Hepatic Profile of Gallus Gallus Linnaeus, 1758 Experimentally
Total Page：16
File Type：pdf, Size：1020Kb
Veterinary Parasitology 175 (2011) 207–211 View metadata, citation and similar papers at core.ac.uk brought to you by CORE Contents lists available at ScienceDirect provided by Elsevier - Publisher Connector Veterinary Parasitology journal homepage: www.elsevier.com/locate/vetpar Hepatic profile of Gallus gallus Linnaeus, 1758 experimentally infected by Plasmodium juxtanucleare Versiani & Gomes, 1941 Usha Vashist a, Aline Duarte Falqueto a, Danilo Lustrino b, Victor Menezes Tunholi a,b, Vinícius Menezes Tunholi-Alves a,b, Marcos Antônio José dos Santos c, Marta D’Agosto d, Carlos Luiz Massard a, Jairo Pinheiro b,∗ a Curso de Pós-Graduac¸ ão em Ciências Veterinárias, Departamento de Parasitologia Animal, Instituto de Veterinária, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ 23.890-000, Brazil b Departamento de Ciências Fisiológicas, Instituto de Biologia, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ 23.890-000, Brazil c Departamento de Biologia Animal, Instituto de Biologia, Universidade Federal Rural do Rio de Janeiro, Seropédica, RJ 23.890-000, Brazil d Departamento de Zoologia, Instituto de Ciências Biológicas, Universidade Federal de Juiz de Fora, Juiz de Fora, MG 36036-900, Brazil article info abstract Article history: One of the species that causes avian malaria is Plasmodium juxtanucleare. It is commonly Received 24 June 2010 found in poultry, especially when the birds receive food free of coccidiostats. Since industrial Received in revised form 14 October 2010 and organic poultry breeding is increasing in the world and few studies have been conducted Accepted 14 October 2010 examining the clinical parameters of both healthy and infected birds, this work evaluated whether the infection caused by P. juxtanucleare in Gallus gallus provokes alterations in the Keywords: birds’ hepatic profile. We analyzed the activity of ALT and AST and carried out histological Avian malaria analyses of liver sections of infected fowls by intracelomic inoculation with infected blood Aminotransferases Plasmodium juxtanucleare from a donor fowl with a parasite load of around 7%. The infected birds’ parasite load was evaluated during 45 days by means of blood smears. There was a positive correlation between the increase in parasite load and higher ALT activity in the infected fowls, but there was no significant variation of the AST activity between the control and infected groups, possibly because of the non-specificity of this enzyme as an indicator of hepatic lesion. The results show that infection caused by P. juxtanucleare in G. gallus provokes hepatic alterations, indicated by the increase in the ALT enzyme activity and by the inflammatory infiltrates found in the liver sections of the infected fowls. © 2010 Elsevier B.V. Open access under the Elsevier OA license. 1. Introduction Besides domestic chickens, there are reports of other fowls being infected by P. juxtanucleare: Gallus lafayet- Plasmodium juxtanucleare Versiani & Gomes, 1941, the tei Linnaeus, 1758 (jungle fowl) in Sri Lanka (Dissanaike, agent that causes avian malaria in Gallus gallus Linnaeus, 1963); Bambusicola thoracica sonorivox Linnaeus, 1758 1758, was observed for the first time in Brazil when (bamboo partridge) in Taiwan (Manwell, 1966); Fran- researchers studying avian spirochetosis at a street mar- colinus spp. (partridge) in Africa (Mohan and Manwell, ket, while examining blood samples drawn from live fowls, 1966); Crysolophus pictus Linnaeus, 1758 (golden pheas- observed a small parasite with rounded or irregular shape, ant), Lophura nyctemera Linnaeus, 1758 (silver pheasant), always found near the blood cell nuclei. For this reason, it Crysolophus amherstiae Linnaeus, 1758 (Lady Amherst’s was named P. juxtanucleare (Versiani and Gomes, 1941). pheasant) and Phasianus colchicus Linnaeus, 1758 (com- mon pheasant) in Brazil (Massard and Massard, 1981); and Meleagris galopavo Linnaeus, 1758 (Turkey), also in ∗ Corresponding author. Tel.: +55 21 26823222; fax: +55 21 26821763. Brazil (Serra-Freire and Massard, 1979). The invertebrate E-mail address: [email protected] (J. Pinheiro). hosts of P. juxtanucleare are mosquitoes of the Culicini 0304-4017 © 2010 Elsevier B.V. Open access under the Elsevier OA license. doi:10.1016/j.vetpar.2010.10.031 208 U. Vashist et al. / Veterinary Parasitology 175 (2011) 207–211 tribe (Versiani and Gomes, 1941; Bennet et al., 1966; were fed Purina Natural® free of antibiotics and coccid- Garnham, 1966; Krettli, 1972; Lourenc¸ o-de-Oliveira and iostats and given water ad libitum. At 60 days-old, the Castro, 1991). birds were separated into two groups of 12 animals each, The pathogenicity of P. juxtanucleare can be related to a control group and a group infected by P. juxtanucleare. the strains in which it produces exoerythrocytes stages, The control group was intracelomically inoculated with which can mainly be