Epidemiology and genetic characterization of <i>Border Disease Virus</i> circulating in Sardinia

Border Disease Virus (BDV), a pestivirus from the Flaviviridae family, is an important pathogen of sheep and goats responsible for significant losses in farms around the world. In spite of the relevance of this pathogen there are only a few epidemiological studies on BDV infection and, as a consequence, the economic impact on small ruminant productions is probably under-estimated. The aims of this study are i) to determine the distribution of BDV in small ruminant farms in Sardinia and genetically characterize circulating strains ii) analyze the relation between seroprevalence, Somatic Cells Count (SCC) an milk yeld. ELISA was performed using “BVDV/MD/BDV p80 Protein Antibody Test Kit” (IDEXX) on serum of bulk tank milk (BTM) samples collected from Sardinian sheep flocks and goat herds between spring 2014 and 2015. The number of sampled farms corresponded to 8.5% of all registered farms in Sardinia. RNA was isolated using Qiamp Viral RNA mini kit from the cellular fraction of each ELISA positive BTV sample and amplified by rt-PCR using complementary primers to a highly conserved region in the untranslated regions (UTRs) of the viral genome. The amplicons were sequenced for phylogenetic analysis. Geographic distribution of collected specimen, seroprevalence and virological positive samples were analyzed via GIS (ESRI ARCGIS 10.3). ELISA screening shows a seroprevalence of 8.3% among goat farms and 10.5% among ovine farms. Ten from the ELISA positive samples were found rt-PCR positive. The sequence analysis indicates that all the amplified samples match with BDV genomes and the phylogenetic analysis revealed that all the viruses clustered in the same group classified as BDV-7. BDV-7 is the only group isolated in Sardinia so far

Development of a Real-Time PCR for detection of <i>Mycoplasma agalactiae</i> in bulk tank milk samples and epidemiology of infection in Sardinia

In this work the Mycoplasma agalactiae p48 gene was used as a diagnostic marker for contagious agalactia (CA) of sheep and goats by Real-Time PCR. The p48 gene encodes an invariable, constantly expressed, immunodominant surface lipoprotein belongs to the basic membrane protein family. The Real-Time PCR test based on p48 resulted specific and sensible. The test performance were evaluated on bulk tank milk samples collected from 1064 ovine and 66 goat farms in sardinian region. 4.8% of sheep farms and 4.5 % of goat farms tested positive. Our results showed that the test based on the p48 gene can be used on bulk tank milk for detection and epidemiological surveillance of Mycoplasma agalactiae infections

In vitro phenotypic characterisation of two genotype I African swine fever viruses with genomic deletion isolated from Sardinian wild boars

African swine fever virus (ASFV) causes a devastating disease affecting domestic and wild pigs. ASF was first introduced in Sardinia in 1978 and until 2019 only genotype I isolates were identified. A remarkable genetic stability of Sardinian ASFV isolates was described, nevertheless in 2019 two wild boar isolates with a sustained genomic deletion (4342 base pairs) were identified (7303WB/19, 7212WB/19). In this study, we therefore performed in vitro experiments with monocyte-derived macrophages (moMФ) to unravel the phenotypic characteristics of these deleted viruses. Both 7303WB/19 and 7212WB/19 presented a lower growth kinetic in moMФ compared to virulent Sardinian 26544/OG10, using either a high (1) or a low (0.01) multiplicity of infection (MOI). In addition, flow cytometric analysis showed that both 7303WB/19 and 7212WB/19 presented lower intracellular levels of both early and late ASFV proteins. We subsequently investigated whether deleted virus variants were previously circulating in wild boars in Sardinia. In the four years preceding the last genotype I isolation (February 2015–January 2019), other eight wild boar isolates were collected, all belonging to p72 genotype I, B602L subgroup X, but none of them presented a sustained genomic deletion. Overall, we observed the deleted virus isolates in Sardinia only in 2019, at the end of a strong eradication campaign, and our data suggest that it might possess an attenuated phenotype in vivo. A better understanding of ASFV evolution in endemic territories might contribute to development of effective control measures against ASF

