Abel Gonzalez OlivaAbel Gonzalez Oliva oliva@itqb.unl.ptoliva@itqb.unl.pt

Abel Gonzalez OlivaAbel Gonzalez Oliva oliva@itqb.unl.ptoliva@itqb.unl.pt

Biosensor / Biomolecular Biosensor / Biomolecular Diagnostics Laboratory Diagnostics Laboratory Biosensor / Biomolecular Biosensor / Biomolecular Diagnostics Laboratory Diagnostics Laboratory

Project Acronyme: ADDAV Contract No: ICA4-CT2000-Project Acronyme: ADDAV Contract No: ICA4-CT2000-300028 300028 INCO 2 PROGRAMMEINCO 2 PROGRAMME - EU- EU

Molecular and immunological characterization of Molecular and immunological characterization of merozoite antigens and their encoding genes of merozoite antigens and their encoding genes of a Theileria species highly pathogenic for small a Theileria species highly pathogenic for small

ruminants in China: application for the development ruminants in China: application for the development of diagnostics and vaccineof diagnostics and vaccine

2001 - 20052001 - 2005

1.1. Prof. J.S. Ahmed (Co-ordinator), Research Center Bortstel,Prof. J.S. Ahmed (Co-ordinator), Research Center Bortstel, Borstel, Germany.Borstel, Germany.

2. Prof. Hong Yin, Lanzhou Veterinary Research Institute 2. Prof. Hong Yin, Lanzhou Veterinary Research Institute (LIVRI), Chinese Academy of Agricultural Sciences, (LIVRI), Chinese Academy of Agricultural Sciences, Xujiaping 11, Yanchangbu, Lanzhou, Gansu, 730046, PR of Xujiaping 11, Yanchangbu, Lanzhou, Gansu, 730046, PR of China.China.

3. Prof. J. Wu Animal Quarantine Institule, Ministry of 3. Prof. J. Wu Animal Quarantine Institule, Ministry of Agriculture (AQIMA), 369 Nanjing Road, Qingdao 266032, Agriculture (AQIMA), 369 Nanjing Road, Qingdao 266032, PR of China.PR of China.

4. Prof. Z. Yuan, Institute of Animal Husbandry of Gannan 4. Prof. Z. Yuan, Institute of Animal Husbandry of Gannan Tibet Nationality Autonomous Prefecture, Gansu Province, Tibet Nationality Autonomous Prefecture, Gansu Province, PR China.PR China.

5. Prof. Abel G. Oliva, Instituto de Biologia Experimental e 5. Prof. Abel G. Oliva, Instituto de Biologia Experimental e Technologica (IBET) Biosensor/Biomolecular Diagnostics Technologica (IBET) Biosensor/Biomolecular Diagnostics Laboratory - Apartado 12 , 2781-901 Oeiras, Portugal.Laboratory - Apartado 12 , 2781-901 Oeiras, Portugal.

Parceiros no ADDAVParceiros no ADDAV

Objectivos do ADDAVObjectivos do ADDAV

Os principais objectivos de este Os principais objectivos de este projecto de investigação foram:projecto de investigação foram:

Desenvolvimento de métodos de Desenvolvimento de métodos de diagnóstico e vacinas, para diagnóstico e vacinas, para espécies altamente patogénicas espécies altamente patogénicas de de Theileria sp.Theileria sp. que afectam que afectam pequenospequenos ruminantesruminantes na China. na China.

Merogony /Merogony /

Geographic distribution of Theileria sp. (China) in the northwest part of the People’s Republic of China. Endemic area surrounded

in black.

Zona endémica deZona endémica de Theileria sp. Theileria sp. na RP China na RP China

Tarefas do IBET no ADDAVTarefas do IBET no ADDAVDeveloped tasks:

•Participation in the infection process and in serum/blood collection•Infected blood characterisation by PCR and Reverse line blot (RLB) technique. •Merozoites isolation and purification studies as well as identification of potential antigens.•Antigens characterisation by mass spectrometry •Establishment of In vitro cultivation of Theileria sp. (China) merozoites in erythrocytes

Treino de pessoalTreino de pessoal

• Coolaboração na preparação do “Workshop in Biochemical & Molecular Biology Techniques” para estudantes Chineses. Lanzhou - 12-26 June 2002

• Joana Miranda (PhD Student) tem realizado numerosas visitas ao Lanzhou Veterinary Research Institute – Lanzhou, República Popular da China, onde desenvolveu trabalho experimental relativo a parasita Theileria sp., sob a supervisão do Prof. Yin Hong), nomeadamente para recolha de amostras de soro e sangue ovino, isolamento de merozoites, purificação de RNA e produção de cDNA, e desenvolvimento e validação de teste de ELISA Junho 2002 e Abril-Maio 2005

WorkshopWorkshop

Workshop “Establishment of tick colonies and tissue cultures of Theileria annulata for the immunisation against Tropical Theilerosis”. CMDT & IBET, Lisbon, Portugal, from the 31st March until the 10th of April, 2003.

