutation but the greatest absolute decrease in MIC (124 mg/L) was for isolates with 2 mutations. Inhibition of the AcrAB efflux pump in isolates characterized by multidrug resistance decreased the MIC of drugs structurally unrelated to fluoroquinolone. CONCLUSIONS AND CLINICAL RELEVANCE: Fluoroquinolone resistance in E coli appeared to be a stepwise phenomenon, with MIC increasing as the number of point mutations in gyrA increased, but high-level resistance and multidrug resistance associated with fluoroquinolone resistance reflected overexpression of the AcrAB efflux pump.

Dual-emission fluorescence resonance energy transfer (FRET) real-time PCR differentiates feline immunodeficiency virus subtypes and discriminates infected from vaccinated cats.

Feline immunodeficiency virus (FIV) is among the most common infectious agents of cats. Five well-characterized FIV subtypes, A, B, C, D, and E, are recognized worldwide. As in HIV diagnosis, serum antibodies against FIV classically serve as an indicator of infection status. After the introduction of an inactivated FIV vaccine, this approach has become problematic, since antibodies generated by vaccination are indistinguishable from antibodies in response to infection. However, PCR detection of host-cell-integrated FIV DNA will differentiate infection-derived antibody from vaccination-derived positivity because presumably the RNA of inactivated vaccine virus will not integrate into the host genome. In this study, we established a gag gene-based dual-emission fluorescence resonance energy transfer (FRET) real-time PCR that amplifies single-target copies of all known FIV strains and differentiates five FIV subtypes. All blood samples from experimentally FIV-infected cats (n=5) were antibody positive and highly positive in the FIV PCR. In contrast, nine cats became antibody positive after FIV vaccination but remained negative in the FIV PCR. Of 101 FIV antibody-positive feline blood specimens submitted for FIV PCR diagnosis, 61 were positive (60%). A total of 23 of the positive PCRs identified subtype A, 11 identified subtype B1, 11 identified subtype B2/E, and 16 identified subtype C. FIV subtype D was not detected in any submitted specimens even though 13 blood specimens were from cats known to have received the FIV vaccine, which contains FIV subtype A and D inactivated virions. Therefore, this PCR quantitatively identifies FIV subtypes and unambiguously discriminates between FIV-vaccinated and FIV-infected cats.

Loss of naïve (CD45RA+) CD4+ lymphocytes during pediatric infection with feline immunodeficiency virus.

Feline immunodeficiency virus (FIV) infection of cats is an animal model for the pathogenesis of CD4+ lymphopenia and thymus dysfunction in HIV-infected humans. Recently, a monoclonal antibody (755) was reported to recognize the feline homologue to CD45RA, allowing the enumeration of naïve T cells in cats. We tested the hypothesis that pediatric FIV infection would be associated with a selective loss of naïve CD4+ lymphocytes by inoculating newborn cats with a pathogenic clone of FIV (JSY3) or a related clone with an inactive ORF-A gene (JSY3-DeltaORFA), and compared the data to age-matched uninfected control cats. Both FIV inocula were associated with a reduction in the CD4-CD8 ratio (p=0.01), which was attributable to a disproportionate loss of naïve CD4+ cells (p=0.01) vs. naïve CD8+ cells. Therefore, the reduced CD4:CD8 ratio in FIV-infected juvenile cats is associated with a selective depletion of naïve CD4+ cells from the blood.

Proliferative sparganosis in a dog.

