**4. Discussion**

In this paper the presence of nodular proliferative and crusted lesions, was observed in feet de 45 (100%) de 60 bay-winged cowbird (*Gnorimopsar chopi*) examined. Some of them also had lesions on the beak and wings.

Fig. 6. Ultrathin sections of the scabs fragments reveling type B electron dense inclusions bodies with viral particles budding of dense amorphous material (arrow). Fig. 6 - Bar: 600

In the immunocytochemistry technique, the antigen-antibody interaction was strongly

In this paper the presence of nodular proliferative and crusted lesions, was observed in feet de 45 (100%) de 60 bay-winged cowbird (*Gnorimopsar chopi*) examined. Some of them also

enhanced by the dense gold particles over the viruses (Fig. 9, arrow).

nm.

Fig. 7. Bar: 430 nm.

**4. Discussion** 

**3.3 Immunocytochemistry technique** 

had lesions on the beak and wings.

Fig. 8. Ultrathin sections of the skin fragments revealing fibrillar inclusions constituted by groups of fibrils or lamelae (arrow). Bar: 320 nm.

Fig. 9. In the immunocitoquemistry technique, the antigen-antibody interaction was strongly enhanced by the dense colloidal gold particles (arrow) over the avipoxvirus particles. Bar: 130 nm.

Lesions similar have been described in other birds (Goodpasture & Anderson, 1961; Trapp, 1980; Van Riper et al., 2002; Catroxo et al., 2009).

Lesions can interfere wich perching when occur on the feet or legs and frequently lead to development of secondary infections (Van Riper III & Forrester, 2007).

Clinical signs of lack appetite, weight lost, difficulties of locomotion, diarrhea, dehidratation and death were observed in all birds of this study. This has been documented in similar occurrence of avianpox in other avian species (Mc Donald et al., 1981; Docherty et al., 1991; Ostrowski et al.., 1995; Gerlach et al., 1998; Kreuder et al., 1999; Catroxo et al., 2009).

Detection of Poxvirus Using Transmission Electron

Assoc., 65 (1): 23-5, 1994.

**5. References** 

2006.

*25:*454-62, 1981.

*35 (1):* 244-7, 1991.

1969. 666p.

9:409-14, 1961.

*Morphol., 27(2)*:577-585, 2009.

Anim. Sci., 29: 449-4, 2005.

Microscopy Techniques During Outbreak in Bay-Winged Cowbird (*Gnorimopsar chopi*) 565

Allwright, D.M.; Burger, W.P.; Geyer, A. & Wessles, J. Avian pox in ostriches. J. S. Afri. Vet.

Brenner, S. & Horne, R. W. A negative staining method for high resolution electron

Bohls, R.L.; Linares, J.A.; Gross, S.L.; Ferro, P.J.; Silvy, N.J.; Collisson, E.W. Phylogenetic

Carter, J.K.V. & Cheville, N.F. Isolation of surface tubules of fowlpox virus. *Avian Dis.,* 

Catroxo, M. H. B.; Pongiluppi T.; Melo, N. A.; Milanelo, L.; Petrella, S.; Martins, A. M. C. P.

Docherty, D.E.; Long, R. I.; Flickinger, E.L. & Locker, L.N. Isolation of poxvírus from

Ensley, P.K.; Anderson, M.P.; Costello, M.L.; Powell, H.C. & Cooper, R. Expornitic of avian

Friend, M.; Franson, J.C. & Ciganovich, E.A. (Eds). *Field Manual of Wildlife Diseases: General* 

Gonzales-Santander, R. Técnicas de microscopia eletrônica en biología. Madrid. Ed. Aguilar,

Goodpasture, E.W. & Anderson, K. Isolation of a wild avian pox virus inducing both cytoplasmatic and nuclear inclusions. Am. J. Pathol., 40:437-53, 1961. Gülbahar, M.Y.; Çabalar, M. & Boynukara, B. Avipoxvirus infection in quails. Turk. J. Vet.

Gustafson, C.R.; Bickford, A.A.; Cooper, G.L. & Charlton, B.R. Sticktight fleas associated with fowl pox in a backya chicken flock in California. Avian. Dis., 41: 1006-9, 1997.

Luft, J. A. Improvements in an epoxy resin embedding methods. J. Biophys. Biochem. Cytol.,

Moss, B. Poxviridae: The viruses and their replication. In: Virology, Philadelphia, USA, 5 th.

