Prevalences of feline leukaemia virus and feline immunodeficiency virus infections in cats in Sydney

Objective To determine prevalences of feline leukaemia virus (FeLV) and feline immunodeficiency virus (FIV) infections in ‘healthy’ cats that, through acute misadventure or other circumstance, were presented to veterinary practitioners. Prevalences of FeLV and FIV in this population were compared to those in a population of predominantly sick cats. Design and procedures Serum specimens were obtained over a 2-year period from 200 cats oldeer than 1 year of age presented to veterinary clinics for routine procedures, including cat fight injuries or abscesses, vehicular trauma, neutering, dental scaling, vaccination, grooming or boarding. An additional 894 sera were obtained over approximately the same period from specimens submitted by veterinarians to a private clinical pathology laboratory, mainly from sick cats suspected of having immune dysfunction, but including some sera from healthy cats being screened prior to FeLV vaccination. FIV antibody and FeLV antigen were detected in samples using commercial enzyme immunoassays. Results Amongst 200 ‘healthy’ cats, the prevalence of FeLV infection was 0 to 2%, and the prevalence of FIV was 6.5 to 7.5%, depending on the stringency of the criteria used to define positivity. FIV infection was significantly more prevalent in cats which resided in an inner city environment (P = 0.013). Of the 894 serum specimens submitted to the laboratory by practitioners, 11/761 (1.4%) were FeLV positive, while 148/711 (20.8%) were FIV positive. The prevalence of FIV was significantly higher in these predominantly ‘sick’ cats than in cats seen for routine veterinary procedures (P < 0.00001), while there was no difference in the prevalence of FeLV (P = 0.75) Conclusions The prevalence of FeLV and FIV in healthy cats may have been substantially overestimated in some previous Australian surveys. FeLV infection would appear to be a rare cause of disease in Australian cats. The higher prevalence of FIV positivity in sick as opposed to healthy cats infers that FIV infection contributes to the development of disease.     

Vaccine protection against feline immunodeficiency virus: setting the challenge

Since feline immunodeficiency virus (FIV) was first isolated, international research efforts have been directed towards developing a protective vaccine, not least because it may provide a model for a candidate human immunodeficiency virus (HIV) vaccine. This article reviews the challenges facing vaccine development, the current state of knowledge and future prospects for FIV vaccination.     

Serological differentiation of FIV-infected cats from dual-subtype feline immunodeficiency virus vaccine (Fel-O-Vax FIV) inoculated cats

Feline immunodeficiency virus (FIV) vaccine, Fel-O-Vax FIV, was released for sale in the US in 2002. The antibodies of vaccinated cats interfere with serological assays by currently available FIV diagnostic kits. In this study, we investigated whether it is possible to distinguish serologically cats vaccinated with Fel-O-Vax FIV from cats experimentally or naturally infected with FIV. A total of 153 sera taken from 97 cats were used as serum samples. Enzyme linked immunosorbent assay (ELISA) was performed using whole FIV antigen and formalin treated whole FIV antigen, recombinant-gag (r-gag) antigen, and transmembrane (TM) peptide. Statistical analysis was performed using ELISA optical density (O.D.) values obtained with each antigen as variables. Except for the ELISA O.D. values obtained with r-gag antigen, a significant difference in ELISA O.D. values was observed between the vaccinated and the infected groups. However, it was not possible to distinguish both groups unequivocally. Using discriminant analysis, it was possible to distinguish the two groups with an accuracy of 97.1% with two discriminating variables (ELISA O.D. values obtained with formalin treated whole FIV antigen, and TM peptide), 97.8% with three discriminating variables (ELISA O.D. values obtained with whole FIV antigen, formalin treated whole FIV antigen, and TM peptide). Therefore, it was considered possible to distinguish cats vaccinated with Fel-O-Vax FIV from FIV-infected cats by ELISA using two types of antigens including formalin treated whole FIV antigen and TM peptide, or three types of antigens including formalin treated whole FIV antigen, TM peptide and whole FIV antigen.     

In vivo CXCR4 expression, lymphoid cell phenotype, and feline immunodeficiency virus infection

Primary isolates of feline immunodeficiency virus (FIV) appear to require binding to CD134 in conjunction with CXCR4(X4) to infect IL-2-dependent T-cell-derived cells in culture. However, much less is known about the role of X4 for the infection of cells in vivo. To investigate the correlation between X4 expression and FIV infection in cats acutely infected with FIV-C-Pgmr we used high-speed fluorescence-activated cell sorting and realtime PCR to co-analyze cell phenotypes from lymph node, thymus, bone marrow and blood for FIV infection and X4 expression. X4 expression was greatest in lymph node, both in frequency and in mean fluorescence intensity. The thymus demonstrated a higher proviral burden in X4+ thymic T cells (14% in X4+ thymic T cells and 7% in X4− cells) whereas, proviral loads were similar between X4+ and X4− cell populations in all other tissues examined. Assuming a minimum of one proviral copy per cell, a maximum of 50% of FIV-positive cells were X4+. The highest fraction of FIV-infected X4− cells was present in bone marrow. Regardless of X4 status, proviral loads were higher in lymph node and blood T cells than in B cells. These studies provide both a positive association between X4 expression and FIV infection and introduce the probability that X4-independent infection occurs in other target cells in vivo.     