CASE DESCRIPTION: A 21-month-old spayed female Border Collie was examined because of progressive right forelimb lameness, signs of pain, and subcutaneous edema. The dog lived in a fenced yard in Tampa, Fla, that contained a small area of marshy terrain. CLINICAL FINDINGS: The subcutis and intermuscular fascia contained multiple cystic cavities filled with larval cestodes (plerocercoids or spargana) and cloudy red fluid. Parasites were identified morphologically and by DNA sequence analysis as pseudophyllidean cestodes, most likely Sparganum proliferum. The dog developed a progressively worsening fever, dyspnea, mature neutrophilia, and hypoproteinemia. Septic pleuritis and peritonitis complicated the later stages of the disease. TREATMENT AND OUTCOME: Treatment with praziquantel, fenbendazole, and nitazoxanide failed to control the proliferation and dissemination of larval cestodes. The dog was euthanatized after 133 days of treatment. At necropsy, numerous parasitic tissue cysts were present in the subcutis and intermuscular fascia; these cysts were most abundant in the soft tissues of the forelimbs and cervical musculature. The pleural and peritoneal cavities contained multiple larval cestodes and were characterized by neutrophilic inflammation and secondary bacterial infection. CLINICAL RELEVANCE: Findings indicated that clinical signs associated with proliferative sparganosis in dogs may be rapidly progressive and that the condition may be refractory to antiparasitic treatment. Veterinarians should be aware of this zoonotic, water-borne agent.

Immunopathogenesis of feline immunodeficiency virus infection in the fetal and neonatal cat.

The global incidence of pediatric HIV infection is estimated at 2.3 million children, most acquiring the infection from their mothers in utero, peripartum, or postpartum. Pediatric HIV infection typically causes a rapidly progressive disease when compared with adult infection, due in part to the profound susceptibility of the neonatal thymus to productive infection or degenerative changes. Failed production of naive T-lymphocytes further limits the success of antiviral therapy to restore immunologic function. In this review, we explore the use of feline immunodeficiency virus (FIV) infection of domestic cats as an animal model for pediatric HIV infection. Cats infected with FIV represent the smallest host of a naturally occurring lentivirus, and the immunodeficiency syndrome elicited by FIV infection is similar to that of HIV-AIDS. The feline-FIV model uniquely reproduces several key aspects of immunosuppressive lentivirus infection of the thymus, allowing investigators to define viral determinants of pathogenicity, influence of host age on disease outcome, and therapeutic strategies to restore thymus function.

Viral gene expression and provirus load of Orf-A defective FIV in lymphoid tissues and lymphocyte subpopulations of neonatal cats during acute and chronic infections.

Neonatal cats were infected with a wild type (JSY3) or orf-A defective (JSY3DeltaORF-A) feline immunodeficiency virus (FIV) to determine the provirus load and level of viral gene expression at the acute versus chronic stages of infection. FIV DNA in the thymus, lymph node, peripheral blood mononuclear cells (PBMCs) and lymphocyte subpopulations at week 8 post-infection was lower in animals infected with JSY3DeltaORF-A as compared to that of JSY3. At week 16 we observed no significant difference in provirus load between the two groups except for B cells where it was higher in the JSY3 infection. In B cells proviral burden was found to be the same in animals infected with JSY3 for both time points. In the chronic stage, therefore, proviral burden dominates in B cells for JSY3, whereas the level of JSY3DeltaORF-A was lower with comparable values for all lymphocytes at both weeks 8 and 16. Gene expression profiles as measured by real time PCR for gag and rev transcripts revealed decreased levels of JSY3DeltaORF-A mRNAs as compared to that of JSY3. The JSY3 chronic phase infection showed viral gene expression to be higher in B cells relative to CD4+ and CD8+ cells. The presence of viral RNA in CD8 and B cells during the chronic infection implicates active virus replication. Hematological profiles revealed that there was a decline in the number of B cells in JSY3DeltaORF-A-infected cats during the chronic stage of infection while no significant change was observed in animals infected with the wild type virus. Comparative analysis of cell numbers to provirus load and levels of viral transcripts in CD4+ and CD8+, however, did not correlate cell numbers to the levels of viral DNA and gene expression. It remains to be determined whether the relatively high virus burden in B cells as compared to CD4+ and CD8+ cells reflects a role for Orf-A in a shift to B cell virus load during the chronic stage of FIV infection.

Calcium hydroxylapatite gel (Radiesse) injection for the correction of postrhinoplasty contour deficiencies and asymmetries.

