Molecular characterisation of 12 Chlamydophila felis polymorphic membrane protein genes.

A group of genes thought to encode members of the unique chlamydial polymorphic membrane protein (pmp) family were recently described in the Chlamydophila felis genome. This study aimed to commence characterisation of a subset of 12 of these putative pmp genes by developing and using gene-specific real-time (Q)PCR assays to confirm their presence in a wide range of C. felis field isolates and laboratory strains, and to look for pmp mRNA expression during in vitro infection. Sequencing of 525-698 base pair regions of pmp genes 7, 9-11, 13-20 for two laboratory strains of C. felis and alignment with the published Fe/C-56 sequence found only a single nucleotide polymorphism present in pmp9. Following the development of gene-specific (Q)PCR assays, analysis of genomic DNA extracted from 40 C. felis field isolates and 4 laboratory strains found that all 12 pmp genes were represented in all cases. Reverse transcription (RT)-QPCR analysis of RNA extracted from cell cultures at 24 and 48 h post inoculation with 1 of 5 different strains of C. felis detected transcripts for all 12 pmp genes at both time points. Analysis of the relative levels of pmp gene transcription suggested that down-regulation of the expression of multiple C. felis pmp genes occurs between 24 and 48 h post inoculation. This study provides the first evidence that 12 of the putative pmp C. felis genes are transcribed during in vitro infection, and shows that these genes are present in a large range of C. felis field isolates and multiple passage laboratory-grown strains.

Diagnosis of feline leukaemia virus infection by semi-quantitative real-time polymerase chain reaction.

In this paper the design and use of a semi-quantitative real-time polymerase chain reaction assay (RT-PCR) for feline leukaemia virus (FeLV) provirus is described. Its performance is evaluated against established methods of FeLV diagnosis, including virus isolation and enzyme-linked immunoassay (ELISA) in a population of naturally infected cats. The RT-PCR assay is found to have both a high sensitivity (0.92) and specificity (0.99) when examined by expectation maximisation methods and is also able to detect a large number of cats with low FeLV proviral loads that were negative by other conventional test methods.

An update on FIV and FeLV test performance using a Bayesian statistical approach.

BACKGROUND: Screening tests for feline retroviruses are thought to have high sensitivity and specificity, although previous studies that evaluated these tests have limitations. Novel statistical approaches have been developed that allow the estimation of sensitivity and specificity in situations where the true state of the disease in individual animals cannot be assured. OBJECTIVE: The purpose of this study was to evaluate the sensitivity and specificity of a variety of retrovirus tests, including some screening tests, in a population of cats potentially infected with either feline leukemia virus (FeLV) and/or feline immunodeficiency virus (FIV) by using a Bayesian statistical approach. METHODS: Four hundred and ninety blood samples from cats being evaluated for FIV infection were tested by 2 rapid immunomigration tests (Witness single [WS], Witness combi [WC]) and a plate-based ELISA (Petcheck) for FIV antibody, and by a newly designed real-time polymerase chain reaction (PCR) assay for FIV provirus. Four hundred and ninety-five blood samples from cats being evaluated for FeLV infection were tested by 2 rapid immunomigration tests (WS, WC) and a plate-based ELISA (Petcheck) for FeLV antigen, and by a FeLV virus isolation technique. Results were then analyzed by using a Bayesian statistical method. RESULTS: For FIV tests, median sensitivity estimates were 0.98 for WS, 0.97 for WC, 0.98 for ELISA, and 0.92 for PCR. Median specificity estimates were 0.96 for WS, 0.96 for WC, 0.93 for ELISA, and 0.99 for PCR. For FeLV tests, median sensitivity estimates were 0.97 for WS, 0.97 for WC, 0.98 for ELISA, and 0.91 for virus isolation. Median specificity estimates were 0.96 for WS, 0.96 for WC, 0.98 for ELISA, and 0.99 for virus isolation. CONCLUSIONS: The use of Bayesian statistical methods overcomes a variety of methodologic problems associated with diagnostic test evaluations, including the lack of a definitive reference test. The sensitivity and the specificity of all 6 evaluated screening tests was high: however, specificity estimates were slightly lower than those reported by most recent studies.

Healthcare professional education: a unique role for hospices.

We sought to describe the educational experiences that hospices currently provide for students in health professions by using a national cross-sectional, Web-based survey of the National Hospice and Palliative Care Organization (NHPCO) and the Population-based Palliative Care Research Network (PoPCRN) hospice sites. Of the 75 responding sites, 71 (95 percent) participated in the education of health profession students, most commonly providing education for bachelor of science in nursing students (73 percent), associate's degree nursing students (61 percent), master's-level social work or counseling students (61 percent), registered nursing students (59 percent), medical students (58 percent), and medical residents (57 percent). Significant staff time was devoted to educating these students (averaging 11 to 30 hours/week), but less than 3 percent received any financial compensation. Therefore, we conclude that hospices provide a significant amount of uncompensated education. This conclusion provides a unique opportunity for hospices to collaborate with healthcare educational institutions and to influence the work force of the future.

Polymerase chain reaction survey of feline haemoplasma infections in Greece.

The aim of this study was to use real-time polymerase chain reaction assays to determine the prevalence of three haemoplasma species in cats from Greece and to evaluate possible associations between haemoplasma infection and age, gender, feline immunodeficiency virus/feline leukaemia virus (FIV/FeLV) status and packed cell volume (PCV). Ninety-seven cats (24 ill anaemic, 55 ill non-anaemic, 18 healthy non-anaemic) were included in the study. Twenty cats (20.6%) were haemoplasma positive; seven cats were infected only with Mycoplasma haemofelis, 10 were infected only with 'Candidatus Mycoplasma haemominutum' and three were co-infected with M haemofelis and 'Candidatus M haemominutum'. 'Candidatus Mycoplasma turicensis' was not detected. Haemoplasma infection was associated with older age (P=0.019). M haemofelis infection tended to be more common in anaemic cats (P=0.058). No association between gender and haemoplasma infection, or haemoplasma relative copy number and PCV, was detected. Retroviral infection rates were very low with only one FeLV proviral positive cat found.

Detection of Chlamydophila felis and feline herpesvirus by multiplex real-time PCR analysis.

Chlamydophila felis and feline herpesvirus (FHV) are pathogens commonly associated with feline respiratory and ocular disease. A real-time multiplex PCR assay was developed to allow detection of these organisms, together with feline 28S ribosomal DNA, in a single tube. Of 538 ocular swab samples tested, 123 were positive for FHV, 97 were positive for C. felis, and 16 were positive for both pathogens.