Giacomo Coppola

Molecular Detection, Epidemiology, and Genetic Characterization of Novel European Field Isolates of Equine Infectious Anemia Virus

ABSTRACT The application of molecular diagnostic techniques along with nucleotide sequence determination to permit contemporary phylogenetic analysis of European field isolates of equine infectious anemia virus (EIAV) has not been widely reported. As a result, of extensive testing instigated following the 2006 outbreak of equine infectious anemia in Italy, 24 farms with a history of exposure to this disease were included in this study. New PCR-based methods were developed, which, especially in the case of DNA preparations from peripheral blood cells, showed excellent correlation with OIE-approved agar gel immunodiffusion (AGID) tests for identifying EIAV-infected animals. In contrast, the OIE-recommended oligonucleotide primers for EIAV failed to react with any of the Italian isolates. Similar results were also obtained with samples from four Romanian farms. In addition, for the first time complete characterization of gag genes from five Italian isolates and one Romanian isolate has been achieved, along with acquisition of extensive sequence information (86% of the total gag gene) from four additional EIAV isolates (one Italian and three Romanian). Furthermore, in another 23 cases we accomplished partial characterization of gag gene sequences in the region encoding the viral matrix protein. Analysis of this information suggested that most Italian isolates were geographically restricted, somewhat reminiscent of the “clades” described for human immunodeficiency virus type 1 (HIV-1). Collectively this represents the most comprehensive genetic study of European EIAV isolates conducted to date.

Quantification of Equid herpesvirus 5 DNA in clinical and necropsy specimens collected from a horse with equine multinodular pulmonary fibrosis

A 15-year-old Belgian gelding was referred for fever, depression, and respiratory distress. Lung biopsy revealed interstitial fibrosis consistent with chronic interstitial pneumonia. Equid herpesvirus 5 (EHV-5) DNA was detected by polymerase chain reaction (PCR) in bronchoalveolar lavage and biopsy specimens. A presumptive diagnosis of equine multinodular pulmonary fibrosis (EMPF) was made, and the horse was administered a systemic treatment with corticosteroids and antiviral drugs. Despite initial clinical improvement, 4 weeks later, the condition of the horse rapidly deteriorated, and the animal was euthanized. Postmortem examination confirmed the presumptive diagnosis of EMPF. The EHV-5 DNA load in different tissues was estimated using a quantitative real-time PCR. Lung had a remarkable viral load, higher than in other organs, especially within the pulmonary fibrotic nodules, and a linkage between high viral burden and the most severely affected tissues was observed. The results suggest that the quantitative real-time PCR is a useful tool to quantify the EHV-5 load in different organs and to understand the relationship between EHV-5 and EMPF. The bronchoalveolar lavage was determined to be a good clinical sample to estimate the EHV-5 load in lung.

Isolation and characterization of β-haemolytic-Streptococci from endometritis in mares.

The objective of this manuscript was to validate published PCR-based methods for detection of β-haemolytic Streptococci by comparison with established bacteriological techniques using 85 clinical isolates recovered from uterine swabs of mares with clinical signs of endometritis and to determine the distribution of SeeL/SeeM and SzeL/SzeM superantigens in isolates of Streptococcus equi subsp. equi (S. equi) and S. equi subsp. zooepidemicus (S. zooepidemicus). The conventional bacteriological techniques showed the vast majority of these isolates (78) were S. zooepidemicus with just 5 Streptococcus dysgalactiae subsp. equisimilis (S. equisimilis) and 2 S. equi strains detected. The PCR analyses confirmed the bacteriological results demonstrating the reliability of the 16S rRNA PCR assay for detecting Streptococci, the multiplex PCR for differentiating between S. zooepidemicus, and S. equi, and PCR assays based on streptokinase genes for identification of S. equisimilis. PCRs for genes encoding superantigens revealed seeL and seeM specific amplicons with size of approximately 800 and 810 bp respectively for the S. equi strains and for 2 S. zooepidemicus strains. To our knowledge, this is the first report of szeL and szeM possession by S. zooepidemicus isolates derived from endometritis in mares.

Pulmonary rhodococcosis in a cat

Feline Rhodococcus equi infection is rare, despite the bacteria is widespread in the environment. R equi infection is typically observed in equine species but the infection has also been reported in dogs, cats and other domestic animals. There are a few reports regarding pulmonary R equi infection in cats and the disease appears to be limited to the skin and the subcutaneous tissue. This report describes the pathological, microbiological and the virulence features associated with an acute necrosuppurative pneumonia in a cat. To the best of our knowledge, this is the first report of feline pulmonary R equi infection in Italy.