Prevalence and risk factors for cats testing positive for feline immunodeficiency virus and feline leukaemia virus infection in cats entering an animal shelter in New Zealand

Abstract

AIMS

To estimate the prevalence of cats testing positive for antibodies to feline immunodeficiency virus (FIV) and feline leukaemia virus (FeLV) antigens in domestic cats entering a New Zealand animal shelter, based on a commercial point-of-care ELISA, to identify risk factors associated with cats testing positive, and to compare the results obtained from the ELISA with those obtained using PCR-based testing.     

Combination chemotherapy in feline lymphoma: treatment outcome, tolerability, and duration in 23 cats

BACKGROUND: Different chemotherapy regimes have been described for feline lymphoma with varying outcomes. HYPOTHESIS: In cats with lymphoma, a long-term, multiagent chemotherapy protocol will be effective and carry acceptable toxicity. ANIMALS: Twenty-three cats with histologically or cytologically confirmed diagnosis of lymphoma. METHODS: Prospective, single-arm clinical trial in which cats were treated with a chemotherapy protocol consisting of a cyclic combination of l-asparaginase, vincristine, cyclophosphamide, doxorubicin, methotrexate, and prednisolone with a planned total treatment time of 122 weeks. RESULTS: Complete remission (CR) rate was 74% (n = 17). Fourteen percent of cats attained partial remission (PR). Median duration of first CR was 264 days (range, 45-2,485 days). Six-month, 1-, and 2-5-year remission rates were 75, 50, and 34%, respectively. Duration of PR ranged between 23 and 63 days. Median survival in cats with CR was 296 days (range, 50-2,520 days). Six-month, 1-, 2-, and 3-5-year survival rates in cats with CR were 82, 47, 34, and 27%, respectively. Survival of cats achieving PR ranged between 38 and 120 days. Of the analyzed variables, only anatomical location had a significant influence on remission duration (P=.022). Actual median treatment time in cats with CR was 128 days (18 weeks). Hematologic and gastrointestinal toxicosis was infrequent and mostly low grade. CONCLUSIONS AND CLINICAL IMPORTANCE: In this population of cats with lymphoma, chemotherapy was effective. With infrequent and mostly low-grade toxicosis, tolerability of the protocol may be considered good.     

Diagnosing feline immunodeficiency virus (FIV) infection in FIV-vaccinated and FIV-unvaccinated cats using saliva

We recently showed that two immunochromatography point-of-care FIV antibody test kits (Witness FeLV/FIV and Anigen Rapid FIV/FeLV) were able to correctly assign FIV infection status, irrespective of FIV vaccination history, using whole blood as the diagnostic specimen. A third FIV antibody test kit, SNAP FIV/FeLV Combo (an enzyme-linked immunosorbent assay [ELISA]), was unable to differentiate antibodies produced in response to FIV vaccination from those incited by FIV infection. The aim of this study was to determine if saliva is a suitable diagnostic specimen using the same well characterized feline cohort. FIV infection status of these cats had been determined previously using a combination of serology, polymerase chain reaction (PCR) testing and virus isolation. This final assignment was then compared to results obtained using saliva as the diagnostic specimen utilizing the same three point-of-care FIV antibody test kits and commercially available PCR assay (FIV RealPCR). In a population of cats where one third (117/356; 33%) were FIV-vaccinated, both immunochromatography test kits accurately diagnosed FIV infection using saliva via a centrifugation method, irrespective of FIV vaccination history. For FIV diagnosis using saliva, the specificity of Anigen Rapid FIV/FeLV and Witness FeLV/FIV was 100%, while the sensitivity of these kits was 96% and 92% respectively. SNAP FIV/FeLV Combo had a specificity of 98% and sensitivity of 44%, while FIV RealPCR testing had a specificity of 100% and sensitivity of 72% using saliva. A revised direct method of saliva testing was trialed on a subset of FIV-infected cats (n = 14), resulting in 14, 7 and 0 FIV positive results using Anigen Rapid FIV/FeLV, Witness FeLV/FIV and SNAP FIV/FeLV Combo, respectively. These results demonstrate that saliva can be used to diagnose FIV infection, irrespective of FIV vaccination history, using either a centrifugation method (Anigen Rapid FIV/FeLV and Witness FeLV/FIV) or a direct method (Anigen Rapid FIV/FeLV). Collection of a saliva specimen therefore provides an acceptable alternative to venipuncture (i) in fractious cats where saliva may be easier to obtain than whole blood, (ii) in settings when a veterinarian or trained technician is unavailable to collect blood and (iii) in shelters where FIV testing is undertaken prior to adoption but additional blood testing is not required.     