OBJECTIVES: To describe our technique for the injection of calcium hydroxylapatite gel (Radiesse) to treat postrhinoplasty contour defects and to evaluate the agent's efficacy, duration of action, required dosage, complication rates, and patient satisfaction. Slight defects or asymmetries are not uncommon, even after well-executed rhinoplasty surgery in the most expert of hands. These contour deformities have been treated with filler agents in the past, but with mixed results. Calcium hydroxylapatite gel was recently introduced as a filler agent in facial plastic surgery, but its use has not yet been described in the correction of postrhinoplasty nasal contour defects. DESIGN: A prospective before-and-after trial conducted in a private-practice facial cosmetic surgery office. Eligible patients had postrhinoplasty contour irregularities or asymmetry. Postrhinoplasty irregularities at the nasal dorsum or tip underwent subcutaneous injection with calcium hydroxylapatite. Main outcome measures included number of treatments, posttreatment injection pain score, required dose and complications, natural feel, patient satisfaction, and length of follow-up. Digital photographs were evaluated by blinded observers. RESULTS: Thirteen patients were followed up prospectively for a mean of 2.5 months. The mean injection pain score was 1.9 (scale, 0-5); the mean dose, 0.19 mL. Patient satisfaction was good to excellent in 11 (85%) of 13 cases. Photographic improvement was seen in 15 (88%) of 17 treatment sites. CONCLUSIONS: Calcium hydroxylapatite gel has been shown in this study to improve postrhinoplasty nasal symmetry and smooth the curves and lines that constitute the contour of the nose. The longevity of nasal augmentation by calcium hydroxylapatite remains unsettled; long-term safety is also unknown. Long-term studies of safety and efficacy are indicated.

Effect of heartworm disease and heartworm-associated respiratory disease (HARD) on the right ventricle of cats.

BACKGROUND: Dirofilaria immitis infection occurs in dogs and cats, both of which species are clinically affected by mature adult infections. Cats are uniquely affected by immature-adult infections with an inflammatory pulmonary disease called Heartworm-Associated Respiratory Disease (HARD). D. immitis infection causes pulmonary parenchymal and vascular pathology in the dog and cat. Dogs develop pulmonary hypertension and cor pulmonale, whereas the development of pulmonary hypertension is rare in the cat. D. immitis infection in the dog causes alteration of the right ventricular (RV) extracellular matrix, including a decrease in myocardial collagen. In this study, the RV myocardial changes of cats infected with adult and immature-adult D. immitis were assessed. METHODS: The cardiopulmonary systems of six groups of SPF cats (n = 9-10 per group) were examined 8 or 18 months after infection with L3 D. immitis. Two groups were untreated and allowed to develop adult HW; two groups were treated with ivermectin starting 3 months post infection, thus allowing HARD but no mature adult heartworms; and two groups were treated with selamectin beginning 1 month post infection, preventing development of L5 or adult heartworms. A group of specific pathogen free (SPF) normal cats was utilized as a negative control (n = 12). Lung pathologic lesions were objectively assessed, and both RV and left ventricular (LV) weights were obtained to calculate an RV/LV ratio. Intramural RV myocardial collagen content was quantitatively assessed. RESULTS: RV/LV weight ratios were not different between groups. Negative control cats had significantly greater RV collagen content than all other affected groups (P = 0.032). Analysis of the RV/LV ratios and collagen content revealed no significant relationship (r = 0.03, P = 0.723, respectively). Collagen content had a modest, but significant, negative correlation, however, with both pulmonary vascular pathology (r = -0.25, P = 0.032) as well as the total pulmonary parenchymal and vascular pathology (r = -0.26, P = 0.025). CONCLUSIONS: Cats infected with mature and immature D. immitis did not develop RV hypertrophy but did demonstrate loss of RV myocardial collagen content. The collagen loss was present at 8 and 18 months after infection in all infected cats. This loss of RV myocardial collagen was correlated with the severity of pulmonary parenchymal and vascular pathology.

The immediate use of a silicone sheet wound closure device in scar reduction and prevention.