found in the spleen, along with 0.5 ml of physiological saline, while the infected group, the liver, lungs, bone marrow and brain (Paraense, 1947; by the same route, received 0.5 ml of blood from a donor Krettli, 1972). The number of parasited erythrocytes is gen- bird infected by P. juxtanucleare, with a parasite load of erally low in naturally infected fowls. For this reason, the around 7%. The parasite load of the birds in the two increase in blood parasite loads is usually slow due to the groups was then monitored by examining blood smears small number of merozoites produced by the schizonts in from a drop of blood drawn from the fine wing capil- relation to the plasmodia that afflict humans (Massard and laries. The blood smears were examined daily during the Massard, 1981). first 15 days after inoculation, the most critical period for Poultry breeding in recent decades has been growing experimental infection caused by P. juxtanucleare accord- in importance in the world, generating large capital move- ing to the literature (Vashist et al., 2008, 2009). Afterward, ments and increasing the number of rural jobs (Mota et al., they were examined every three days until the 42nd day 1998). Besides this, ecological problems related to cat- post-inoculation, the period in which according to the lit- tle raising, such as soil compactation and release of large erature the parasitemia tends to become chronic (Silveira amounts of methane by cattle, lead many experts to predict et al., 2009; Vashist et al., 2008, 2009). The smears were that the poultry industry will gain even more importance taken to the laboratory, fixed in methanol for 3 min and in the coming years. In addition, birds have shorter breed- stained with Giemsa stain, diluted in distilled water (1:4) ing cycles, require less space, a reduced amount of money for 45 min. A hundred fields per slide were observed using directed to its growth and they are an excellent source of a light microscope at 1000×. The total number of evo- protein for humans. But because industrial-scale breeding lutive forms of P. juxtanucleare found in each smear was operations make it easier for disease to spread among the recorded. Each week about 1 ml of blood was drawn from animals, there is a need for better methods to diagnose each fowl for hematocrit determination, by the microhe- avian diseases and to study the ways these diseases can matocrit technique, and for analysis of the activity of the affect the birds’ productivity. aspartate aminotransferase (AST) and alanine aminotrans- Biochemical variables have been used to diagnose dis- ferase (ALT) enzymes. For the aminotransferase analyses, eases in pets and producing animals (Borsa et al., 2006). 0.5 ml of ALT or AST substrate (a solution of 0.2 M l-alanine In the current literature there are no studies on the hep- or 0.2 M l-aspartate, respectively, 0.002 M ␣-ketoglutarate atic profile of birds infected by P. juxtanucleare, although and 0.1 M sodium phosphate buffer, pH 7.4) was incu- this is an important parameter that can be used to evaluate bated at 37 ◦C for 2 min. Then, 100 lor200l of serum poultry health. (for ALT or AST, respectively) was added and the solution Aminotransferases (ALT and AST) are a group of was homogenized and incubated again at 37 ◦C for 30 min. enzymes that catalyze the interconversion of amino acids After that, 0.5 ml of 0.001 M 2.4-dinitrophenylhydrazine into ␣-ketoacids by transfer of amine groups (Moss and was added and maintained at 25 ◦C for 20 min. The reaction Henderson, 1998). Aminotransferases play an important was finalized by adding 5 ml of 0.4 M NaOH. The read- role in the link between the amino acids and carbohy- ings were taken in a spectrophotometer at 505 nm and drates metabolism. They are an essential group of enzymes the results were expressed as URF/ml (Kaplan and Pesce, for gluconeogenesis, besides being excellent indicators of 2003). hepatic lesions (Pinheiro et al., 2001). To study the possible types of hepatic lesions result- This article reports an experiment to verify changes ing from the infection by P. juxtanucleare, liver fragments in the hepatic profile of G. gallus in response to infection were taken from two fowls from each group at the end caused by P. juxtanucleare. of the 45-day experiment. For this, liver fragments of about 1 cm in diameter were taken during necropsy from 2. Materials and methods the left liver lobe of each fowl. The fragments were fixed in 4% formalin for 24 h at 4 ◦C and then main- This experiment was performed on 24 hens of the Cobb tained in 70% ethanol. These tissues were processed breed, purchased as day-old chicks. The chicks were vac- according to routine histological techniques, embedded cinated against fowl pox, gumboro disease and Marek’s in paraffin, sliced into 5-m sections with a microtome disease. The chicks were taken from the commercial estab- and mounted on glass slides. The sections were stained lishment and transferred to the W.O.