Pages Circo Pozzi

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Pozzi S. P. , Yadin H. ; Lavi J ; Pacciarini M . Alborali L . 'Milano, Italy; Kimron Veterinary Institute, Bet Dagan, Israel; Shavey Zion, Israel; IZSLER Animal Health Institute, Brescia, Italy
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Correspondence: paolo.pozzi@, tel: +39 335 7720466, fax + 39 02 4045549
Keywords: Swine, PCV2, P M W S , PDNS, serology, virology, PCR, IHC.
During the years 2006-2007, an increase of respiratory and enteric problems in piglets after weaning was reported from pig herds in western Galilee. These problems were accompanied by increased mortality and a lower growth rate of the affected piglets. A reduced growth rate was also noted in older pigs at these farms. Sick piglets were bled on three affected farms, and on necropsy, lungs, inguinal and mesenteric lymph nodes were collected from carcasses, and submitted to laboratory examination. Evidence for the presence of porcine circovirus type 2 (PCV2), was based on positive ELISA serology; virus isolation; polymerase chain reaction (PCR) and immunohisto chemistry (IHC) on the lymphoid tissues. To our knowledge this is the first case on PCV2 identification in Israel pigs.
Porcine circovirus type 2 (PCV2) is a single-stranded DNA virus, belonging to genus C i r c o v i r u s , family C i r c o v i r i d a e . It is considered ubiquitous and both domestic and wild pigs are the natural hosts (1). Oral-nasal infection is the most frequent and natural route of transmission (1). Alternatively, infected sows may shed virus into the colostrum and the milk, thereby infecting piglets by the oral route (2). PCV2 is resistant at acid p H (pH<3) and stable for 15' at 70°C (3). Chlorexidine, formaldehyde, iodine and alcohol-based disinfectants can reduce the virus titer at room temperature within 10 minutes (1). Despite the ubiquity of PCV2 in almost all pig herds worldwide, only in some cases does clinical disease develop presenting 5-30% morbidity and 50-60% mortality among the affected pigs. In such cases, there is a dramatic worsening of the health of weaned piglets, from between 8 to 16 weeks of age, represented by a post-weaning multi-systemic wasting syndrome (PMWS) with emaciation, jaundice, reduced growth (Fig. 1,3), hind legs and dermatitis (porcine dermatitis associated with a nephropathy syndrome (PDNS) (Fig. 4,5), (1,4,5,6). Both respiratory and enteric disease appear in fattening animals, while in adult herds, ,reproductive problems affect breeders. A l l these syndromes are clinically indistinguishable from other diseases. Thus, P C V 2 involvement in P C V A D , PMWS, PDNS must be confirmed by the virus isolation (1). The virus may be isolated on a pig kidney cell line, and a polymerase chain reaction (PCR) is available. Immuno histochemistry (IHC); immunofluorescence (IF) and serology (ELISA competition) can be used to detect P C V 2 infection. Histological evaluation of lymphoid cells depletion in PCV2-target lymphoid organs
such as the inguinal or mesenteric lymph nodes has been reported. The purpose of this article is to describe PCV2-associated clinical signs in 3 swine farms of north Israel.
MATERIALS AND Herd history
In 3 far-to-finish swine farms located in western Galilee, an increased incidence of respiratory and enteric problems was encountered from weaning at 5 weeks old and 12-14 weeks. There was high mortality (12-20%), and a decreased growth rate in piglets. A n increased proportion of fattening pigs suffered from dermatitis that was unresponsive to antibiotic treatment. A tentative diagnosis of PCV2 infection was made
Six blood samples from affected chronic piglets aged 8 to 12 weeks were collected from each farm.. In addition, 10 dead piglets were necropsied; and inguinal and mesenteric lymph nodes and lung tissue were collected from each piglet .The samples were processed at the Virology Department of the Kimron Veterinary Institute, Bet Dagan, and then submitted for further laboratory investigation at the IZSLER Animal Health Institute , Brescia, Italy.
