Microbes, as the most abundant sediment organisms, play a major role in the fate of pollutants. Many pollutant-related microbes have been studied, yet the entire microbial community profiles in subtropical marine environments have not been reported. The focus of this thesis study was on the impacts of a large sewage pollution reduction program, Harbor Area Treatment Scheme (HATS) implemented in Victoria Harbor, on sediment microbes and the main goal of this thesis research was to investigate the diversity, community structure, and the spatial-temporal variability of prokaryotes (bacteria and archaea in general, sulfate-reducing bacteria in particular) in sediments of Victoria Harbor, Hong Kong. Various molecular methods, including clone library construction, terminal restriction fragmen...[ Read more ]

Microbes, as the most abundant sediment organisms, play a major role in the fate of pollutants. Many pollutant-related microbes have been studied, yet the entire microbial community profiles in subtropical marine environments have not been reported. The focus of this thesis study was on the impacts of a large sewage pollution reduction program, Harbor Area Treatment Scheme (HATS) implemented in Victoria Harbor, on sediment microbes and the main goal of this thesis research was to investigate the diversity, community structure, and the spatial-temporal variability of prokaryotes (bacteria and archaea in general, sulfate-reducing bacteria in particular) in sediments of Victoria Harbor, Hong Kong. Various molecular methods, including clone library construction, terminal restriction fragment length polymorphism (T-RFLP), archaeal phylum-scale primed PCR (ASP-PCR), genus specific PCR (GS-PCR), and web-base Phylogenetic Assignment Tools (PAT) analysis were used to accomplish this goal. Sediment samples were collected bimonthly from three polluted (VHE, VH, and VHW) and two adjacent (open oceanic, TLC; estuary discharge affected, PC) sites from August 2004 to October 2006.

Bacterial communities in samples from different sites showed high 16S rDNA richness and were significantly different from each other. Phylogenetic analysis of full-length 16S rDNA revealed 19 bacterial phyla in Victoria Harbor sediments. γ- and δ-proteobacteria were dominant at all sites (33.33-11.67%). In addition, γ- and δ-proteobacteria, holophaga/acidobacteria, and planctomycetales were recorded in all the libraries. Bacterial abundance in the surface sediments from all the sites, except the PC site, showed a clear seasonal change during the three-year monitoring period. The changes of bacterial community composition in surface sediments collected from five sites were investigated from 2004 to 2006. Irrespective of the sampling time (except in winter 2005), the chronically contaminated site, VH, showed the largest deviation in bacterial community structure from the adjacent four sites. Surprisingly, bacterial communities in the sediments from two contrasting environments, TLC (oceanic) vs. PC (river runoff affected), shared high similarity throughout the study period. Results further showed that the disposal of treated sewage into the western side of Victoria Harbor (VHW) since 2001 had an insignificant effect on bacterial community composition. A striking seasonal trend was observed at the central harbor area (i.e. summer samples formed a separate group) but not at PC, where there was a strong seasonal gradient in salinity and water quality. Although factors other than pollution gradients may explain the observed bacterial community patterns, the information presented here may be useful in predicting long-term effects of anthropogenic pollution on benthic bacterial community.

Substantial variations were detected in both the presence and the type of archaeal phyla (including Crenarchaeota, Euryarchaeota, Korarchaeota, and Nanoarchaeota) at different sediment depths (0-54 cm) at both VH and PC using archaea phylum scaled PCR. The sequences of 155 unique phylotypes were affiliated with the phylum Crenarchaeota and Euryarchaeota. The archaeal diversity at VH and PC displayed high phylotype richness (193 to 698) and evenness (0.8766 to 0.9141). The archaeal community compositions from the two sites were significantly different: 88.4% of the phylotypes at VH were detected as Crenarchaeota members while 53.3% of the phylotypes at PC were identified as Euryarchaeota members. Phylotypes of the two sites belonged to separate clusters within the phyla Crenarchaeota or Euryarchaeota. Judging by genetic distance, phylotypes were more diverse at PC than VH.

The most diversified sulfate-reducing bacteria (SRB) were found at polluted sites (VH and VHW). Although the compositions of the SRB communities were different at the two polluted sites, they were still more similar to each other than to the two reference sites (TLC and PC). Based on a dsrAB clone library constructed at VHW, five clades of SRB were found and three clades belonged to the known families Desulfobacteraceae, Desulfobulbaceae, and Syntrophobacteriaceae. The majority of sequenced phylotypes constituted the remaining two unclassified groups, suggesting unique and novel SRB members related to the polluted harbor environment. In addition, SRB richness was correlated with sulfur content, acid volatile sulfate, and redox potential. The spatial-temporal changes of SRB in surface sediments were monitored in nine sampling occasions using genus-specific PCR. Desulfosarcina, Desulfobacter, Desulforhopalus, Desulfobulbus, and Desulfovibro decreased in quantity gradually during this period. Integrated density values of SRB were significantly higher at the polluted site than at adjacent nutrient-rich site, irrespective of sampling time. Sampling time and site interacted strongly and both affected SRB relative quantities. Negative correlations between sampling time and SRB integrated density value reflected the adverse impacts of decreasing pollutant, derived from HATS, on sediment SRB communities.

This thesis study disclosed a portion of the clones that were distantly related to sequences in the GenBank, suggesting microbes in Victoria Harbor sediments were unique and diversified. The results of this study indicated that the microbial communities in Victoria Harbor sediments were highly dynamic and reflected contrasting marine environmental conditions. The results from this study also implied that the contrasting marine environments, even within a small geographic area in the subtropical region, could harbor highly diversified and distinct microbial communities. Unfortunately, no significant temporal changes in the microbial communities were observed over the monitoring period in response to the abatement of direct pollutant discharges and government management, indicating three-year period may not be enough to monitor the impacts of sewage pollution reduction program on sediment microbes during environmental recovery processes. However, this thesis research did prove that the disposal of treated sewage into the western harbor since 2001 did not significantly affect the bacterial community composition in the sediments, at least within 5 years (2001-2006).