Abstract Book of EAVLD2012 - eavld congress 2012

Recommendations

Info

S3 - P - 11 DOLPHIN MORBILLIVIRUS INFECTION IN A CAPTIVE HARBOR SEAL (PHOCA VITULINA) Sandro Mazzariol 1 , Francesco Grande 2 , Cristina Pilenga 2 , Paola Modesto 3 , Cristina Biolatti 3 , Giovanni Di Guardo 4 , Alessandra Mondin 5 , Simone Peletto 3 , Pier Luigi Acutis 3 1 Department of Comparative Biomedicine and Nutrition (BCA), University of Padua, Legnaro, Italy 2 Zoomarine Italia, Rome, Italy 3 Istituto Zooprofilattico Sperimentale del Piemonte, Liguria e Valle d’Aosta, Turin, Italy 4 Department of Comparative Biomedical Sciences, University of Teramo, Teramo, Italy 5 Department of Animal Medicine and Health, University of Padua, Legnaro, Italy morbillivirus, DMV, harbor seal, transmission, phylogeny Introduction During the last 25 years morbilliviral infections have caused dramatic mortalities among several aquatic mammal species and populations worldwide (1). In particular, Dolphin Morbillivirus (DMV) represents a major biological threat for free-ranging cetaceans. In the Mediterranean Sea, DMV has been responsible for two epidemics, the first of which caused a dramatic mortality among striped dolphins (Stenella coeruleoalba) between 1990 and 1992, while the second one was firstly reported in the western Mediterranean in 2006-2008, moving thereafter eastward along the French and the Italian coastlines. In June 2011, one bottlenose dolphin (Tursiops truncatus) and one striped dolphin stranded along the Tyrrhenian coastline, close to Rome. Both animals were found molecularly (RT-PCR) positive to Morbillivirus. Personnel from a marine park helped the stranding network with the bottlenose dolphin, which stranded alive and died after one day of rehabilitation. Standard quarantine protocols were applied, in order to avoid any contact with cetaceans kept in the park. On July 25 th , 2011, an 8-years-old male harbor seal (Phoca vitulina) hosted inside the aforementioned park was submitted for post-mortem examination after a short history of anorexia, tremors, abdominal contractions, and polyuria with hypothermia and vomit in the last phases. Materials & methods A complete post-mortem investigation was carried out the day after death. Samples from main organs (brain, lungs, heart, liver, kidneys, GI tract, spleen, lymph nodes) were fixed in 10% neutral buffered formalin, paraffin-embedded and routinely processed for histopathology. Samples from intestine and kidneys were submitted for routine microbiological exams while cerebral, hepatic, cardiac, splenic, pulmonary and lymph node tissues were preserved frozen for molecular analyses. Molecular investigations were carried out to look for the presence of Canine Distemper Virus (CDV), Phocine Distemper Virus (PDV) and Cetacean Morbillivirus (CeMV) nucleic acids. Multiple sequence alignment was performed with the software BioEdit using CLUSTAL W. An identity matrix was created to compare sequenced products with morbillivirus sequences deposited in GenBank. Phylogenetic analysis was performed using MEGA 5.0 software. The tree topology was inferred by the maximum likelihood method with Kimura 2-parameter model. The reliability of the topology was tested by bootstrapping 1,000 replicates. Results Necropsy revealed a mild gastro-enteritis and splenitis due to Aeromonas hydrophyla, along with hepatic and renal necrosis presumably due to endotoxemia. Besides these pathological findings, a generalized and diffuse lymph node enlargement was observed, along with a severe meningeal hyperemia and choroidal edema. Histologically, a severe and diffuse reactive lymphadenopathy was apparent, with lesions being characterized by lymphoid cell depletion, hyalinosis, and several multinucleate syncytia. A mild, multifocal, chronic choriomeningitis with white matter spongiosis and demyelination were also apparent, with eosinophilic inclusion bodies being also found in the cytoplasm of