Nitrogenase gene expression in the Chesapeake Bay Estuary

Like many estuaries, the Chesapeake Bay has pronounced gradients in salinity and nutrients. Previous studies have shown that there is a high diversity of nitrogenase (nifH) genes in the estuary, and that there are specific distributions of individual nifH phylotypes. In contrast to previous work that revealed the remarkable diversity of nifH phylotypes in the Chesapeake estuary, in this study of nifH expression we only detected two phylotypes, and both were phylogenetically related to cyanobacterial nifH genes. One of the phylotypes was closely related to a nifH sequence from the filamentous, heterocystous cyanobacterium Anabaena cylindrica, and was found at the head of the estuary. The other phylotype was found in a sample collected near the mouth of the estuary and was closely related to nifH sequences from Group A unicellular cyanobacteria, which has previously been reported in oceanic waters only. These nifH phylotypes had distinct patterns of expression that were restricted to different regions of the Chesapeake Bay. This study provides the first evidence of nifH expression in the Chesapeake Bay, and suggests that diazotrophic unicellular cyanobacteria have a broader distribution and activity than previously recognized.     

Global survey of diversity among environmental saltwater Bacteriovoracaceae

Halophilic Bacteriovorax (Bx), formerly known as the marine Bdellovibrio, are Gram-negative, predatory bacteria found in saltwater systems. To assess their genetic diversity and geographical occurrence, the small subunit rRNA (ssu-rRNA) gene sequences were analysed from 111 marine, salt lake and estuarine isolates recovered from 27 locations around the world. Phylogenetic analysis of these isolates using Geobacter as the outgroup revealed eight distinct ribotype clusters each with at least two isolates. Each cluster was composed of isolates with >or= 96.5% similarity in ssu-rRNA sequences. Three single isolate outliers were observed. Many of the Bx ribotypes were widely dispersed among different types of ecosystems (e.g. cluster III was recovered from the Great Salt Lake, the Atlantic Ocean, Pacific Ocean, Chesapeake Bay and gills of aquarium fish). However, cluster V was only recovered from a single ecosystem, estuaries. Cluster V was originally detected in the Chesapeake Bay and subsequently in the Pamlico Sound/Neuse River system. Principal coordinate analysis revealed that the sequences of the isolates from different environments were distinct from each other. The results of this study reveal the saltwater Bx to be phylogenetically and environmentally more diverse than was previously known.     

Diversity and Seasonality of Bioluminescent Vibrio cholerae Populations in Chesapeake Bay

Association of luminescence with phenotypic and genotypic traits and with environmental parameters was determined for 278 strains of Vibrio cholerae isolated from the Chesapeake Bay during 1998 to 2000. Three clusters of luminescent strains (A, B, and C) and two nonluminescent clusters (X and Y) were identified among 180 clonal types. V. cholerae O1 strains isolated during pandemics and endemic cholera in the Ganges Delta were related to cluster Y. Heat-stable enterotoxin (encoded by stn) and the membrane protein associated with bile resistance (encoded by ompU) were found to be linked to luminescence in strains of cluster A. Succession from nonluminescent to luminescent populations of V. cholerae occurred during spring to midsummer. Occurrence of cluster A strains in water with neutral pH was contrasted with that of cluster Y strains in water with a pH of >8. Cluster A was found to be associated with a specific calanoid population cooccurring with cyclopoids. Cluster B was related to cluster Y, with its maximal prevalence at pH 8. Occurrence of cluster B strains was more frequent with warmer water temperatures and negatively correlated with maturity of the copepod community. It is concluded that each cluster of luminescent V. cholerae strains occupies a distinct ecological niche. Since the dynamics of these niche-specific subpopulations are associated with zooplankton community composition, the ecology of luminescent V. cholerae is concluded to be related to its interaction with copepods and related crustacean species.     

