Chadwick, N.C.; (1998) Molecular strategies for the detection of measles virus in inflammatory bowel disease. Doctoral thesis , University of London.

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Abstract

Hypotheses. i) Atypical exposure to measles virus is a factor in the aetiology of inflammatory bowel disease (IBD). ii) Measles, mumps and rubella (MMR) vaccination is a factor in the aetiology of autistic enteropathy. Aims. i) To compare a range of molecular techniques for measles RNA amplification. ii) To develop a sensitive and robust method for the detection of measles RNA. iii) To analyse clinical samples from IBD patients for the presence of measles RNA. iv) To analyse clinical samples from autistic enteropathy patients for the presence of measles, mumps and rubella RNA. Methods development. Three RNA amplification methods were compared in terms of their sensitivity and fidelity for the detection of measles RNA and nucleic acid sequence-based amplification (NASBA) was found to be the most sensitive. In a preliminary study, NASBA did not detect any measles RNA in a coded series of IBD and control intestinal tissues. In order to improve the detection sensitivity, the use of hybrid capture, using measles-specific oligonucleotides linked to paramagnetic solid phase supports, was investigated. Hybrid capture was found to increase the measles RNA detection sensitivity 100-fold when followed by RT-PCR. An internal modified transcript was developed which could be co-amplified with measles RNA as an internal positive control. IBD samples. Resection samples from 20 IBD and control patients were used for measles hybrid capture followed by RT-PCR, in addition to peripheral blood mononuclear cells (PBMCs) from 13 IBD and control patients. Autistic enteropathy samples. Biopsies, PBMCs and Vero/PBMC cocultures were analysed from 22 patients with autistic enteropathy and 6 controls. Results. Hybrid capture and RT-PCR could detect 10⁴ molecules of a measles RNA transcript added to control tissue homogenates. The fidelity of NASBA, in terms of its nucleic acid error rate, was found to be comparable with that of RTPCR. All samples were found to be positive for a housekeeping RNA species and internal modified positive control RNA. None of the samples tested positive for measles, mumps or rubella RNA, although viral RNA was successfully amplified in positive control samples. Conclusion. The results do not support previous data implicating persistent measles virus infection with the aetiology of IBD or autistic enteropathy.

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