Serological survey of canine parvovirus 2 antibody titres in breeding kennels in northern Italy.

BACKGROUND: Current guidelines recommend parvovirus revaccination of adult dogs no more frequently than every 3 years. The aim of this study was to determine the prevalence of dogs showing protective serum antibody titres against canine parvovirus 2 in breeding kennels in Northern Italy and to assess the effect of time from vaccination and the sex of the dog on antibody titres. The study was carried out on 370 animals of different breeds kept in 33 breeding kennels. Antibodies to canine parvovirus 2 in serum samples were measured with an indirect immunoenzymatic assay validated by the manufacturer in relation to the 'gold standard' haemagglutination inhibition test. The number of months that had elapsed since the last vaccination was calculated for each animal and categorized into the following classes: < 12 months; 13-24 months; 25-36 months; 37-48 months; and > 49 months. RESULTS: The prevalence of 'unprotected' dogs was 4.6%. A satisfactory solid herd immunity was present in the majority of breeding kennels, although some vaccination failures were detected. A significant negative correlation was found between antibody titre and months since last vaccination. Comparable antibody titres were found in the first 3 years after vaccination. Although the antibody titre over time was not affected by the sex of the dog, 'unprotected' females had been vaccinated more recently than males with analogous low titres. CONCLUSIONS: Parvovirus revaccination of adult dogs every 3 years, as currently recommended, is also the appropriate recommendation for breeding kennels. Serological tests could be a useful tool to assess the effectiveness of vaccination.

Field application of an indirect gE ELISA on pooled milk samples for the control of IBR in free and marker vaccinated dairy herds.

BACKGROUND: The aim of the present study was to assess the reliability of a new strategy for monitoring the serological response against Bovine Herpesvirus 1 (BoHV1), the causative agent of infectious bovine rhinotracheitis (IBR). Bulk milk samples have already been identified as cost effective diagnostic matrices for monitoring purposes. Nevertheless, most eradication programs are still based on individual standard assays. In a region of northwestern Italy (Piedmont), the voluntary eradication program for IBR has become economically unsustainable. Being the prevalence of infection still high, glycoprotein E-deleted marker vaccines are commonly used but gE blocking ELISAs are less sensitive on bulk milk samples compared to blood serum. RESULTS: A recently developed indirect gE ELISA showed high versatility when applied to a wide range of matrices. In this study, we applied a faster, cost effective system for the concentration of IgG from pooled milk samples. The IgG enriched fractions were tested using a gE indirect ELISA for monitoring purposes in IBR-positive and IBR-marker-vaccinated herds. Official diagnostic tests were used as gold standard. During a 3 years study, a total 250 herds were involved, including more than 34,500 lactating cows. The proposed method showed a very good agreement with official diagnostic protocols and very good diagnostic performances: only 37 positive animals were not detected across the entire study. CONCLUSIONS: The results highlighted the ability of the proposed method to support the surveillance of IBR in the Piedmont region, reducing the costs without affecting the diagnostic performances.

Development and validation of an indirect ELISA as a confirmatory test for surveillance of infectious bovine rhinotracheitis in vaccinated herds.

BACKGROUND: Bovine herpesvirus 1 (BoHV1) is a member of the viral subfamily of Alphaherpesvirinae that infects various species, including cattle, sheep, and goats. The virus causes infectious bovine rhinotracheitis (IBR), which is included in a European list of diseases that may require control and eradication programs. The lack of confirmatory tests affects the validity of diagnostic tools, especially those used for vaccinated herds. In this study, we report the development and validation of an indirect enzyme-linked immunosorbent assay (ELISA) based on BoHV1 glycoprotein E, which was expressed as a secreted recombinant antigen in a mammalian cell system. The performance of the new rec-gE ELISA was compared with that of commercially available indirect and/or blocking ELISAs. RESULTS: The sample set included blood sera from animals from IBR-positive farms, IBR-free farms, and marker-vaccinated farms. The indirect ELISA proposed in this study is based on antibody reactivity against BoHV1 gE, and showed high sensitivity and specificity (98.41 and 99.76 %, respectively). CONCLUSIONS: The ELISA performed well, in terms of both its diagnostic sensitivity and specificity, and as a confirmatory methodology, and therefore should improve the diagnostic protocols used for IBR surveillance.

Feline morbillivirus in northwestern Italy: first detection of genotype 1-B.

OBJECTIVES: A novel morbillivirus was recently described in stray and domestic cats in Asia, the USA and Europe. Most cats infected with feline morbillivirus (FeMV) showed lower urinary tract or kidney disease. Although the association of FeMV infection and kidney diseases has been suggested, the virus pathogenicity remains unclear. The present study aimed to investigate the distribution of FeMV infection, as well as the relationship between FeMV infection and kidney diseases in cats from northwestern Italy. METHODS: A total of 153 urine samples (150 individuals and three pools) and 50 kidney samples were collected and included in the study; total RNA was extracted and a reverse transcription quantitative PCR (RT-qPCR) was performed in order to identify FeMV. Kidneys were also submitted to anatomopathological examination. Phylogenetic analysis and isolation attempts were carried out on positive samples. In FeMV-positive cats, urinalysis and blood analysis were performed. RESULTS: FeMV RNA was detected in 7.3% of urine samples and in 8% of kidney samples, both in healthy cats and in cats with clinical signs/post-mortem lesions compatible with kidney disease. At histopathological examination, tubulointerstitial nephritis (TIN) was shown in 3/4 positive kidney samples, but a clear relationship between FeMV and TIN was not observed. Isolation attempts were unsuccessful, although the urine sample of one castrated male cat hosted in a cattery showed a positive signal in RT-qPCR until the fourth cell passage. Phylogenetic analysis revealed that this FeMV strain belonged to genotype 1-B. In the same cattery, a second genotype 1-B variant was detected from a urine pool. Urinalysis showed proteinuria in three cats, while at blood analysis three cats presented altered creatinine levels. CONCLUSIONS AND RELEVANCE: Data reported suggest the presence of a FeMV sub-cluster distinct from the strain previously isolated in Italy, whose role in renal disorders remains uncertain.

Surveillance of Infectious Bovine Rhinotracheitis in marker-vaccinated dairy herds: Application of a recombinant gE ELISA on bulk milk samples.

Infectious Bovine Rhinotracheitis (IBR) occurs worldwide, requiring significant resources for eradication programs or surveillance purposes. The status of infection is usually detected by serological methods using the virus neutralization test (VNT) or enzyme-linked immunosorbent assay (ELISA) on individual sera. The gE DIVA (Differentiating Infected from Vaccinated Animals) vaccines approach, adopted in order to reduce the virus circulation and prevent clinical signs, have tightened the range of available methods for the serological diagnosis. Different gE blocking ELISA could be performed to detect specific antibodies in sera of infected or whole virus-vaccinated animals but with less sensitivity if applied to bulk milk samples, especially in marker-vaccinated herds. A new rec-gE ELISA was recently developed in Italy and applied with good performances on blood serum samples. The present paper focuses on the application of a rapid protocol for purification/concentration of immunoglobulin G (IgG) from bulk milk and on the use of the new rec-gE indirect ELISA. The study involved three different partners and 225 herds (12,800 lactating cows) with different official IBR diagnostic statuses. The diagnostic specificity of the method was demonstrated closed to 100% while the diagnostic sensitivity was strictly related to the herd-seroprevalence. Considering 2.5% as the limit of detection of within-herd seropositivity prevalence, the diagnostic sensitivity showed by the proposed method was equal to 100%. A single reactivation of a whole strain vaccine in an old cow was detected inside a group of 67 lactating cows, showing the field applicability of the method.