Spatial and Temporal Distribution of Two Diazotrophic Bacteria in the Chesapeake Bay

The aim of this study was to initiate autecological studies on uncultivated natural populations of diazotrophic bacteria by examining the distribution of specific diazotrophs in the Chesapeake Bay. By use of quantitative PCR, the abundance of two nifH sequences (907h22 and 912h4) was quantified in water samples collected along a transect from the head to the mouth of the Chesapeake Bay during cruises in April and October 2001 and 2002. Standard curves for the quantitative PCR assays demonstrated that the relationship between gene copies and cycle threshold was linear and highly reproducible from 1 to 10⁷ gene copies. The maximum number of 907h22 gene copies detected was approximately 140 ml⁻¹ and the maximum number of 912h4 gene copies detected was approximately 340 ml⁻¹. Sequence 912h4 was most abundant at the mouth of the Chesapeake Bay, and in general, its abundance increased with increasing salinity, with the highest abundances observed in April 2002. Overall, the 907h22 phylotype was most abundant at the mid-bay station. Additionally, 907h22 was most abundant in the April samples from the mid-bay and mouth of the Chesapeake Bay. Despite the fact that the Chesapeake Bay is rarely nitrogen limited, our results show that individual nitrogen-fixing bacteria have distinct nonrandom spatial and seasonal distributions in the Chesapeake Bay and are either distributed by specific physical processes or adapted to different environmental niches.     

Ecology, serology, and enterotoxin production of Vibrio cholerae in Chesapeake Bay.

A total of 65 isolates of Vibrio cholerae, serotypes other than O--1, have been recovered from water, sediment, and shellfish samples from the Chesapeake Bay. Isolations were not random, but followed a distinct pattern in which salinity appeared to be a controlling factor in V. cholerae distribution. Water salinity at stations yielding V. cholerae (13 out of 21 stations) was 4 to 17 0/00, whereas the salinity of water at stations from which V. cholerae organisms were not isolated was less than 4 or greater than 17 0/00. From results of statistical analyses, no correlation between incidence of fecal coliforms and V. cholerae could be detected, whereas incidence of Salmonella species, measured concurrently, was clearly correlated with fecal coliforms, with Salmonella isolated only in areas of high fecal coliform levels. A seasonal cycle could not be determined since strains of V. cholerae were detectable at low levels (ca. 1 to 10 cells/liter) throughout the year. Although none of the Chesapeake Bay isolates was agglutinable in V. cholerae O group 1 antiserum, the majority for Y-1 adrenal cells. Furthermore, rabbit ileal loop and mouse lethality tests were also positive for the Chesapeake Bay isolates, with average fluid accumulation in positive ileal loops ranging from 0.21 to 2.11 ml/cm. Serotypes of the strains of V. cholerae recovered from Chesapeake Bay were those of wide geographic distribution. It is concluded from the data assembled to date, that V. cholerae is an autochthonous estuarine bacterial species resident in Chesapeake Bay.     

Abundance and distribution of Synechococcus spp. and cyanophages in the Chesapeake Bay

Despite the increasing knowledge of Synechococcus spp. and their co-occurring cyanophages in oceanic and coastal water, little is known about their abundance, distribution, and interactions in the Chesapeake Bay estuarine ecosystem. A 5-year interannual survey shows that Synechococcus spp. and their phages are persistent and abundant members of Chesapeake Bay microbial communities. Synechococcus blooms (10⁶ cells ml⁻¹) were often observed in summer throughout the Bay, contributing 20 to 40% of total phytoplankton chlorophyll a. The distribution of phycoerythrin-containing (PE-rich) Synechococcus cells appeared to mostly correlate with the salinity gradient, with higher abundances at higher salinities. Cyanophages infectious to Synechococcus were also abundant (up to 6 × 10⁵ viruses ml⁻¹ by the most probable number assay) during summer months in the Bay. The covariation in abundance of Synechococcus spp. and cyanophages was evident, although the latitude of observed positive correlation varied in different years, mirroring the changing environmental conditions and therefore the host-virus interactions. The impacts of cyanophages on host Synechococcus populations also varied spatially and temporally. Higher phage-related Synechococcus mortality was observed in drought years. Virus-mediated host mortality and subsequent liberation of dissolved organic matter (DOM) may substantially influence oceanic biogeochemical processing through the microbial loop as well as the microbial carbon pump. These observations emphasize the influence of environmental gradients on natural Synechococcus spp. and their phage population dynamics in the estuarine ecosystem.     