Ritorna a far paura la lingua blu

Dopo le epidemie verificatesi sia in Europa che in America, la Bluetongue (lingua blu) ha ripreso a manifestare la sua pericolosità, per quanto riguarda il Continente europeo, oltre che in Italia anche in Spagna, Francia, Grecia e Bulgaria. La profilassi si sta dimostrando alquanto complessa e per ottenere risultati efficaci occorre applicare più metodi contemporaneamente

Expression and antigenic characterization of recombinant <i>Mycoplasma agalactiae</i> P48 major surface protein

The gene encoding the P48 major surface lipoprotein of M. agalactiae has been recently characterised. Since its product plays an important role in the immune response of infected animals, in this study we analysed a recombinant P48 expressed in E. coli. Multiple point mutations were introduced by site directed mutagenesis in order to convert four tryptophan TGA codons, which are a typical feature of the mycoplasma genetic code, into the standard TGG. The mutated p48 gene was subcloned into pGex-2T and expressed in fusion with glutathione-S transferase. Following purification steps, P48 was eluted from carrier protein by thrombin digestion and used in Western blot and indirect ELISA using well-characterised sheep sera. Results demonstrate that specific antibodies against P48 are detected 3 weeks after onset of clinical disease and the recombinant P48 is a diagnostically relevant marker of M. agalactiae infection

P48 major surface antigen of <i>Mycoplasma agalactiae</i> is homologous to a <i>malp</i> product of <i>Mycoplasma fermentans</i> and belongs to a selected family of bacterial lipoproteins

A major surface antigenic lipoprotein of Mycoplasma agalactiae, promptly recognized by the host's immune system, was characterized. The mature product, P48, showed significant similarity and shared conserved amino acid motifs with lipoproteins or predicted lipoproteins from Mycoplasma fermentans, Mycoplasma hyorhinis, relapsing fever Borrelia spp., Bacillus subtilis, and Treponema pallidum

<i>Dermatophilosis</i> in sheep: first report in Italy, experimental reproduction and evaluation of immune response

A severe exudative-crustous and proliferous dermatitis in a 2 year old sheep caused by Dermatophilus congolensis (observed for the first time in Italy), is reported. The disease was reproduced experimentally in sheep, goats, rabbits and guinea pigs, whose skin was treated in different ways before infection. E.L.I.S.A. and Immunoblotting tests carried out in experimentally infected sheep, showed the antigenic complexity of the pathogen and the existence of cross-immunity to the protein components. Intradermoreaction tests were carried out in all animals. The development of a positive reaction only in rabbits and guinea pigs, confirmed that these animals have a cellular immunity against Dermatophilus congolensis

Malattie infettive degli animali

Le malattie infettive degli animali rappresentano una parte rilevante della sanità pubblica, sia per motivi sanitari sia economici, e storicamente sono state tra i primi problemi di popolazione a dover essere gestiti. I contenuti del libro, scelti in base ai programmi didattici svolti in diverse Facoltà italiane, comprendono un’ampia serie di argomenti: una parte introduttiva sui concetti di epidemiologia, sorveglianza, diagnosi, profilassi, controllo ed eradicazione delle malattie infettive, per poi passare in rassegna le diverse patologie che colpiscono gli animali da reddito, da compagnia, i lagomorfi e gli esotici. Vengono anche trattati argomenti riguardanti le principali malattie emergenti e le zoonosi. La stesura del volume ha coinvolto un gran numero di docenti, ricercatori (di diverse sedi universitarie) e professionisti, ciascuno con la sua specifica competenza, con l’obiettivo di produrre un testo condiviso. “Malattie infettive degli animali”, volume di agile e facile consultazione, mette a disposizione di studenti, ricercatori e docenti un’opera completa e di indubbia utilità, con lo scopo di fornire gli strumenti teorici e pratici per la sorveglianza, la diagnosi e il controllo delle malattie infettive