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Executive summary

Project Background

Haemorrhagic septicaemia is one of the important diseases of cattle and buffaloes not only in Malaysia but also in many parts of the world particularly Asia. It is caused by Pasteurella multocida B:6 leading to disease outbreaks. In Malaysia alone, outbreaks of this disease led to an estimated annual loss of RM2.4 million (Joseph, 1989). Thus, understandably, most initial research activities on haemorrhagic septicaemia in Malaysia were focussed on the control of haemorrhagic septicaemia through vaccine development, which was carried out since 1960s (Thomas, 1968). Eventually, an effective oil-adjuvant vaccine against haemorrhagic septicaemia was produced locally in 1978 (Chandrasekaran & Yeap, 1978). Although the vaccine was found to be effective in preventing haemorrhagic septicaemia for 9-12 months, the disease could not be effectively contained. This is evidenced when approximately 42,000 deaths were recorded from outbreaks of this disease between 1977-1987, leading to a loss of approximately RM38 million (Joseph, 1989). The main reason for these outbreaks was the low vaccination coverage. Only 26% (range between 12-60%) of cattle and buffaloes were vaccinated (Saharee & Salim, 1989). This is due to difficulties in vaccine administration particularly on the animals that are kept under extensive system, while in fact, approximately 70% of the 950,000 heads of cattle and buffaloes in Malaysia are kept under this system (Saharee & Fatimah, 1993). With this difficult scenario, a different approach should be studied and adapted to effectively increase the vaccination coverage of cattle against haemorrhagic septicaemia.

Scope: This programme focuses mainly on an alternative prevention of haemorrhagic septicaemia. Since lipopolysaccharide (LPS) and the outer membrane proteins (OMP) have been shown to provide inconsistent protection to the host, detailed study on other proteins such as the fimbria, which enhance adherence of the organism to the host cells, may be beneficial. Currently, the gene that encodes the fimbrial protein is not known although the organism is extremely important in Malaysian cattle. At the same time, there are not many vector and delivery systems for vaccine that are available for ruminants in Malaysia. Ideally the system should be cost-effective and applicable to large ruminants including animals maintained under extensive, semi-wild environment. Pseudomonas aeroginosa. Staphylococcus aureus, Lactobacillus sp. and non-pathogenic Pasteurella multocida are among the suitable candidate vectors for further study.

Since the currently available haemorrhagic septicaemia vaccine is less favourable due to the animal restraining problems, an alternative method of vaccine delivery such as spray or oral vaccination should also be studied. A Pasteurella multocida DNA vaccine, when available will be introduced intranasally or orally before the protection against challenge by field strain of Pasteurella multocida is determined.

Objectives:

1. To identify and characterise genes encoding immunogenic proteins
2. To study the suitability of Pseudomonas aeruginosa or other suitable bacteria as vaccine delivery system for vaccination against haemorrhagic septicaemia
3. To study the pathogenesis of haemorrhagic septicaemia in laboratory animal model and in cattle, particularly on the role of fimbria in disease development.
4. To study the development of mucosal immunity of cattle following intranasal or oral introduction of Pasteurella multocida antigen.

Methodology:

Objective 1: The gene(s) that encodes the fimbrial proteins and other proteins that play important role in virulence and immunity will be amplified by polymerase chain reaction (PCR), cloned and characterized by sequence comparison and phylogenetic analysis.

Objective 2: Suitable bacterial vector(s) will be tested as vector for the selected gene. This study will focussed particularly on the detailed stability and suitability of Pseudomonas aeruginosa, Staphylococcus aureus, Lactobacillus and Pasteurella multocida as vector since this bacteria is recently being found to be vector for certain foreign gene.

Objective 3: Animal models such as rabbits or mice will be infected by Pasteurella multocida and the lesions and disease development will be studied. Similarly, the role of fimbria of Pasteurella multocida in the disease development, particularly in the attachment ability of Pasteurella multocida will be studied using both electron microscopy and antiserum techniques. This study will be re-applied in cattle.

Objective 4: Suitable antigens (whole cell or gene) recognised from earlier studies will be introduced either orally or intranasally before the development of mucosal immunity both in the respiratory and gastro-intestinal systems are studied. These include the level of secretory IgA, number and size of associated lymphoid tissues and the inter-relationship of lymphocytes between the two systems. Challenge trials will be conducted to determine protective level and mechanisms.

Research Projects:

Project 1: "Pathogenesis of haemorrhagic septicaemia and the role of fimbria of Pasteurella multocida in disease development - Mohd Zamri Saad, Shahirudin Shamsudin, Jasni Sabri, Muthafar Haddawi (UPM, VRI, KUSTEM)

Project 2: "Identification of gene(s) associated with immunity, particularly the fimbria of Pasteurella multocida and a suitable vector(s) for vaccine delivery using Pseudomonas aeruginosa as model -Siti Khairani Bejo, Zunita Zakaria, Sharifah Syed Hassan, Abd. Rahman Omar (UPM & VRI)"

Project 3: "Epidemiological and pathological pattern of haemorrhagic septicaemia in Malaysia - S. Chandrasekaran, Abd. Aziz Saharee, Latiffah Hassan (VRI & UPM)"

Project 4: "Study on the development of respiratory and gastro-intestinal mucosal immunity following administration of Pasteurella multocida antigens either orally or intranasally - Mohd Effendy Abd. Wahid, Muthafar Haddawi, Mohd Zamri Saad (KUSTEM & UPM)"

Research Status: New Programme

Expected Outcome:

1. DNA vaccine against haemorrhagic septicaemia
2. Alternative vaccine and vaccination in cattle and buffaloes

Period of the project : 3 Years (2003 - 2005)

Project Status

- On-going