Predictability of Vibrio cholerae in Chesapeake Bay

Vibrio cholerae is autochthonous to natural waters and can pose a health risk when it is consumed via untreated water or contaminated shellfish. The correlation between the occurrence of V. cholerae in Chesapeake Bay and environmental factors was investigated over a 3-year period. Water and plankton samples were collected monthly from five shore sampling sites in northern Chesapeake Bay (January 1998 to February 2000) and from research cruise stations on a north-south transect (summers of 1999 and 2000). Enrichment was used to detect culturable V. cholerae, and 21.1% (n = 427) of the samples were positive. As determined by serology tests, the isolates, did not belong to serogroup O1 or O139 associated with cholera epidemics. A direct fluorescent-antibody assay was used to detect V. cholerae O1, and 23.8% (n = 412) of the samples were positive. V. cholerae was more frequently detected during the warmer months and in northern Chesapeake Bay, where the salinity is lower. Statistical models successfully predicted the presence of V. cholerae as a function of water temperature and salinity. Temperatures above 19°C and salinities between 2 and 14 ppt yielded at least a fourfold increase in the number of detectable V. cholerae. The results suggest that salinity variation in Chesapeake Bay or other parameters associated with Susquehanna River inflow contribute to the variability in the occurrence of V. cholerae and that salinity is a useful indicator. Under scenarios of global climate change, increased climate variability, accompanied by higher stream flow rates and warmer temperatures, could favor conditions that increase the occurrence of V. cholerae in Chesapeake Bay.     

Evolutionary Dynamics of Ralstonia solanacearum

We investigated the genetic diversity, extent of recombination, natural selection, and population divergence of Ralstonia solanacearum samples obtained from sources worldwide. This plant pathogen causes bacterial wilt in many crops and constitutes a serious threat to agricultural production due to its very wide host range and aggressiveness. Five housekeeping genes, dispersed around the chromosome, and three virulence-related genes, located on the megaplasmid, were sequenced from 58 strains belonging to the four major phylogenetic clusters (phylotypes). Whereas genetic variation is high and consistent for all housekeeping loci studied, virulence-related gene sequences are more diverse. Phylogenetic and statistical analyses suggest that this organism is a highly diverse bacterial species containing four major, deeply separated evolutionary lineages (phylotypes I to IV) and a weaker subdivision of phylotype II into two subgroups. Analysis of molecular variations showed that the geographic isolation and spatial distance have been the significant determinants of genetic variation between phylotypes. R. solanacearum displays high clonality for housekeeping genes in all phylotypes (except phylotype III) and significant levels of recombination for the virulence-related egl and hrpB genes, which are limited mainly to phylotype strains III and IV. Finally, genes essential for species survival are under purifying selection, and those directly involved in pathogenesis might be under diversifying selection.     

Diversity of ammonia monooxygenase (amoA) genes across environmental gradients in Chesapeake Bay sediments

Chesapeake Bay, the largest estuary in North America, encompasses a wide range of nutrient loading and trophic levels from the rivers and upper Bay to the sea, providing an ideal natural environment in which to explore relationships between functional diversity, physical/chemical complexity and ecosystem function (e.g. nitrification). In this study, amoA gene fragments (encoding subunit A of the key nitrification enzyme, ammonia monooxygenase) were PCR‐amplified from DNA extracted from sediment cores collected at five stations spanning gradients of salinity, ammonium, nitrate, oxygen and organic carbon along the Bay and Choptank River, a subestuary of the Bay. Phylogenetic analysis of ～30 amoA clones from each station revealed extensive diversity within the β‐Proteobacteria group of ammonia‐oxidizing bacteria (AOB), with the vast majority of sequences falling into coherent phylogenetic clusters distinct from sequences of cultivated AOB. Over 70% of the clones fell into two major phylogenetic clusters that appear to represent novel groups of Nitrosomonas‐like and Nitrosospira‐like amoA sequences that may be specific to estuarine and marine environments. Rarefaction analysis, estimators of genetic variation and dissimilarity indices all revealed differences in the relative amoA‐based diversity and/or richness among most of the stations, with the highest diversity at the North Bay station and the lowest at the mesohaline stations. Although salinity appears to play a role, no single physical or chemical parameter entirely explains the pattern of diversity along the estuary, suggesting that a complex combination of environmental factors may shape the overall level of AOB diversity in this dynamic environment.     