Laboratory investigations:
A l l sera were submitted to serological testing for Porcine Respiratory and Reproductive Syndrome virus (PRRSV), Aujeszky Disease virus (ADV), Swine Influenza virus (SIV), PCV2 and erysipelas. The pooled blood samples were also tested for PRRSV-antigen.
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Virology: PCV2 was performed using an in-house sandwich-ELISA antigen test using a monoclonal antibody at the second passage on a pig kidney cell line. Polymerase Chain Reaction: The P C R test was performed on pooled tissue homogenates from each piglet (9). The following primers were designed to amplify 263 nucleotides sequence from opening reading frame 2 (ORF 2) of PCV2 associated with P M W S (10): - CF8 5 ' - T A G G T T A G G G C T G T G G C C T T - 3 ' 1323 to 1342) - CR8 5 ' - C C G C A C C T T C G G A T A T A C T G - 3 ' 1567 to 1586). (nucleotides
Laboratory results: Virus isolation: The test could be performed only on tissues from 5 out of 10 piglets and from only one farm, due to poor preservation of the material. A l l the 5 piglets yielded PCV2 virus.. PCR: Four out of the 10 pooled lymphoid tissue homogenates originating from the 3 farms were PCR positive; IHC Two of the 10 piglets were PCV2 positive; representing 2 out of the three farms. Serology: A l l sera were PCV2-Elisa positive, average titer = log 3.72 sd = 0.57. A l l the 3 farms tested positive.
Immunohistochemistry: The IHC test were performed with a PCV2 monoclonal antibody on 4 jx tissue slices, then compared with PCV2-PCR and i n s i t u hybridization (ISH) positive and negative reference samples (1,7). Serology: Sera were tested using an in-house PCV2 ELISA-competition test,. The results are given as L o g of reciprocal of last dilution, starting 1:10=1 L o g . The results are summarized as mean titer; standard deviation (sd) and percentage of positive samples.
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The wasting disease of the pigs in the weaning area of the farms, together with the poor performance of the fattening pigs, the presence of dermal lesions and necropsy findings indicated the involvement of PCV2 From an epidemiological point of view, it can be considered that more than 50% of the sow population and the related pig production are concentrated in a small and densely populated area (11). Moreover, the farms are located fence to fence and are so interconnected that they represent a single epidemiological entity (12). This explains how P C V 2 could spread among them so rapidly. In connection with the clinical signs induced by PCV2, PDNS must be differentiated from: Classical and African Swine Fever (1) both are notifiable diseases, also considering interstitial nephritic lesions induced by PCV2 in PDNS; Erysipelas, by E . r h u s i o p a t h i a e , a notifiable, zoonotic disease Pityriasis rosea (pustular psoriaform dermatitis), an apparently inherited disease, self-limiting, but not resembling human pityriasis. Inactivated vaccines against the P C V 2 for the use in breeders and piglets are registered and used in several European countries, as well as the USA and Canada. They are generally characterized by their cost-effective results in terms of mortality and in reduction of the incidence in clinical signs. The sow vaccines may be indicated for the passive protection
The PRRSV, ADV, SIV, and Erysipelas examinations were negative. Clinical signs: The weaned piglets appeared apathic and emaciated (fig.l). They were coughing; and had enlarged inguinal lymph nodes at palpation (fig 2). Runts, poor doers and anemic pigs were seen in the fattening areas (fig 3). Examination of the herds showed a few cases of fattening pigs affected by irregular, purple,necrotizing dermal lesions, primarily over the hind legs and perineum (fig. 4 and 5). Post mortem of dead piglets showed poor bodily condition, enlargement of inguinal and the mesenteric lymph-nodes (fig 6, 7), interstitial pneumonia (fig 8), and in some cases, enteritis.
Table 1: A summary of PCV2 laboratory tests performed on piglet tissues.