brain neurons and glial cells. Furthermore, the lung showed a mild, chronic bronchointerstitial pneumonia, with simultaneous evidence of bronchiolar epithelial cell hyperplasia/hypertrophy. Pathological findings suggested a morbilliviral infection, which was confirmed by RT-PCR, with CeMV-specific genome sequences being subsequently amplified from the brain of this seal. CDV and PDV molecular analyses were negative. Similarity was 97.6-99.5% with known DMV sequences and 85.9-89.6% with other CeMV members (Porpoise Morbillivirus and the tentatively named Pilot Whale Morbillivirus). Phylogenetic analysis confirmed clustering of the harbor seal CeMV sequence with previously reported DMV sequences from cetacean species (Figure 1). 53 75 DMV AJ608288 95 DMV AY586536 DMV Z36978 DMV AJ224705 94 DMV Sc/2007 HQ829973 75 DMV Gme/2007 HQ829972 Harbor seal PWMV 20E FJ842382 90 PMV IRL88 FJ648457 PMV 2990 AY586537 MeV UK140H94 U29285 69 RPV LA96 JN234010 PPRV ICV89 EU267273 PDV AF479277 CDV EU716337 SeV X56131 0.2 Figure 1: Maximum Likelihood dendrogram, based on the Kimura 2-parameter model, illustrating haemagglutinin gene (H) nucleotide sequence relationships of morbilliviruses. Discussion & conclusions During the last epidemic outbreak, morbilliviral infection was reported in several species besides striped dolphins, namely pilot whales (Globicephala melas), bottlenose dolphins, Risso’s dolphins (Grampus griseus), and fin whales (Balaenoptera physalus). Although the susceptibility of these cetacean species was already known, no reports other than the present one have ever described DMV transmission to harbor seals. Indeed, infection of pinniped species with morbilliviruses of cetacean origin was previously speculated on the basis of serological cross-reactivity and sequence homology (2). On a global scale, mortality events in both this and in other pinniped species such as grey seals (Halichoerus grypus), Bajkal seals (Pusa siberica), and Caspian seals (P. caspica) have been respectively linked to PDV and to CDV, with the former agent being more closely related to CDV than to any other known morbillivirus, while DMV appears to be more closely related to Rinderpest Virus (RPV). Apart from their diverging phylogeny and their phylogenetic distances, DMV and CDV could share similar epidemiologic features. This hypothesis is strongly supported by the simultaneous evidence of morbilliviral infection in the two freeranging dolphins found beached ashore near Rome before this captive seal died. Therefore, a possible route of virus (DMV) entry into the facility could have been represented by either the personnel and/or the instruments used for the rehabilitation of the Morbillivirus-infected bottlenose dolphin that was found stranded alive one month before. In conclusion, this report emphasizes that, apart from cetaceans, also pinniped species such as harbor seals are susceptible to DMV infection and related mortality. Therefore, strict quarantine procedures should be enforced also in structures where pinnipeds are kept when injured or stranded cetaceans are hospitalized within these facilities. References 1. Di Guardo G, Mazzariol S, Fernández A. (2011). Biologically threatened dolphins and whales. Environmental Microbiology, 13(11), 2833-4. 2. Van de Bildt MWG, Martina BEE, Vedder EJ et al. (2000). Identification of morbilliviruses of probable cetacean origin in carcases of Mediterranean monk seals (Monachus monachus). Veterinary Record, 10, 691-4.