Differential effects of abiotic factors and host plant traits on diversity and community composition of root-colonizing arbuscular mycorrhizal fungi in a salt-stressed ecosystem

Arbuscular mycorrhizal fungi (AMF) were investigated in roots of 18 host plant species in a salinized south coastal plain of Laizhou Bay, China. From 18 clone libraries of 18S rRNA genes, all of the 22 AMF phylotypes were identified into Glomus, of which 18 and 4 were classified in group A and B in the phylogenetic tree, respectively. The phylotypes related to morphologically defined Glomus species occurred generally in soil with higher salinity. AMF phylotype richness, Shannon index, and evenness were not significantly different between root samples from halophytes vs. non-halophytes, invades vs. natives, or annuals vs. perennials. However, AMF diversity estimates frequently differed along the saline gradient or among locations, but not among pH gradients. Moreover, UniFrac tests showed that both plant traits (salt tolerance, life style or origin) and abiotic factors (salinity, pH, or location) significantly affected the community composition of AMF colonizers. Redundancy and variation partitioning analyses revealed that soil salinity and pH, which respectively explained 6.9 and 4.2 % of the variation, were the most influential abiotic variables in shaping the AMF community structure. The presented data indicate that salt tolerance, life style, and origin traits of host species may not significantly affect the AMF diversity in roots, but do influence the community composition in this salinized ecosystem. The findings also highlight the importance of soil salinity and pH in driving the distribution of AMF in plant and soil systems.     

Diversity and composition of sulfate- and sulfite-reducing prokaryotes as affected by marine-freshwater gradient and sulfate availability

Sulfate- and sulfite-reducing prokaryotes (SSRP) communities play a key role in both sulfur and carbon cycles. In estuarine ecosystems, sulfate concentrations change with tides and could be limited in tidal freshwater reach or deep sediments. In a subtropical estuary of northern Taiwan in December 2007, we examined the compositional changes of SSRP communities. We examined three sites: from the lower estuarine brackish-water reach (site GR and mangrove vegetation site, GM) to the upper estuarine tidal freshwater reach (site HR), as well as from surface to a 50-cm depth. The partial sequence of sulfite reductase (dsrB) genes was used as a molecular marker of SSRP, linked to polymerase chain reaction and denaturing gradient gel electrophoresis (DGGE) techniques. SSRP communities of the DGGE profiles varied with sites according to one-way analyses of similarities (Global R = 0.69, P = 0.001). Using cluster analysis, the DGGE profile was found to show site-specific clusters and a distinct depth zonation (five, six, and two SSRP communities at the GM, GR, and HR sites, respectively). SSRP composition was highly correlated to the combination of salinity, reduced sulfur, and total organic carbon contents (BIO-ENV analysis, r _s  = 0.56). After analyzing a total of 35 dsrB sequences in the DGGE gel, six groups with 15 phylotypes were found, which were closely related to marine-freshwater gradient. Moreover, sequences neighboring sulfite-reducing prokaryotes were observed, in addition to those affiliated to sulfate-reducing prokaryotes. Four phylotypes harvested in HR resembled the genus Desulfitobacterium, a sulfite-reducing prokaryote, which failed to use sulfate as an electron acceptor and were active in freshwater and sulfate-limited habitat. The other five phylotypes in the HR reach belonged to the sulfate-reducing prokaryotes of the genera Desulfatiferula, Desulfosarcina, Desulfovibrio, and Desulfotomaculum, which appeared to tolerate low salinity and low sulfate supply. SSRP phylotypes at the mangrove-vegetated GM site (five phylotypes in two groups) were phylogenetically less diverse, when compared with those at the non-mangrove-vegetated GR site (three phylotypes in three groups) and the tidally influenced freshwater HR site (nine phylotypes in five groups). Phylotypes found at GR and GM were all affiliated to marine sulfate-reducing prokaryote strains of the genera Desulfofaba, Desulfobotulus, Desulfatiferula, Desulfosarcina, and Desulfotomaculum. Notably, a phylotype recorded in the surface sediment at GR resembled the genus Desulfobulbus, which was recorded from freshwater environment consisting of the freshwater input at GR during ebb tides.     

