Ghaderpour, Aziz (2013) Evaluation of microbiological quality and safety in Kuala Sepetang estuaries (Perak, Malaysia) using cultivation-dependent method / Aziz Ghaderpour. Masters thesis, University of Malaya.
Abstract
Estuaries are crucial coastal and marine habitats of various aquaculture organisms. However, uncontrolled development has caused contamination in many coastal areas all over the world that will eventually bring negative effects to aquaculture industry and public health. Bacteriological study based on culturing methods is appropriate not only for routine monitoring in laboratories, but also for making inferences on the physiological and metabolic properties of the organisms. Therefore the applicability of CHROMagarTM Orientation agar was evaluated in microbiological survey of estuarine waters and sediments as well as to detect and isolate human-associated bacterial pathogens in the surface waters and sediments along Kuala Sepetang estuary. Genomic diversity of the isolated E. coli strains based on CHROMagarTM Orientation agar was determined from Kuala Sepetang estuary. Water and sediment samples were collected from Kuala Sepetang estuary. The total culturable bacteria (TCB) was determined by direct-plating on both nutrient agar and CHROMagarTM Orientation. Nutrient agar, in all cases, yielded a higher TCB count than CHROMagarTM Orientation. The cultivable bacteria on CHROMagarTM Orientation could be characterized into four major color morphotypes: purple, green, blue and colorless. The green morphotype was found to be more dominance in the sediment of upper estuary while the colorless morphotype dominated the bottom sediment of lower estuary. Identification of representative isolates of green morphotype (n=107) using Biolog gen III and biochemical assays showed that green morphotype comprised of Enterococcus sp., Exiguobacterium sp., Paenibacillus sp., Stenotrophomonas sp. and Staphylococcus scirius. The colorless morphotype comprised of Pseudomonas sp., and Acinetobacter sp. Direct-plating on CHROMagarTM Orientation yielded low recovery of Enterobacteriaceae, specifically E. coli, Klebsiella and Enterobacter, which could be due to cell stress and injury while these enteric iv bacteria were exposed to the unfavourable environment. Detection and isolation of these bacteria were achieved by using direct plating method or by incorporating pre-enrichment before plating onto CHROMagarTM Orientation. A wide range of could identify, specifically disease-associated bacteria, such as E. coli, Pseudomonas sp., Serratia sp., Enterococcus sp., Staphylococcus sp., Klebsiella sp. and Enterobacter sp. REP-PCR genotyping of E. coli isolates (n=53) revealed high genetic diversity along the estuaries. The REP-PCR profiles were grouped into clusters: I, II, III, IV, V, VI, VII, VIII and IX. Cluster III was major cluster consisting of 41% (n=22/53) of total isolates. Generally, since water is streaming from upstream to downstream of estuary and due to geographical location of the water which is in interconnected in Kuala Sepetang estuary most of the isolates clustered together, even from different Rivers (Kuala Selinsing and Kuala Sangga Besar River). In conclusion, CHROMagarTM Orientation was found to be a useful medium in the estuary study as it allows simultaneous isolation and identification of waterborne bacterial pathogens within two to three days. Kuala Sepetang estuary, as important area for one of the major estuary supporting numerous aquacultural activities, was found to be heavily contaminated with various types of waterborne bacterial pathogens.
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