The molecular biology and evolution of feline immunodeficiency viruses of cougars

Feline immunodeficiency virus (FIV) is a lentivirus that has been identified in many members of the family Felidae but domestic cats are the only FIV host in which infection results in disease. We studied FIVpco infection of cougars (Puma concolor) as a model for asymptomatic lentivirus infections to understand the mechanisms of host–virus coexistence. Several natural cougar populations were evaluated to determine if there are any consequences of FIVpco infection on cougar fecundity, survival, or susceptibility to other infections. We have sequenced full-length viral genomes and conducted a detailed analysis of viral molecular evolution on these sequences and on genome fragments of serially sampled animals to determine the evolutionary forces experienced by this virus in cougars. In addition, we have evaluated the molecular genetics of FIVpco in a new host, domestic cats, to determine the evolutionary consequences to a host-adapted virus associated with cross-species infection. Our results indicate that there are no significant differences in survival, fecundity or susceptibility to other infections between FIVpco-infected and uninfected cougars. The molecular evolution of FIVpco is characterized by a slower evolutionary rate and an absence of positive selection, but also by proviral and plasma viral loads comparable to those of epidemic lentiviruses such as HIV-1 or FIVfca. Evolutionary and recombination rates and selection profiles change significantly when FIVpco replicates in a new host.     

Deep and superficial skin scrapings from a feline immunodeficiency virus-positive cat

An 8-year-old, neutered male, domestic shorthair cat housed at the North Carolina State University, College of Veterinary Medicine, Laboratory Animal Research facility as part of a research colony was examined because of mulifocal skin lesions. The lesions consisted of patchy alopecia with mild crusting of the periauricular region, neck, and dorsum; periauricular excoriations; marked dorsal seborrhea and scaling; and generalized erythematous papules. A moderate amount of ceruminous exudate was present in both ear canals. Results of testing for feline immunodeficiency virus (FIV) were positive. An ear swab specimen and superficial and deep skin scrapings were obtained, mounted with oil on glass slides, and coverslipped for microscopic examination. Two populations of mites were observed: a large population of slender, long (approximately 200 microm), adult mites with long, tapering abdomens that comprised two-thirds of the total body length; and a smaller population of more translucent and shorter mites (approximately 100 microm) with wide, blunt abdomens that had prominent transverse ridges. The interpretation was demodicosis, with Demodex cati and D gatoi co-infection. Histologic sections of biopsies from skin lesions on the neck, dorsum, and periauricular area contained a mild perivascular and perifollicular inflammatory infiltrate composed predominantly of histiocytes, lymphocytes, and plasma cells. Diffusely within the follicular lumina and occasionally within the superficial keratin, a myriad of Demodex organisms were observed. Intrafollicular mites were compatible in appearance with D cati whereas those in the corneal layer were suggestive of D gatoi. Demodicosis is an uncommon disease of cats, and rare cases of dual infection have been documented, occasionally in FIV-infected cats. The dual infection emphasizes the importance of doing both superficial and deep skin scrapings and of recognizing the unique microscopic features of different Demodex mites.     

小反刍兽疫病毒H、F基因转基因苜蓿表达载体的构建

为了有效预防小反刍兽疫,试验以pBI121为载体,PPRV-H、PPRV-F为目的基因,并插入绿色荧光蛋白(GFP)基因、MAR序列,构建5种小反刍兽疫病毒(PPRV)苜蓿表达载体。结果表明:成功构建小反刍兽疫表达载体,再将表达载体转入苜蓿,饲喂动物,选择其中免疫效果最好的表达载体,为该病的预防工作提供有效的技术支持。     

东北虎源猫免疫缺陷病毒pol基因遗传进化分析

为了解中国东北虎源猫免疫缺陷病毒(FIV)的感染情况,采集海林市横道河子东北虎林园东北虎全血样本50份,通过套式PCR对样本进行FIV pol基因扩增,并对阳性样本进行测序,再运用生物信息学软件进行遗传进化分析.结果检测出1只虎为FIV阳性,其pol基因与参考毒株的核苷酸和氨基酸序列同源性分别为70.0％～99.8％和...     