Contrasting recombination patterns and demographic histories of the plant pathogen Ralstonia solanacearum inferred from MLSA

We used multilocus sequence analysis (MLSA) on a worldwide collection of the plant pathogenic Ralstonia solanacearum (Betaproteobacteria) to retrace its complex evolutionary history. Using genetic imprints left during R. solanacearum evolution, we were able to delineate distinct evolutionary complex displaying contrasting dynamics. Among the phylotypes already described (I, IIA, IIB, III, IV), eight groups of strains with distinct evolutionary patterns, named clades, were identified. From our recombination analysis, we identified 21 recombination events that occurred within and across these lineages. Although appearing the most divergent and ancestral phylotype, phylotype IV was inferred as a gene donor for the majority of the recombination events that we detected. Whereas this phylotype apparently fuelled the species diversity, ongoing diversification was mainly detected within phylotype I, IIA and III. These three groups presented a recent expanding population structure, a high level of homologous recombination and evidences of long-distance migrations. Factors such as adaptation to a specific host or intense trading of infected crops may have promoted this diversification. Whether R. solanacearum lineages will eventually evolve in distinct species remains an open question. The intensification of cropping and increase of geographical dispersion may favour situations of phylotype sympatry and promote higher exchange of key factors for host adaptation from their common genetic pool.     

Fingerprinting Diazotroph Communities in the Chesapeake Bay by Using a DNA Macroarray

Investigations of the distribution and diversity of nitrogen-fixing microorganisms in natural environments have often relied on PCR amplification and sequence analysis of a portion of one of the key enzymes in nitrogen fixation, dinitrogenase reductase, encoded by nifH. Recent work has suggested that DNA macroarrays provide semiquantitative fingerprints of diversity within mixtures of nifH amplicons (G. F. Steward, B. D. Jenkins, B. B. Ward, and J. P. Zehr, Appl. Environ. Microbiol. 70:1455-1465, 2004). Here we report the application of macroarrays for a study in the Chesapeake Bay. Samples from different locations in the bay yielded distinct fingerprints. Analysis of replicates and samples from different locations by cluster analysis showed that replicates clustered together, whereas different samples formed distinct clusters. There was a correspondence between the hybridization pattern observed and that predicted from the distribution of sequence types in a corresponding clone library. Some discrepancies between the methods were observed which are likely a result of the high nifH sequence diversity in the Chesapeake Bay and the limited number of sequences represented on this version of the array. Analyses of sequences in the clone library indicate that the Chesapeake Bay harbors unique, phylogenetically diverse diazotrophs. The macroarray hybridization patterns suggest that there are spatially variable communities of diazotrophs, which have been confirmed by quantitative PCR methods (S. M. Short, B. D. Jenkins, and J. P. Zehr, Appl. Environ. Microbiol., in press). The results show that DNA macroarrays have great potential for mapping the spatial and temporal variability of functional gene diversity in the environment.     

High Temporal but Low Spatial Heterogeneity of Bacterioplankton in the Chesapeake Bay

Compared to freshwater and the open ocean, less is known about bacterioplankton community structure and spatiotemporal dynamics in estuaries, particularly those with long residence times. The Chesapeake Bay is the largest estuary in the United States, but despite its ecological and economic significance, little is known about its microbial community composition. A rapid screening approach, ITS (internal transcribed spacer)-LH (length heterogeneity)-PCR, was used to screen six rRNA operon (16S rRNA-ITS-23S rRNA) clone libraries constructed from bacterioplankton collected in three distinct regions of the Chesapeake Bay over two seasons. The natural length variation of the 16S-23S rRNA gene ITS region, as well as the presence and location of tRNA-alanine coding regions within the ITS, was determined for 576 clones. Clones representing unique ITS-LH-PCR sizes were sequenced and identified. Dramatic shifts in bacterial composition (changes within subgroups or clades) were observed for the Alphaproteobacteria (Roseobacter clade, SAR11), Cyanobacteria (Synechococcus), and Actinobacteria, suggesting strong seasonal variation within these taxonomic groups. Despite large gradients in salinity and phytoplankton parameters, a remarkably homogeneous bacterioplankton community was observed in the bay in each season. Stronger seasonal, rather than spatial, variation of the bacterioplankton population was also supported by denaturing gradient gel electrophoresis and LH-PCR analyses, indicating that environmental parameters with stronger seasonal, rather than regional, dynamics, such as temperature, might determine bacterioplankton community composition in the Chesapeake Bay.     

