Patrizia Bandecchi

Prevalence of feline immunodeficiency virus and other retroviral infections in sick cats in Italy

Two hundred and seventy-seven sick pet cats living in Italy were tested for antibodies to feline immunodeficiency virus (FIV) and for feline leukemia virus (FeLV) antigen. Overall, 24% of the cats resulted positive for anti-FIV antibody and 18% for FeLV antigen. FIV was isolated from the peripheral mononuclear blood cells of ten out of 15 seropositive cats examined and from one out of eight saliva samples. No FIV isolations were obtained from six serum samples cultured. Feline syncytium forming virus (FeSFV) could be isolated from blood and/or saliva in ten out of 11 FIV seropositive cats examined, in six out of nine FeLV antigen positive cats, in two cats found positive for both infection markers, and in three out of 11 cats negative for both markers. Thus, the probability of isolating FeSFV was enhanced by infection with other exogenous retroviruses.

Simple in vitro methods for titrating Feline Immunodeficiency Virus (FIV) and FIV neutralizing antibodies

The feline immunodeficiency virus (FIV) readily produced syncytia in Crandell feline kidney (CrFK) cells adapted to a medium containing 0.5% fetal calf serum, a variety of growth factors and other supplements. This finding has been exploited to develop simple and sensitive virus titration and neutralization assays. High titre neutralizing antibodies were detected in cats infected naturally and experimentally with FIV, but not in uninfected animals.

In situ PCR-associated immunohistochemistry identifies cell types harbouring the Maedi-Visna virus genome in tissue sections of sheep infected naturally

Maedi-Visna virus (MVV) is a non-oncogenic ovine lentivirus whose main targets are the lung, mammary gland, central nervous system and joints. Cells of the monocyte-macrophage lineage are the major viral target in vivo; other cell types are infected as well, as indicated by several studies, largely based on the examination of animals infected experimentally or on the in vitro infection of cultured cells. Aim of this study was to investigate the cell types harbouring the viral genome in lungs and mammary glands of animals infected naturally by using in situ PCR-associated immunohistochemistry. Several types of cells were infected: in the lung type I and II pneumocytes, interstitial and alveolar macrophages, endothelial cells and fibroblast-like cells. Epithelial cells, macrophages, endothelial cells and fibroblast-like cells were infected also in the mammary gland. These results indicate that the in situ PCR, a powerful technique which combines the high sensitivity of the conventional PCR with the ability to localise the cellular targets within a tissue, can be improved further by its association with the immunohistochemistry. This can be especially advantageous when the presence and localisation of the target sequence are investigated in the context of a tissue with its complex cellular organisation.

Feline leukaemia virus (FeLV) and feline immunodeficiency virus infections in cats in the Pisa district of Tuscany, and attempts to control FeLV infection in a colony of domestic cats by vaccination

The seroprevalence of feline immunodeficiency virus (FIV) in 203 apparently healthy domestic cats living in the district of Pisa, central Italy, was 11·3 per cent, and the prevalence of feline leukaemia virus (FeLV) was 8·4 per cent. The prevalence of FIV depended significantly on the lifestyle and age of the cats; cats living outdoors were more likely to be FIV-positive than cats living indoors, and the proportion of FIV-positive cats increased with age. In contrast, there was no significant relationship between these variables and the prevalence of FeLV. There was no significant relationship between the cats’ seropositivity for FIV and FeLV. The results of a five-year field study to control FeLV infection by vaccination in a colony of 30 domestic adult cats naturally exposed to the infection suggest that the vaccination was effective in FIV-negative cats, but failed to protect FIV-positive cats against FeLV.