OIE. Manual of Diagnostic Tests & Vaccines for Terrestrial Animal. In: Fowlpox. França,

Madeley, C. R. Electron microscopy and virus diagnosis. J. Clin. Pathol., 50:454-6, 1997. Metz, A.L.; Hatcher, L.; Newman, A.J. & Halvorson, D.A. Venereal pox in breeder turkeys in

Hayat, M. A. & Miller, S. E. Negative Staining. Mc. Graw-Hill Publ. Company., 1990. 235p. Kim, T. & Tripathy, D.N. Evatuation of pathogenicity of avian poxvirus isolates from endangered Hawaiian wild birds in chickens. Avian Dis., 50(2):288-91, 2006. Knutton, S. Electron microscopical methods in adhesion. Meth. Enzymol., 253:145-58, 1995. Kulich, P.; Roubalová, E.; Dubská, L.; Sychra, O.; Smid,. B.; Literák, I. Avipoxvirus in

blakcaps (Sylvia atricapilla). Avian Pathol., 37(1):101-7, 2008.

pox in a zoo. *J. Am. Vet. Med. Assoc., 173:*1111-4, 1978.

States Geological Survey, 1999. pp.163-170.

Minnesota. Avian Dis., 29 (3): 850-3, 1985.

Paris, 6 th ed., 2008. pp. 531-7.

Ed. Lippincot Williams & Wilkins, 2007. pp.2905-75.

analyses indicate little variation among reticuloendotheliosis viruses infecting avian species, including the endangered Attwater's prairie chicken. *Virus Res., 119:*187-94,

F.; Rebouças, M. M. Identification of poxvirus under transmission electron microscopy during outbreak period in wild birds, in São Paulo, Brazil. *Int. J.* 

debilitating cutaneous lesions on four immature grackles (*Quiscalus sp*). *Avian Dis.,* 

*Field Procedures and Diseases of Birds.* Washington, Ed.Washington D.C., United

microscopy of viruses. Biochem. Biophys. Acta., 34:103, 1959.

By means of the negative staining technique, poxvirus particles were identified in suspension of skin fragments, as been seen in other cases (Tantawi et al., 1981; Terragino et al., 1981; Docherty et al.,1991; Allwright et al., 1994; Gulbahar et al., 2005; Kulich et al., 2008; Catroxo et al., 2009; Shivaprasad et al., 2009).

In the present ultrastructural study, intracytoplasmic inclusions bodies of different types were found. The analysis of ultrathin sections of skin lesions showed the presence of type A or Bollinguer intracytoplasmic inclusion bodies, containing oval, mature or complete viral particles, measuring on the average, 330 x 260 nm, with an inner dumbell-shaped core. These morphological characteristics described by us were similar to those found in other birds species (Tripathy & Reed, 2003; Smits et al., 2005; Beyut & Haligur, 2007; Kulich et al., 2008; Saito et al., 2009; Shivaprasad et al., 2009; Catroxo et al., 2009).

The observation of immature intracytoplasmic particles, budding of a dense amorphous material or type B inclusion body, agrees with findings of other authors (Purcell et al., 1972; Thiele et al., 1979, Sadosiv et al., 1985; Catroxo et al., 2009).

We also observed fibrilar intracytoplasmic inclusions, constituted by groups of fibrils or lamellae, disposed in groups varying from 2 to 7, occasionally showing cross striations. These ultrastructural aspects were also reported by other authors in avipoxvirus studies (Catroxo et al., 2009).

The avian poxvirus were intensely labeled by collodial gold, when we applied the immunocitoquemistry method, also used to detect these viruses in skin lesions in other passerines (Catroxo et al., 2009).

All birds in our study were sudden death. During avipox outbreak , mortality can reach 80 to 100% in canaries and other finches in contrast with lower mortality observed in chicken and turkey (Tripathy & Reed, 2003).

The increased population of mosquitoes during the summer at the place of the outbreak probably contributed to the sudden onset of the outbreak. Other sucking insects such flea and mite are considered to be important vectors for the spread of avian poxvirus (Ritchie et al., 1994; Gustafson et al., 1997).

Prevention and control measures, such vaccination, quarantine for new birds, control of arthropods, prevention and reduction of stress, cleaning and disinfecting of cages should be instituted to farms and zoos (Ritchie et al., 1994).

It is very important to confirm the presence of the ethiological viral agent in the samples. Infection as other agents, such pappiloma virus, mites, flies and mycotoxins may produce like lesions in the skin (Trypathy & Reed, 2003).

Our results obtained through the techniques mentioned in this study to detect avipoxvírus particles in these birds, allowed rapid preventive measures to control the disease, preventing spread to commercial breeder farms and aviaries with economic losses thus protecting and preserving these endangered species.

To our knowledge, this is the first report of avian poxvirus infection in bay-winged cowbird (*Gnorimopsar chopi*).

