Project description:To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild.

Project description:The federally endangered Okaloosa darter (Etheostoma okaloosae) is found almost exclusively on the Eglin Air Force Base in the Choctawhatchee Bay watershed of Florida. Portions of this limited habitat are threatened with erosion of soils, altered hydrology, and impaired water quality. One stream reach in particular, East Turkey Creek, has demonstrated potential water quality problems including poor invertebrate bioassessment scores (IBI), uncharacteristically high conductivity values, and low numbers of Okaloosa darters. General water quality (dissolved oxygen, specific conductance, pH, temperature, and relative turbidity and primary productivity) was characterized in both the potentially impacted East Turkey Creek and a reference stream (Long Creek). Water quality was assessed during a 30 day exposure using passive samplers for both non-polar and polar effluent parameters. Metal loading in the system was assessed via fish tissue burdens in resident Pteronotropis hypseleotris. Additionally, microarray analysis was performed on gonad and liver tissue from fathead minnows, Pimephales promelas, after 48-h exposures to water collected from the two creeks and brought into the laboratory. Gene expression changes were evident at the site below the influence of a wastewater spray field sited along East Turkey Creek, suggesting that anthropogenic compounds in the effluent waters may have affected both liver and testis function and could be related to account the general decrease in populations of the Okaloosa darter. Array hybridizations were performed using a single color design. Four biological replicates consisting of four different individuals were analyzed for each of the treatments (University of Florida control, and each of five sites). Site C was left out of subsequent analysis for publication, due to poor characterization of the site. Two Samples were left out of the normalization/analysis due to QC failure. Therefore, there are only 3 replicates of the "testis, site A water, exposed 48 h." and "liver, site E water, exposed 48 h." groups.

Project description:To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. To gain improved temporal, spatial and phylogenetic resolution of marine microbial communities, in this study we expanded the original prototype genome proxy array (an oligonucleotide microarray targeting marine microbial genome fragments and genomes), evaluated it against metagenomic sequencing, and applied it to time series samples from the Monterey Bay long term ecological research site. The expanded array targeted 268 microbial genotypes (vs. 14 in the original prototype) across much of the known diversity of cultured and uncultured marine microbes. The target abundances measured by the genome proxy array were highly correlated to pyrosequence-based abundances (linear regression R2 = 0.85-0.91, p<0.0001). Fifty-seven samples from ~4-years in Monterey Bay were examined with the array, spanning the photic zone (0m), the base of the surface mixed layer (30m), and the subphotic zone (200m). A significant portion of the expanded genome proxy array’s targets showed signal (95 out of 268 targets present in ≥ 1 sample). The multi-year community survey showed the consistent presence of a core group of common and abundant targeted taxa at each depth in Monterey Bay, higher variability among shallow than deep samples, and episodic occurrences of more transient marine genotypes. The abundance of the most dominant genotypes peaked after strong episodic upwelling events. The genome-proxy array’s ability to track populations of closely-related genotypes indicated population shifts within several abundant target taxa, with specific populations in some cases clustering by depth or oceanographic season. Although 51 cultivated organisms were targeted (representing 19% of the array) the majority of targets detected and of total target signal (85% and ~92%, respectively) were from uncultivated lineages, often those derived from Monterey Bay. The array provided cost-effective (~$15 per array, for construction and use) insights into the natural history of uncultivated lineages in the wild. The expanded genome proxy array was designed as in (Rich et al., 2008). Briefly, each genotype was targeted using suites of ~20 70-mer oligonucleotide probes designed using the program ArrayOligoSelector (Zhu et al., 2003). Probes had approximately the same %GC (40%) and were distributed across the target genome or genome fragment, with no more than one probe per gene and avoiding 16S and 23S rRNA genes. The array included positive and negative control probes designed using the same method, to Halobacterium salinarum NRC-1 and a random genome sequence, respectively. The expanded array had a broader scope than the prototype of Rich et al., 2008 (268 target genotypes, as opposed to the prototype’s 14) and included a co-spot oligo for spot alignment and gridding purposes (using the “alien” oligo sequence of (Urisman et al., 2005). The targets were selected from fully-sequenced marine microbial genomes, publicly-available marine-derived BAC and fosmid clone sequences, and fully-sequenced clones from the lab’s Monterey Bay and Hawaii environmental BAC- and fosmid-based genomic libraries. Targeted genotypes are detailed in Table S1, summarized in Table S2, and presented in a schematic phylogenetic overview in Fig. 1. Previously-unpublished sequences used for array design were submitted to Genbank under accession numbers GU474833-GU474949. This submission presents the analysis of 57 samples, each hybridized to >= 3 arrays. In addition to negative control probes, for each hyrbdization date and run a negative control array was hyrbdized for which the input DNA was TE, taken through the entire aplification, labeling, and hybridization process.

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Surface waters in Zhanjiang Bay

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