Silicone has been used successfully postoperatively in the prevention of hypertrophic and other types of adverse scars. The Silicone Suture Plate (SSP) is a new, minimally invasive, sterile wound closure device that is applied intraoperatively to prevent adverse scarring. The SSP device permits immediate application of silicone while concurrently allowing for wound-edge tension redistribution. In this prospective, controlled, single-blinded clinical study, 8 consecutive patients undergoing deep-plane rhytidectomy were selected. SSP devices were placed on the patients' posterior rhytidectomy hairline incision; the mirror-image control site underwent standard suturing techniques. Three blinded, independent raters assessed the treatment and control sides at 6-week and 4-month follow-up visits, using the Objective Scar Assessment Scale (OSAS), a validated scar assessment tool. The 6-week OSAS scores revealed an 18.4% improvement on the side with the SSP device (13.3) when compared to the control side (16.3). The 4-month OSAS scores showed a 27.3% improvement on the treatment side from 12.7 (control) to 9.2 (SSP). These OSAS results were found to be statistically significant when taken as an aggregate of the observers' scores, but not when observers' scores were measured individually (p < 0.05). In our series of patients, we showed promising results with the use of the SSP device. Early silicone application and tissue tension distribution contributed to an overall more aesthetically pleasing scar compared to those seen with standard suturing techniques, although more testing is required.

Abnormal coagulation factor VIII transcript in a Tennessee Walking Horse colt with hemophilia A.

Hemophilia A is an X-chromosome-linked disorder caused by a deficiency in factor VIII (FVIII). Although foals have been diagnosed with hemophilia A based on deficiency in FVIII activity, causative gene mutations have not been identified. The genomic DNA and cDNA encoding FVIII of a Tennesee Walking Horse colt affected with hemophilia A and the genomic DNA of his dam and a normal unrelated horse were analyzed with no splice site or coding sequence abnormalities identified in any of the horses. Polymerase chain reactions (PCR) were then performed on hepatic cDNA from the affected colt and an unrelated normal horse, and no product was obtained for the sequence between and including exon 1 and exon 2 in the affected colt. Based on these results, suspected mutations were identified in the noncoding region of FVIII (intron 1), and genomic sequencing of intron 1 in the dam and the affected colt suggested maternal inheritance.

Pulmonary arterial disease in cats seropositive for Dirofilaria immitis but lacking adult heartworms in the heart and lungs.

OBJECTIVE: To determine the prevalence and severity of pulmonary arterial lesions in cats seropositive for heartworms (Dirofilaria immitis) but lacking adult heartworms in the heart and lungs during necropsy. ANIMALS: 630 adult cats from an animal control shelter in Florida. PROCEDURE: Cats were tested for adult heartworms in the heart and pulmonary arteries and antibody against heartworms in the serum. Histologic examination was conducted on the right caudal lung lobe of 24 heartworm- and antibody-positive cats; 24 heartworm-negative and antibody-positive cats; and 24 heartworm-, antibody-, and antigen-negative cats. Wall areas of 10 small to medium-sized pulmonary arteries of each cat were measured and expressed as a proportion of total cross-sectional area. RESULTS: Heartworm infection or seropositive status was significantly and strongly associated with seventy of medial hypertrophy of pulmonary arterial walls. Heartworm- and antibody-positive cats and heartworm-negative and antibody-positive cats had a significant increase in wall thickness, compared with wall thickness for heartworm- and antibody-negative cats. Heartworm- and antibody-positive cats had the most severe hypertrophy. The proportion with occlusive medial hypertrophy was significantly higher in heartworm- and antibody-positive cats (19/24 [79%]) and heartworm-negative and antibody-positive cats (12/24 [50%]), compared with heartworm- and antibody-negative cats (3/24 [13%]). CONCLUSIONS AND CLINICAL RELEVANCE: Cats with serologic evidence of exposure to heartworms, including those without adult heartworms in the lungs and heart, have a greater prevalence of pulmonary arterial lesions than heartworm-negative cats without serologic evidence of exposure. Additional studies are needed to define the pathogenesis, specificity, and clinical importance of these lesions.

Can functional septorhinoplasty independently treat obstructive sleep apnea?