## **5. References**

564 A Bird's-Eye View of Veterinary Medicine

By means of the negative staining technique, poxvirus particles were identified in suspension of skin fragments, as been seen in other cases (Tantawi et al., 1981; Terragino et al., 1981; Docherty et al.,1991; Allwright et al., 1994; Gulbahar et al., 2005; Kulich et al., 2008;

In the present ultrastructural study, intracytoplasmic inclusions bodies of different types were found. The analysis of ultrathin sections of skin lesions showed the presence of type A or Bollinguer intracytoplasmic inclusion bodies, containing oval, mature or complete viral particles, measuring on the average, 330 x 260 nm, with an inner dumbell-shaped core. These morphological characteristics described by us were similar to those found in other birds species (Tripathy & Reed, 2003; Smits et al., 2005; Beyut & Haligur, 2007; Kulich et al.,

The observation of immature intracytoplasmic particles, budding of a dense amorphous material or type B inclusion body, agrees with findings of other authors (Purcell et al., 1972;

We also observed fibrilar intracytoplasmic inclusions, constituted by groups of fibrils or lamellae, disposed in groups varying from 2 to 7, occasionally showing cross striations. These ultrastructural aspects were also reported by other authors in avipoxvirus studies

The avian poxvirus were intensely labeled by collodial gold, when we applied the immunocitoquemistry method, also used to detect these viruses in skin lesions in other

All birds in our study were sudden death. During avipox outbreak , mortality can reach 80 to 100% in canaries and other finches in contrast with lower mortality observed in chicken

The increased population of mosquitoes during the summer at the place of the outbreak probably contributed to the sudden onset of the outbreak. Other sucking insects such flea and mite are considered to be important vectors for the spread of avian poxvirus (Ritchie et

Prevention and control measures, such vaccination, quarantine for new birds, control of arthropods, prevention and reduction of stress, cleaning and disinfecting of cages should be

It is very important to confirm the presence of the ethiological viral agent in the samples. Infection as other agents, such pappiloma virus, mites, flies and mycotoxins may produce

Our results obtained through the techniques mentioned in this study to detect avipoxvírus particles in these birds, allowed rapid preventive measures to control the disease, preventing spread to commercial breeder farms and aviaries with economic losses thus

To our knowledge, this is the first report of avian poxvirus infection in bay-winged cowbird

Catroxo et al., 2009; Shivaprasad et al., 2009).

(Catroxo et al., 2009).

passerines (Catroxo et al., 2009).

and turkey (Tripathy & Reed, 2003).

al., 1994; Gustafson et al., 1997).

(*Gnorimopsar chopi*).

instituted to farms and zoos (Ritchie et al., 1994).

like lesions in the skin (Trypathy & Reed, 2003).

protecting and preserving these endangered species.

2008; Saito et al., 2009; Shivaprasad et al., 2009; Catroxo et al., 2009).

Thiele et al., 1979, Sadosiv et al., 1985; Catroxo et al., 2009).


**29** 

**Doppler Ultrasonography for Evaluating** 

Ergot alkaloids are produced by non-spore producing fungal endophytes that infect certain species of grasses, most notably tall fescue [*Lolium arundinaceum* (Schreb.) Darbysh.] and perennial ryegrass (*Lolium perenne* L.), and the spore producing *Claviceps* spp. that infect seed heads of certain grasses and particularly the cereal grains [rye (*Secale cereal* L.), barley (*Hordeum vulgare* L.), wheat (*Triticum aestivum* L.), , and oats (*Avena sativa* L.)] (Strickland et al., 2011). Ergot alkaloids induce a toxicosis in grazing livestock, with symptoms in cattle that include rough hair coats during the warm season, severe heat stress in warm and humid temperatures, reduced dry matter intake, agalactia, and poor reproductive and weight gain performance (Porter & Thompson, 1992; Paterson et al., 1995). Sheep grazing endophyte-infected fescue also can have elevated core body temperatures in warm and humid environments (Zanzalari et al., 1989; Hanna et al., 1990), and reduced dry matter intakes (Chestnut et al., 1992; Aldrich et al., 1993). The most pronounced effect on horses is observed with broodmares, which can exhibit prolonged gestation and agalactia (Cross et al., 1995). Symptomatology of the malady are reflective of alterations in hormone profiles (Porter & Thomson, 1992; Browning et al., 1997, 1998) and reductions in blood flow to peripheral tissues caused by interactions of ergopeptine ergot alkaloids with biogenic amine receptors in the vasculature (Oliver et al., 1998) to induce persistent vasoconstriction and restrict regulation of core body temperature by the sympathetic nervous system (Oliver, 1997). Consequently, lack of thermoregulation by livestock exposed to ergopeptines are extremely vulnerable to heat and cold stresses (Hemken et

Ergot alkaloids contain a common tetracycline ergoline ring structure and there are 3 different classes: 1) clavine alkaloids, 2) lysergic acid and its derivatives, and 3) ergopeptines (Lyons et al., 1986; Bush and Fannin, 2009; Strickland et al. 2011). Ergopeptines exert the greatest influence on the vasculature, with ergovaline being the ergopeptine of highest concentration in tall fescue (Lyons et al., 1986) and with a demonstrated high potency as a

**1. Introduction** 

al., 1981; Al-Haidary et al., 2001).

vasoconstrictor (Klotz et al., 2006).

**Vascular Responses to Ergopeptine** 

**Alkaloids in Livestock** 

G.E. Aiken and J.R. Strickland *USDA-Agricultural Research Service, Forage-Animal Production Research Unit,* 

*Lexington, KY,* 

*USA* 