Establishment of continuous in vitro culture of erythrocytic stages of a Establishment of continuous in vitro culture of erythrocytic stages of a Theileria Theileria species highly pathogenic for small ruminants in Chinaspecies highly pathogenic for small ruminants in China.. Miranda J., Nascimento E., Cruz H., Zweigarth E., Oliva A. (2006) Vet Parasit. (submitted)Identification of homologous genes of Identification of homologous genes of T. annulata T. annulata proteins in the genome of proteins in the genome of Theileria spTheileria sp. (China).. (China). Miranda J., Bakheit M., Schneider I.., Haller D., Ahmed J., Hong Y., Oliva A., Seitzer U. Ann NYAcad Sciences (submitted) Development of a recombinant indirect ELISA for the diagnosis of Development of a recombinant indirect ELISA for the diagnosis of TheileriaTheileria sp. (China) infection in small ruminantssp. (China) infection in small ruminants.. Miranda J, Bakheit MA, Liu Z, Yin H, Yongjuan M, Guo S, Beyer D, Oliva A, Ahmed JS, Seitzer U (2006) - Parasitol Res (in press)Identification and characterization of merozoite antigens of a Theileria Identification and characterization of merozoite antigens of a Theileria species highly pathogenic for small ruminants in Chinaspecies highly pathogenic for small ruminants in China. Miranda J, Nascimento E, Cruz H, Hong Y, A. Coelho, Ahmed J, Oliva A (2006) Journal of Veterinary Pathology Ann NYAcad Sci (in press).Identification of antigenic proteins of a Theileria species pathogenic for Identification of antigenic proteins of a Theileria species pathogenic for small ruminants in China: characterization of the immune response and small ruminants in China: characterization of the immune response and assessment of diagnostic potentialassessment of diagnostic potential J.Miranda , B.Stumme, D. Beyer, H. Cruz, A.G. Oliva, D. Wicklein, H. Yin, L. Jianxun, J.S. Ahmed, U. Seitzer. (2004) Ann NYAcad Sci, 1026:1-4 Immunosensors for diagnostic applicationsImmunosensors for diagnostic applications. Helder J. Cruz, Carla C. Rosa, Abel G. Oliva (2002) Parasitol Res 88: S4-S.

Artigos científicosArtigos científicos

Presentações em Conferências Int.Presentações em Conferências Int.

Establishment of Continuous Establishment of Continuous in vitroin vitro Culture of Erythrocytic Stages of a Culture of Erythrocytic Stages of a Theileria Theileria species Highly Pathogenic for Small Ruminants in Chinaspecies Highly Pathogenic for Small Ruminants in China .. JMiranda, ENascimento, HCruz, YHong, EZweygarth, AOliva June 26 -July 1st 2005, Hanoi/Vietnam.Identification and characterization of merozoite antigens of Identification and characterization of merozoite antigens of TheileriaTheileria species 2 highly pathogenic for small ruminants in China.species 2 highly pathogenic for small ruminants in China. Miranda, J.; Nascimento, E.; Cruz, H.; HYin.; Ahmed, J.; Oliva A. June 26 -July 1st 2005, Hanoi/Vietnam.Identification of homologous genes of Identification of homologous genes of T. annulata T. annulata proteins in the proteins in the genome of genome of Theileria spTheileria sp. (China).. (China). JMiranda, MBakheit, ISchneider, DHaller, JAhmed, HYin, Abel G. Oliva, Ulrike Seitzer June 26 -July 1st 2005, Hanoi/Vietnam.Identification of antigenic proteins of a Theileria species pathogenic Identification of antigenic proteins of a Theileria species pathogenic for small ruminants in China: characterisation of the immune response for small ruminants in China: characterisation of the immune response and assessment of diagnosis potentialand assessment of diagnosis potential JMiranda, BStumme, DBeyer, HCruz, AOliva, DWicklein, HYin, LJianxun, JAhmed, USeitzer (2003) 7th Biennal Conference STVM (Society for Tropical Veterniary Medicine) 22nd – 28th June 2003 Foz do Iguaçu, Brasil.

Identification and characterization of different Theileria china antigen/sera Identification and characterization of different Theileria china antigen/sera preparations from small ruminantspreparations from small ruminants Helder J.S. Cruz, Joana Miranda, Abel G. Oliva (2002) Central Asia and European Union Commission International Conference on Ticks and tick-borne Diseases, Tashkent, Republic of Uzbekistan, 13-15 Mai. 2002.

Antigen characterization and purification of Theileria china infected sera Antigen characterization and purification of Theileria china infected sera from small ruminantsfrom small ruminants Helder J.S. Cruz, Joana Miranda, Luo JX, Yin Hong and Abel G. Oliva (2001) INCO-DEV Meeting: Hypodermosis and Theileria diseases International Conference, Lanzhou, Republic of China, 16th -19th Oct. 2001.

Immunosensors for diagnosticImmunosensors for diagnostic Oliva, A. 2nd.Internation Conference on Ticks and Tick-borne diseases (ICTTD). Shanghai, China, September 11th- 15th, 2000.

Presentações em Conferências Int.Presentações em Conferências Int.

Identification and characterization of merozoite antigens of Identification and characterization of merozoite antigens of Theileria Theileria species highly pathogenic for small ruminants in China.species highly pathogenic for small ruminants in China. (oral presentation) J. Miranda, E. Nascimento, H. Cruz, H. Yin, A. Oliva. EMOP XI, Valencia, Spain, 2004

Lanzhou Veterinary Research Institute (LIVRI)Lanzhou Veterinary Research Institute (LIVRI)