BACKGROUND: The purpose of this study was to determine whether functional rhinoplasty alone results in a significant improvement in obstructive sleep apnea parameters in patients with nasal obstruction. METHODS: Records of consecutive adult patients with nasal obstruction who underwent surgery to repair their nasal inlet and completed preoperative and postoperative polysomnography were reviewed. Patients underwent polysomnography before and after functional septorhinoplasty. Long-term follow-up using Nasal Obstruction Symptom Evaluation scores was conducted. Statistical analysis was performed using the Wilcoxon signed rank sum test. A Holm-Bonferroni sequential correction was also used because of multiple statistical comparisons being made. RESULTS: Twenty-six patients were included in this study. Mean apnea-hypopnea index scores preoperatively was 24.7, which dropped to a mean postoperative apnea-hypopnea index of 16, a reduction of 35 percent (p = 0.013). Excluding patients with a body mass index greater than 30 resulted in improved apnea-hypopnea index scores, from 22.5 to 9.6, a mean 57 percent reduction (p < 0.01). CONCLUSIONS: Functional rhinoplasty may have the potential to significantly improve the severity of obstructive sleep apnea for select patients with nasal obstruction. The nasal airflow improvement may modify pharyngeal aerodynamics. This is a fast and minimally invasive approach to consider in patients with obstructive sleep apnea and nasal obstruction, especially in patients with a body mass index less than 30. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Hematopoiesis in the feline fetal liver: an assessment by flow cytometry.

The domestic cat is an excellent model for the development of therapeutic protocols that target hematopoietic stem cells (HSCs) because it is relatively resistant to complications related to bone marrow transplantation. To identify a plentiful source of HSC that could be used as targets for gene transduction and transplantation, the livers of 28 mid-gestation fetuses (28-52 days) and late-gestation fetuses (53 days-term) were analyzed for erythroid, myeloid, lymphoid, and uncommitted hematopoietic progenitor cells by flow cytometry. We found that the fetal liver mononuclear cells (FLMCs) contained 57% erythroid progenitors during mid-gestation, but this percentage declined to 43% as gestation progressed. Myelomonocytic cells within FLMC were more numerous in late-gestation (31%) than in mid-gestation (18%). Two monoclonal antibodies (mAb), CH 152 and CH 755, which recognize cells with the potential to reconstitute multilineage hematopoiesis in cats, were tested. Approximately, 32% of FLMC from late-gestation fetuses expressed the epitope recognized by mAb CH 152, a significant increase above the 12% positive cells in mid-gestation fetuses. Approximately, 33% of hepatic mononuclear cells expressed the epitope recognized by mAb CH 755 in both mid-term and late-term fetuses. When expressed in absolute numbers, medians of 2.7 x 10(7) CH 152-positive cells and 3.2 x 10(7) CH 755-positive cells were extracted from the late-term fetal livers of individual cats. T-lymphocytes were a minor component (<3%) of FLMC, despite their presence in the thymus and spleen. These data suggest that the late-term feline fetal liver is a suitable source of mutipotential hematopoietic cells that could be used for gene therapy protocols in the cat.

In vitro development and characterization of a manatee bronchial cell line.

In the present study, culture conditions that promote the growth and differentiation of manatee respiratory tract epithelial cells toward a mucociliary phenotype were determined. Characterization of a manatee-specific cell line enables investigators to conduct in vitro testing where live-animal experimentation is not possible. Cell cultures were established from both explants and enzymatically dissociated cells that were isolated from manatee bronchial tissue. To modulate their differentiation, bronchial epithelial cells were grown on Transwell collagen membranes either submerged or at an air-liquid interface. Growth on a collagen membrane at an air-liquid interface and medium supplemented with retinoic acid was required to promote a mucociliary phenotype. When cells were grown in submerged cultures without retinoic acid, they appeared more squamous and were not ciliated. Intracellular keratin proteins were detected in both submerged and interface cultures. Cultured manatee bronchial epithelial cells will facilitate future studies to investigate their potential role in pulmonary disease associated with brevetoxicosis after exposure to the red-tide organism, Karenia brevis.