EIAV基因转移载体的优化

本研究在马传染性贫血病毒（EIAV）基因转移载体pcPPTPRE（＋）的基础上[含有报告基因增强绿色荧光蛋白（EGFP）和人乙型肝炎病毒转录后调控元件（HARE）1,对其转录后调控元件和外源基因启动子进行优化。参考土拨鼠肝炎病毒（Woodchuck hepatitis virus,WHV）基因组序列和人巨细胞病毒增强子／鸡B—actin启动子序列（CAGp）,设计合成2对引物,分别扩增WHV转录后调控元件（WARE）和CAGp片段,正向插入pcPPTPRE（＋）获得重组质粒pcPPTWARE和pcPPTWCAG。将获得的EIAV转移载体pcPPTWARE和pcPPTWCAG以及pcPPTPRE（＋）,脂质体法分别转染人胚肾细胞HEK293和鸡胚成纤维细胞系DF-1,转染后12h、24h、36h、48h在荧光显微镜下观察特异性绿色荧光的表达。48h后收获细胞,利用流式细胞仪检测转染细胞中表达EGFP的阳性细胞。结果表明,在HEK293细胞中pcPPTWARE和pcPPTWCAG的阳性细胞平均百分率均极显著高于pcPPTPRE（＋）（P〈0．01）,pcPPTWCAG显著高于pcPPTWARE（P〈0．05）;pcPPTWCAG的平均道数值极显著高于pcPPTPRE（＋）和pcPPTWARE,pcPPTWPRE显著高于pcPPTPRE（＋）。在DF-1细胞中pcPPTWARE和pcPPTWCAG的阳性细胞平均百分率均显著高于pcPPTPRE（＋）,而pcPPTWARE和pcPPTWCAG的平均道数值均极显著高于pcPPTPRE（＋）,pcPPTWCAG显著高于pcPPTWARE。本研究利用WARE和CAGp优化构建了EIAV基因转移载体pcPPTWCAG,获得了较强的基因表达能力,为以鸡为宿主进行转基因研究提供了条件。     

Novel diabetes mellitus treatment: mature canine insulin production by canine striated muscle through gene therapy

Muscle-targeted gene therapy using insulin genes has the potential to provide an inexpensive, low maintenance alternative or adjunctive treatment method for canine diabetes mellitus. A canine skeletal muscle cell line was established through primary culture, as well as through transdifferentiation of canine fibroblasts after infection with a myo-differentiation gene containing adenovirus vector. A novel mutant furin-cleavable canine preproinsulin gene insert (cppI4) was designed and created through de novo gene synthesis. Various cell lines, including the generated canine muscle cell line, were transfected with nonviral plasmids containing cppI4. Insulin and desmin immunostaining were used to prove insulin production by muscle cells and specific canine insulin ELISA to prove mature insulin secretion into the medium. The canine myoblast cultures proved positive on desmin immunostaining. All cells tolerated transfection with cppI4-containing plasmid, and double immunostaining for insulin and desmin proved present in the canine cells. Canine insulin ELISA assessment of medium of cppI4-transfected murine myoblasts and canine myoblast and fibroblast mixture proved presence of mature fully processed canine insulin, 24 and 48 h after transfection. The present study provides proof of principle that canine muscle cells can be induced to produce and secrete canine insulin on transfection with nonviral plasmid DNA containing a novel mutant canine preproinsulin gene that produces furin-cleavable canine preproinsulin. This technology could be developed to provide an alternative canine diabetes mellitus treatment option or to provide a constant source for background insulin, as well as C-peptide, alongside current treatment options.     

Preparation and diagnostic utility of a hemagglutination inhibition test antigen derived from the baculovirus-expressed hemagglutinin-neuraminidase protein gene of Newcastle disease virus

A recombinant hemagglutinin-neuraminidase (rHN) protein from Newcastle disease virus (NDV) with hemagglutination (HA) activity was expressed in Spodoptera frugiperda cells using a baculovirus expression system. The rHN protein extracted from infected cells was used as an antigen in a hemagglutination inhibition (HI) test for the detection and titration of NDV-specific antibodies present in chicken sera. The rHN antigen produced high HA titers of 2¹³ per 25 µL, which were similar to those of the NDV antigen produced using chicken eggs, and it remained stable without significant loss of the HA activity for at least 12 weeks at 4℃. The rHN-based HI assay specifically detected NDV antibodies, but not the sera of other avian pathogens, with a specificity and sensitivity of 100% and 98.0%, respectively, in known positive and negative chicken sera (n = 430). Compared with an NDV-based HI assay, the rHN-based HI assay had a relative sensitivity and specificity of 96.1% and 95.5%, respectively, when applied to field chicken sera. The HI titers of the rHN-based HI assay were highly correlated with those in an NDV-based HI assay (r = 0.927). Overall, these results indicate that rHN protein provides a useful alternative to NDV antigen in HI assays.