Epitope mapping of the V3 domain of feline immunodeficiency virus envelope glycoprotein by monoclonal antibodies

A panel of six IgG monoclonal antibodies (MAbs) was produced by immunizing mice with a 22 amino acid synthetic peptide, designated V3.3, of the third variable region of feline immunodeficiency virus (FIV) envelope glycoprotein. This peptide is known to induce neutralizing antibodies in cats. In ELISA all MAbs reacted with purified SDS-disrupted FIV and in flow cytometry all MAbs stained permeated, persistently infected FL4 cells but not unfixed FL4 cells; this indicated that the MAbs recognize essentially cryptic epitopes of the gp100 V3 loop. By direct ELISA using partially overlapping synthetic peptides and by competition binding studies, the anti-V3.3 MAbs were shown to detect at least four distinct epitopes, two located in the amino-terminal half and two in the carboxy-terminal half of the sequence. When tested for neutralizing activity by the syncytium inhibition assay in Crandell feline kidney cells, all anti-V3.3 MAbs neutralized FIV at high dilution. However, at low dilution two MAbs exhibited much less neutralizing activity. These results indicate that the V3 region of FIV contains multiple epitopes involved in neutralization.

Development and comparison of strain specific gag and pol real-time PCR assays for the detection of Visna/maedi virus

The aim of this study was the development of gag and pol dual labelled probe real-time PCR and RT PCR assays to quantify the proviral load and the transcripts of the British Visna/maedi virus EV1 strain. Primers and probes were chosen based on the consensus sequences of gag and pol clones representative of EV1 genetic variants. Both PCRs had a detection limit of 3 copies of target gene, with a linearity over 6 orders of magnitude. The performances of the two PCRs in vivo were evaluated and compared on a panel of DNAs extracted from blood of sheep infected experimentally with EV1. The pol assay detected in most cases lower numbers of viral molecules than gag assay, yielding some false negative results. The gag real-time RT PCR had a detection limit of 100 RNA molecules with a linearity over 5 orders of magnitude. This did not result in a lower performance of the RT PCR compared to the PCR in cells permissive for virus replication, which contain higher numbers of viral transcripts than proviral genomes. The real-time assays developed in this study, particularly the gag assay, provide a sensitive tool which can be used to quantify the viral load in experimental infections.

Evaluation of an ELISA to detect antibodies to maedi-visna virus in individual and pooled samples of milk from sheep.

An ELISA was used to detect antibodies to maedi-visna virus in samples of serum and milk from individual sheep; the results obtained indicated that the ELISA can be used to detect antibodies in milk. The assay was also applied to samples of bulk-tank milk; a standard curve was created and used to calculate the seroprevalence of maedi-visna in 11 flocks of sheep and the results were compared with the results obtained by applying the ELISA to individual serum samples. There was good agreement between the seroprevalences calculated from the standard curve for bulk-tank milk and from the individual serum samples.

Antigenic variability of ovine lentivirus isolated in Italy

S. Rosati1*, M Profiti1, E. Grego, M.L. Carrozza2, M. Mazzei3 and P. Bandecchi3 1Dipartimento di Produzioni Animali Epidemiologia ed Ecologia, Università di T orino; 2Scuola Normale Superiore, Pisa; 3Dipartimento di Patologia Animale, Università di Pisa *Correspondence: Dipartimento di Produzioni Animali, Epidemiologia ed EcologiaFacoltà di Medicina veterinaria V ia L eonardo da V inci, 44 10095 Grugliasco (TO), Italy e-mail: sergio.rosati@unito.it

In vitro antiviral effects of an aqueous extract of Artemisia verlotorum Lamotte (Asteraceae)

Plants represent a possible source of new interesting antiviral drugs. An aqueous extract of Artemisia verlotorum exhibited, in vitro, strong activity against the feline immunodeficiency virus (FIV), which can be considered a reliable model of the human immunodeficiency virus type 1, the aetiological agent of AIDS. The extract determined a decrease of the virus‐induced syncytia in the cultured cells, an inhibition of the viral reverse transcriptase activity and a reduction of the viral capsid protein P24 expression. Copyright

Small Animal Model of AIDS and the Feline Immunodeficiency Virus

Many aspects of the virology, epidemiology and prevention of AIDS are fairly well known, but the mechanisms whereby HIV produces the characteristic severe immune deficiency remain essentially obscure. Even more importantly, the armamentarium for controlling HIV infection is still extremely limited. For these reasons it is generally believed that AIDS research might profit greatly from the study of animal models.