Piglets Test performed Virological * Immunofluorescence Immuno Histo Chimic Polimerase Chain Reaction * two p a s s a g e s o n p i g k i d n e y c e l l s l i n e
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neg neg
10 7 ! 8 1 9 not performed \ p o s pos pos p o s pos neg neg posIneg neg pos p o s pos neg pos p o s j n e g neg neg neg]neg neg pos pos p o s j n e g pos neg neg i neg neg , i
i LL i
ARTICLES of the piglets in the presence of early clinical signs of PCV2 in the weaning period, and up to the age of 6-8 weeks. The vaccination of piglets at the third week of age is strongly indicated when there are PCV2 problems at the growingfattening stages. As an alternative to vaccination, the application of good management practice (13), rescheduling of other vaccination times (1) and dietary changes to include certain supplements (14), could give some benefit in affected farms. To the authors' knowledge, this is the first report of PCV2associated disease of Israel pigs.
1. Segales J, Allan G, Domingo M , "Porcine Circovirus Diseases", in Diseases of swine, 9th Ed., 14:299-307, 2006. Shibata I , Okuda Y , Kitajima K , Asai T , "Shedding of P C V into colostrum of sows." J. Vet Med B 53(6):278-280 ,2006. Allan G, Phenix K , Todd D, McNulty M , "Some biological and physico-chemical properties of porcine circovirus". J. Vet. Med. B 41:17-26, 1994. Segales J, Allan G , Domingo M , " P C V D " , in Animal Health Research reviews - Conference of Research Workers in Animal Diseases, 6(2):119-142, 2005. Calsamiglia M , Fraile L, Espinal A , Cuxart A , Seminati C, Martin M , Mateu M , Domingo M , Segales J, "Sow P C V 2 status effect on litter mortality in P M W S " , Res in Vet Sci 82(3):299-304, 2007. Martinez J, Segales J, Aduriz G, Atxaerandio R, Jaro P, Ortega J, Peris B , Corpa J, "Pathological and aetiological studies of multifocal interstial nephritis in wasted pigs at slaughter", Res in Vet Sci 81(1): 92-98, 2006. Sarli G , Ostanello F, Morandi F, Nigrelli A , Alborali L , Dottori M , Vezzoli F, Barigazzi G, Sala V, Leotti G , "Application of a protocol for the P M W S diagnosis" X X X I I I SIPAS Meeting, 189-202, 2007. Opriessnig T, Halbur P, " P C V A D in North America" X X X I I I SIPAS Meeting, 91-96,2007. Alborali G , Salogni C, Zanoni M , Pacciarini M , Cordioli P, Martelli P, "Comparative study and virological investigations on P M W S suspected and non suspected herds in Italy", 4th Emerging and Re-emerging Pig Dis. Symp. 216-217, 2003. Larochelle R, Bielanski A , Muller P, Magar R, " P C R detection and evidence of shedding of PCV2 in boar semen" J. Clin. Microb. 38, 12:4629-4632,2000. Israel Veterinary Services .Annual Report, 2006. Elad D , Samina I, Nankin M , Barigazzi G, Foni E , Guazzetti S, Pozzi SP, "Serological monitoring towards antigens responsible of respiratory diseases in fattening pigs in Israel", Proc X X V I I I SIPAS, Piacenza (I), 155-160, 2002. Madec F, Eveno E, Morvan P, Hamon L , Blanchard P, Cariolet R, Amenna N , Morvan M , Truong C, Mahe D , Albina E , Jestin A " P M W S in pigs in France: clinical observation from follow-up studies on affected farms" Livest. Prod. Sci. 63:223-233, 2000. Baebko P, Hassing A , Olsen P, Lorenzen B , Wachman H , Lauridsen C, "Vitamin E and postweaning mortality in P M W S affected herds" 18th IPVS, 1-62, 2004.
Ackowledgment: Dr. E . Greva is acknowledged for the review of the text.
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11. 12.
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Fig 6: Enlarged mesenteric lymph nodes
Fig 2: Enlarged inguinal lymph nodes.
Fig 8: Interstitial pneumonia
Fig 3: Growth retardation at fattening (approx 4 months of age) all the pigs in the picture are the same age.
Fig 5: Severe signs of PDNS in fattening pigs
Fig. 4: Mild signs of PDNS in fattening pigs
Fig 7: Enlarged inguinal lymph nodes
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