S3 - P - 12 PRELIMINARY VALIDATION OF THE ID SCREEN AFRICAN SWINE FEVER INDIRECT ELISA BASED ON THREE RECOMBINANT ASF PROTEINS Pourquier, P. 1 , Loic, C. 1, 1 IDvet, Montpellier, France African Swine Fever, diagnostics, serology, ELISA Introduction African swine fever (ASF) is a highly contagious, viral haemorrhagic disease of pigs, warthogs, European wild boar and American wild pigs caused by the African Swine Fever Virus (ASFV). There is neither treatment nor vaccine for ASF. To prevent introduction and spread of the disease, countries implement strict surveillance, control and eradication programs which require accurate and reliable diagnostic tests. In this context, IDvet has launched the ID Screen ® African Swine Fever Indirect ELISA. This test detects anti-ASFV antibodies in domestic or wild pigs, in serum, plasma or blood filter paper samples. Unique features of the ID Screen ® African Swine Fever Indirect ELISA include the coating of three recombinant ASFV antigens (P32, P62, and P72), and the ability to use the test with blood filter paper samples as well as serum and plasma. This study presents the preliminary validation data obtained for this ELISA kit on both serum and filter paper samples. References 1. Barderas MG et al, 2000. Serodiagnosis of African Swine Fever using the recombinant protein p30 expresses in insect larvae. Journal of virological methods, 89:129-136. 2. Gallardo C et al, 2009. Recombinant antigen targets for serodiagnosis of African Swine Fever. 16 :1012-1020. 3. Gallardo C et al, 2006. Antigenic properties and diagnostic potential of African Swine Fever virus protein p62 expressed in insect cells. Journal of clinical microbiology, 44 : 950-956. 4. Hutching GH et al, 2006. Indirect sandwich ELISA for antigen detection of African Swine Fever virus : comparison of polyclonal and monoclonal antibodies. Journal of virological methods, 131:213-217. 5. OIE, 2008. Manuel terrestre de l’OIE : African Swine Fever. Chapitre 2.8.1. 6. Sanz et al, 1985. Monoclonal antibodies specific for African Swine Fever Virus proteins. Journal of virology, 54:199-206. Material & methods The ID Screen ® African Swine Fever Indirect is based on three recombinant ASFV antigens (P32, P62, and P72) and an antiswine-HRP conjugate. Analyses were performed as per manufacturer’s specifications. Results Specificity - 556 disease-free sera from domestic pigs (France and Norway), wild boars (France), and Iberian pigs (Spain) were tested. All samples were found negative. - 90 negative animals were tested by both the serum and filter paper protocols. All sera were found negative by both protocols. Sensitivity - 3 sera were tested from pigs vaccinated on day 0 and day 24 with the ASF strain Ourt88/3, challenged on day 42 with a mild ASF strain (ANSES, Ploufragan, France), and bled on day 62 or 63. After challenge, all three animals gave strong positive results with the ID Screen ® African Swine Fever Indirect ELISA. - 8 reference sera provided by the Community Reference Laboratory (CRL), CISA-INIA, Spain were analysed. The ID Screen® African Swine Fever Indirect ELISA accurately identified all sera. - 92 sera from infected herds (Sassari, Sardinia, Italy) were tested in parallel by the ID Screen ® ELISA and the CRL ELISA reagents (CISA-INIA, Spain). Test correlation was found to be 95.7%. The relative sensitivity and specificity were 97.67% and 97.83%, respectively. - 3 sera were titrated and tested by both the serum and filter paper protocols. The measured analytical sensitivity was similar regardless of the sample type tested (serum, Whatman 1 or Whatman 3 filter paper). Discussion & conclusions The ID Screen ® African Swine Fever Indirect ELISA is an efficient and reliable tool for the diagnosis of ASF in wild and domestic species. It is the only commercial ELISA based on 3 recombinant antigens. Both the serum and filter paper test protocols show excellent sensitivity and specificity Collecting blood using filter paper facilitates sample collection in the field. With the ID Screen protocol, filter paper samples may be tested in a 96-well deep-well plate, making sample processing faster and less prone to mix-ups.
Page 2 and 3:
2 nd CONGRESSOFTHEEUROPEAN ASSOCIAT
Page 4:
Welcome Dear colleagues, Welcome to
Page 7 and 8:
worked with many diagnostic compani
Page 9 and 10:
Products and services for scientist
Page 11 and 12:
16:4518.15 20:0023:00 [S1.]Oralpres