Prey bacteria shape the community structure of their predators

Although predator–prey interactions among higher organisms have been studied extensively, only few examples are known for microbes other than protists and viruses. Among the bacteria, the most studied obligate predators are the Bdellovibrio and like organisms (BALOs) that prey on many other bacteria. In the macroscopical world, both predator and prey influence the population size of the other''s community, and may have a role in selection. However, selective pressures among prey and predatory bacteria have been rarely investigated. In this study, Bacteriovorax, a predator within the group of BALOs, in environmental waters were fed two prey bacteria, Vibrio vulnificus and Vibrio parahaemolyticus. The two prey species yielded distinct Bacteriovorax populations, evidence that selective pressures shaped the predator community and diversity. The results of laboratory experiments confirmed the differential predation of Bacteriovorax phylotypes on the two bacteria species. Not only did Bacteriovorax Cluster IX exhibit the versatility to be the exclusive efficient predator on Vibrio vulnificus, thereby, behaving as a specialist, but was also able to prey with similar efficiency on Vibrio parahaemolyticus, indicative of a generalist. Therefore, we proposed a designation of versatilist for this predator. This initiative should provide a basis for further efforts to characterize the predatory patterns of bacterial predators. The results of this study have revealed impacts of the prey on Bacteriovorax predation and in structuring the predator community, and advanced understanding of predation behavior in the microbial world.     

Spatial and temporal distribution of two diazotrophic bacteria in the Chesapeake Bay

The aim of this study was to initiate autecological studies on uncultivated natural populations of diazotrophic bacteria by examining the distribution of specific diazotrophs in the Chesapeake Bay. By use of quantitative PCR, the abundance of two nifH sequences (907h22 and 912h4) was quantified in water samples collected along a transect from the head to the mouth of the Chesapeake Bay during cruises in April and October 2001 and 2002. Standard curves for the quantitative PCR assays demonstrated that the relationship between gene copies and cycle threshold was linear and highly reproducible from 1 to 10(7) gene copies. The maximum number of 907h22 gene copies detected was approximately 140 ml(-1) and the maximum number of 912h4 gene copies detected was approximately 340 ml(-1). Sequence 912h4 was most abundant at the mouth of the Chesapeake Bay, and in general, its abundance increased with increasing salinity, with the highest abundances observed in April 2002. Overall, the 907h22 phylotype was most abundant at the mid-bay station. Additionally, 907h22 was most abundant in the April samples from the mid-bay and mouth of the Chesapeake Bay. Despite the fact that the Chesapeake Bay is rarely nitrogen limited, our results show that individual nitrogen-fixing bacteria have distinct nonrandom spatial and seasonal distributions in the Chesapeake Bay and are either distributed by specific physical processes or adapted to different environmental niches.     

Temporal and Spatial Variability in the Distribution of Vibrio vulnificus in the Chesapeake Bay: A Hindcast Study

Vibrio vulnificus, an estuarine bacterium, is the causative agent of seafood-related gastroenteritis, primary septicemia, and wound infections worldwide. It occurs as part of the normal microflora of coastal marine environments and can be isolated from water, sediment, and oysters. Hindcast prediction was undertaken to determine spatial and temporal variability in the likelihood of occurrence of V. vulnificus in surface waters of the Chesapeake Bay. Hindcast predictions were achieved by forcing a multivariate habitat suitability model with simulated sea surface temperature and salinity in the Bay for the period between 1991 and 2005 and the potential hotspots of occurrence of V. vulnificus in the Chesapeake Bay were identified. The likelihood of occurrence of V. vulnificus during high and low rainfall years was analyzed. From results of the study, it is concluded that hindcast prediction yields an improved understanding of environmental conditions associated with occurrence of V. vulnificus in the Chesapeake Bay.     

