| Summary: | 碩士 === 國立中山大學 === 海洋資源學系研究所 === 89 === Abstract
Recent researches have pointed out that most of marine bacteria are uncultivable. However, majority of prior researches about bacteria flora in cultivated ponds used cultivating method to researches. That means those researches ignored uncultivable bacteria in ponds and caused inaccuracy in counting bacteria. Therefore we used analyzing bacteria 16S rDNA sequences to study composition of bacteria in cultivated ponds in place of traditional bacteria taxonomy. The phylogenetic diversity of bacteria examined by analyzing the 16S rDNA sequences permits the characterization of environmental bacteria community without culturing and has been used widely.
This research is to adopt both MMA medium cultivation and direct recovery of bacteria 16S rDNA sequences to investigate bacteria flora in a closed Penaeus monodon pond. We sampled from A2 (10×8×1.5m) test pond at the Department of Marine Resource in Sun Yat-Sen University on August 17, 1999. Then, we adopted AO (Acridine Orange) epifluorescent microscopic technique to count total direct count (TDC) and direct count of viable bacteria cell (DVC). Respective results were 2.846×107/ml, 1.029×107/ml, and plate count (PC) determined by MPN count method were 1.130×105cfu/ml.
In the part of cultivable bacteria, they could be separated into 9 groups by their morphologic after culturing in MMA plate at 25℃for 5 days. We isolated 15 strains to analyze their 16S rDNA sequences, and separated respectively into 4 groups after comparing with the genebank. Those four groups are CFB group, low G+C Gram-positive bacteria, Alpha proteobacteria and Gamma proteobacteria. The genus Vibrio (47%) in the Gamma proteobacteria group is the dominant. In the part of uncultivable bacteria, we filtered bacteria from the water in the same pond, amplified the 16S rDNA by the polymerase chain reaction (PCR) and then cloned. After that, we randomly isolated 40 clones for sequence analysis. The bacteria belong to following groups, cyanobacteria, CFB group, Verrucomicrobia, Gram-positive eubacteria, Alpha proteobacterium, Beta proteobacterium, Gamma proteobacterium and Delta proteobacterium.
|
|---|