Page 13 and 14:
Day4 Wednesday,4July2012 4 th sessi
Page 15 and 16:
Oral presentations “General Sessi
Page 17 and 18:
antigens or attenuated vaccines can
Page 19 and 20:
conditions, e.g. Staphylococcus aur
Page 21 and 22:
S1 - O - 1 ORAL FLUIDS - SAMPLE MAT
Page 23 and 24:
S1 - O - 03 INTRODUCTION RATE OF A
Page 25 and 26:
S1 - O -05 VALIDATION OF A COMPETIT
Page 27 and 28:
S1 - O - 07 SUBTYPING OF SWINE INFL
Page 29 and 30:
S1 - O - 09 VIRAL DIAGNOSIS USING T
Page 31 and 32:
S1 - O - 11 USING ORAL FLUID FOR TH
Page 33 and 34:
S1 - O - 13 THE FIRST YEAR OF OBLIG
Page 35 and 36:
S1 - O - 15 SEROLOGICAL ANALYSIS AN
Page 37 and 38:
S1 - O - 17 RING TEST EVALUATION FO
Page 39 and 40:
Oral presentations “Diagnostics a
Page 41 and 42:
S2 - O - 01 DEVELOPMENT OF A RAPID
Page 43 and 44:
S2 - O - 03 EVALUATION OF RAPID HRS
Page 45 and 46:
Oral presentations “Emerging, re-
Page 47 and 48:
advances, the fundamental tools of
Page 49 and 50:
S3 - O - 02 25 YEARS OF PASSIVE SUR
Page 51 and 52:
S3 - O - 04 EQUINE NOCARDIOFORM PLA
Page 53 and 54:
S3 - O - 06 DETECTION OF SCHMALLENB
Page 55 and 56:
S3 - O - 08 SCHMALLENBERGVIRUS: SER
Page 57 and 58:
S3 - O - 10 DIAGNOSTIC ASPECTS OF S
Page 59 and 60:
Oral presentations “New technique
Page 61 and 62:
acid and organic solvent. Then a dr
Page 63 and 64:
S4 - O - 02 BIOLOG GENERATION III,
Page 65 and 66:
S4 - O - 04 MATRIX-ASSISTED LASER D
Page 67 and 68:
S4 - O - 06 15 MINUTE ELISA USING A
Page 69 and 70:
S1 - P - 01 LAWSONIA INTRACELLULARI
Page 71 and 72:
S1 - P - 03 DETECTION OF PRRSV ANTI
Page 73 and 74: S1 - P - 05 DEVELOPMENT AND EVALUAT
Page 75 and 76: S1 - P - 07 ANALYTICAL EVALUATION O
Page 77 and 78: S1 - P - 09 PTS FOR BVDV ANTIGEN DE
Page 79 and 80: S1 - P - 11 ANTIMICROBIAL SUSCEPTIB
Page 81 and 82: S1 - P - 13 IDENTIFICATION OF GLOBI
Page 83 and 84: S1 - P - 15 ACCURACY OF PARATUBERCU
Page 85 and 86: S1 - P - 17 VIROLOGICAL SURVEILLANC
Page 87 and 88: S1 - P - 19 CHARACTERIZATION OF EXT
Page 89 and 90: S1- P - 21 ANALYSIS OF THE EFFICIEN
Page 91 and 92: S1 - P - 23 RELIABLE AND EARLY DETE
Page 93 and 94: S1 - P - 25 GENOTYPIC VARIABILITY D
Page 95 and 96: S1 - P - 27 PROFICIENCY TESTING (PT
Page 97 and 98: S1 - P - 29 LOW PATHOGENIC AVIAN IN
Page 99 and 100: S1 - P - 31 PRRSV DIAGNOSTICS BY RT
Page 101 and 102: S1 - P - 33 APPLICATION OF A HERD P
Page 103 and 104: S1 - P - 35 DEVELOPMENT OF A BROADL
Page 105 and 106: S1 - P - 37 MONITORING OF BOVINE VI
Page 107 and 108: S1 - P - 39 BOVINE BLOOD DENDRITIC
Page 109 and 110: Poster presentations “Diagnostics
Page 111 and 112: S2 - P - 02 NEW IMMUNOASSAYS FOR DI
Page 113 and 114: Poster presentations “Emerging, r
Page 115 and 116: S3 - P - 02 LIPOPOLYSACCHARIDE-CAPT
Page 117 and 118: S3 - P - 04 DEVELOPMENT OF A SCHMAL
Page 119 and 120: S3 - P - 06 COMPARISON OF CULTURE A
Page 121 and 122: S3 - P - 08 EMERGENCE OF SCHMALLENB
Page 123: S3 - P - 10 SIMULTANEOUS DETECTION
Page 127 and 128: S3 - P - 14 DEVELOPMENT OF MULTISPE
Page 129 and 130: S3 - P - 16 DETECTION OF IBV QX IN
Page 131 and 132: S3 - P - 18 PREVALENCE OF COXIELLA
Page 133 and 134: S3 - P - 20 INTERFERON-GAMMA ASSAY
Page 135 and 136: S3 - P - 22 COMPARISON OF THE PERFO
Page 137 and 138: S3 - P - 24 DEVELOPMENT OF A VIRUS
Page 139 and 140: S3 - P - 26 DETECTION OF EQUINE FOA
Page 141 and 142: S4 - P - 01 MOLECULAR CHARACTERIZAT
Page 143 and 144: S4 - P - 03 USEFULNESS OF LIVE/DEAD
Page 145 and 146: S4 - P - 05 EVALUATION OF THREE ELI
Page 147 and 148: S4 - P - 07 EVALUATION OF DIFFERENT
Page 149 and 150: S4 - P - 09 VALIDATION OF THE ID SC
Page 151 and 152: S4 - P - 11 EFFECTIVE TESTING STRAT
Page 153 and 154: List of Abstracts (in order of numb
Page 155 and 156: S1-P-03 BALLAGI ANDREA S1-P-04 BENI
Page 157 and 158: S3-P-06 KELLER SELINA S3-P-07 KÖNI
Page 159 and 160: List of abstracts (alphabetical ord
show all

Abstract Book of EAVLD2012 - eavld congress 2012

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?