Predatory Bacteriovorax communities ordered by various prey species

The role of predation in altering microbial communities has been studied for decades but few examples are known for bacterial predators. Bacteriovorax are halophilic prokaryotes that prey on susceptible gram-negative bacteria. We recently reported novel observations on the differential selection of Bacteriovorax phylotypes by two different prey, Vibrio parahaemolyticus and Vibrio vulnificus. However, the conclusion is restricted by the limited number of prey tested. In this study, we have conducted two independent investigations involving eight species of prey bacteria while using V. vulnificus and V. parahaemolytics as reference strains. Water samples collected from Dry Bar, Apalachicola Bay were used to establish microcosms which were respectively spiked with prey strains Vibrio cholerae, Escherichia coli or Pseudomonas putida to examine the response of native Bacteriovorax to freshwater bacteria. Indigenous Vibrio sp., Pseudoalteromonas sp., Photobacterium sp. and a clinical strain of V. vulnificus were also tested for the impact of saltwater prey on the Bacteriovorax community. At 24 hour intervals, optical density of the microcosm samples and the abundance of Bacteriovorax were measured over five days. The predominant Bacteriovorax plaques were selected and analyzed by 16S rRNA gene amplification and sequencing. In addition, the impacts of prey on predator population and bacterial community composition were investigated using culture independent denaturing gradient gel electrophoresis. Strikingly, Cluster IV was found consistently as the predominant phylotype produced by the freshwater prey. For all saltwater prey, subgroups of Bacteriovorax phylotype IX were the major predators recovered. The results suggest that prey is an important factor along with temperature, salinity and other environmental parameters in shaping Bacteriovorax communities in aquatic systems.     

Spatial-temporal variability in diazotroph assemblages in Chesapeake Bay using an oligonucleotide nifH microarray

The distribution of nitrogen-fixing microorganisms in the Chesapeake Bay was investigated using fingerprints from a nifH microarray comprised of 706 60-mer oligonucleotide nifH probes representing cultivated organisms and environmental clones from different nifH clusters. Diverse nifH targets, amplified from samples using degenerate nifH primers, were detected in water column and sediment samples collected in April and October, 2001-2002. Total nifH richness and diversity (Simpson's and Shannon indices) were highest at the most riverine, oligohaline North Bay station. In most samples, the highest diversity was in nifH Cluster 3, which includes many anaerobes, while Cluster 1 (alpha-, beta- gamma- Proteobacteria, Cyanobacteria) targets had the greatest microarray signal intensities. In a multidimensional scaling analysis, deep water communities from April and October were similar within each of the sampling sites, while the surface communities had more variability. Diazotroph communities in the water column in the North Bay were distinct from the Mid- and South Bay communities, and there was a gradual change in sediment diazotroph assemblages from the North to the South Bay. Diazotrophic assemblages from the majority of the water column samples from the Mid- and South Bay clustered with the sediment assemblage in Mid-Bay. Dissolved inorganic nitrogen, salinity, dissolved organic carbon and dissolved organic phosphorus had a significant relationship with the diazotrophic bacterioplankton community. Higher diversity in the freshwater end of the system may reflect variability in disturbance rates and environmental conditions such as forms and concentrations of organic matter, nutrients and oxygen.     

Influence of Sire Breed on the Interplay among Rumen Microbial Populations Inhabiting the Rumen Liquid of the Progeny in Beef Cattle

This study aimed to evaluate whether the host genetic background impact the ruminal microbial communities of the progeny of sires from three different breeds under different diets. Eighty five bacterial and twenty eight methanogen phylotypes from 49 individuals of diverging sire breed (Angus, ANG; Charolais, CHA; and Hybrid, HYB), fed high energy density (HE) and low energy density (LE) diets were determined and correlated with breed, rumen fermentation and phenotypic variables, using multivariate statistical approaches. When bacterial phylotypes were compared between diets, ANG offspring showed the lowest number of diet-associated phylotypes, whereas CHA and HYB progenies had seventeen and twenty-three diet-associated phylotypes, respectively. For the methanogen phylotypes, there were no sire breed-associated phylotypes; however, seven phylotypes were significantly different among breeds on either diet (P<0.05). Sire breed did not influence the metabolic variables measured when high energy diet was fed. A correlation matrix of all pairwise comparisons among frequencies of bacterial and methanogen phylotypes uncovered their relationships with sire breed. A cluster containing methanogen phylotypes M16 (Methanobrevibacter gottschalkii) and M20 (Methanobrevibacter smithii), and bacterial phylotype B62 (Robinsoniella sp.) in Angus offspring fed low energy diet reflected the metabolic interactions among microbial consortia. The clustering of the phylotype frequencies from the three breeds indicated that phylotypes detected in CHA and HYB progenies are more similar among them, compared to ANG animals. Our results revealed that the frequency of particular microbial phylotypes in the progeny of cattle may be influenced by the sire breed when different diets are fed and ultimately further impact host metabolic functions, such as feed efficiency.