Typing of Aeromonas isolates from children with diarrhoea & water samples by randomly amplified polymorphic DNA polymerase chain reaction & whole cell protein fingerprinting.

BACKGROUND & OBJECTIVES: Aeromonas spp. are water-borne organisms, often associated with childhood diarrhoea. The present study was conducted to examine the epidemiological relationship among the Aeromonas spp. isolated from water and children with acute diarrhoea in Chennai. METHODS: Thirty six Aeromonas isolates inclusive of 16 from children with diarrhoea, 15 from domestic water samples and 5 reference strains were studied by randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR). Twenty eight Aeromonas isolates, 15 from children with diarrhoea, 10 from domestic water samples and three reference strains were analysed by SDS-PAGE for their whole cell protein profiles. RESULTS: The 36 Aeromonas isolates examined by RAPD-PCR generated RAPD fingerprints with majority of the bands ranging from about 250 to 2800 bp. The RAPD fingerprints did not correspond with the phenospecies and varied greatly among the strains within the phenospecies. Cluster analysis revealed two major groups at 75 per cent hierarchical level, comprising 18 Aeromonas isolates, mainly recovered from domestic water samples, while the clinical isolates were scattered in different hierarchical levels in the dendrogram. The whole cell protein fingerprints examined by SDS-PAGE did not correspond with the phenospecies. Only four isolates of A. caviae were found to produce similar protein fingerprints allowing them to form a cluster at about 90 per cent hierarchical level, while the rest of the isolates were scattered at various hierarchical levels in the dendrogram. INTERPRETATION & CONCLUSIONS: In the present study, RAPD fingerprinting was found to be useful in distinguishing Aeromonas isolates recovered from clinical and domestic water supplies. However, RAPD-PCR could not distinguish the phenospecies of the genus Aeromonas. Whole cell protein fingerprinting and cluster analysis could neither differentiate isolates from clinical and domestic water sources nor the phenospecies of the genus Aeromonas.

Development of a computer software for analysis of SDS-PAGE protein fingerprints of bacterial isolates.

Protein fingerprinting is a widely used technique in epidemiological studies for typing bacterial strains. This study reports the development of a computer based gel analysis system. The system has the capability to analyse SDS-PAGE whole-cell protein profiles using digital image processing techniques. The software incorporates spatial and frequency domain operators for image enhancement, support for geometric correction of images and new algorithms for identification of strain tracks and protein bands. The system also provides facilities for correcting imaging defects for inter-gel comparison, similarity analysis, clustering and pictorial representation of results as a dendrogram. The software is highly interactive, user-friendly and can produce accurate results for differentiation of bacterial strains with minimal overhead of time.