Niche Partition of Bacteriovorax Operational Taxonomic Units Along Salinity and Temporal Gradients in the Chesapeake Bay Reveals Distinct Estuarine Strains

The predatory Bacteriovorax are Gram-negative bacteria ubiquitous in saltwater systems that prey upon other Gram-negative bacteria in a similar manner to the related genus Bdellovibrio. Among the phylogenetically defined clusters of Bacteriovorax, cluster V has only been isolated from estuaries suggesting that it may be a distinct estuarine phylotype. To assess this hypothesis, the spatial and temporal distribution of cluster V and other Bacteriovorax phylogenetic assemblages along the salinity gradient of Chesapeake Bay were determined. Cluster V was expected to be found in significantly greater numbers in low to moderate salinity waters compared to high salinity areas. The analyses of water and sediment samples from sites in the bay revealed cluster V to be present at the lower salinity and not high salinity sites, consistent with it being an estuarine phylotype. Cluster IV had a similar distribution pattern and may also be specifically adapted to estuaries. While the distribution of clusters V and IV were similar for salinity, they were distinct on temperature gradients, being found in cooler and in warmer temperatures, respectively. The differentiation of phylotype populations along the salinity and temporal gradients in Chesapeake Bay revealed distinct niches inhabited by different phylotypes of Bacteriovorax and unique estuarine phylotypes.     

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.     

Nitrogen-fixing phylotypes of Chesapeake Bay and Neuse River estuary sediments

Sediments often exhibit low rates of nitrogen fixation, despite the presence of elevated concentrations of inorganic nitrogen. The organisms that potentially fix nitrogen in sediments have not previously been identified. Amplification of nifH genes with degenerate primers was used to assess the diversity of diazotrophs in two distinct sediment systems, anoxic muds of Chesapeake Bay and shallow surficial sediments of the Neuse River. Phylogenetic analysis revealed that sequences obtained from mid-Chesapeake Bay, which receive high organic loading and are highly reducing, clustered closely with each other and with known anaerobic microorganisms, suggesting a low abundance of aerobic or facultative diazotrophs in these sediments. Sulfate reduction dominates in the surface, but methanogenesis becomes more important with depth. A thin (<1 cm) oxidized layer is present only in the spring. No archaeal nifH sequences were obtained from Chesapeake Bay. Sequences of nifH amplified from surficial sediments of the Neuse River were distant from Chesapeake Bay sequences and included nif phylotypes related to sequences previously reported from marine mats and the Spartina rhizosphere. Differences in environmental site characteristics appear to select for different types of sediment diazotrophs, which is reflected in the phylogenetic composition of amplified nifH sequences.     

Actinomycetal Community Structures in Seawater and Freshwater Examined by DGGE Analysis of 16S rRNA Gene Fragments

The actinomycetal community structures in marine and freshwater environments (the Pacific Ocean, East China Sea, Tokyo Bay, and Arakawa River) were investigated by a culture-independent molecular method to clarify spatial and seasonal distributions. Deoxyribonucleic acid (DNA) was extracted from environmental water samples, and a community analysis was carried out on polymerase chain reaction-amplified 16S ribosomal DNA. The amplified DNA fragments were analyzed by denaturing gradient gel electrophoresis (DGGE) and nonmetric multidimensional scaling analysis, followed by sequencing analysis. The actinomycetal community structures were different at each station in the Pacific Ocean, the East China Sea, Tokyo Bay, and Arakawa River, and different populations predominated in each area. There were vertical variations in actinomycetal communities in the Pacific Ocean and East China Sea between the surface and 100-m depth, but communities were similar from 200- to 1,000-m depths. There were also distinct seasonal variations in communities in Tokyo Bay. Phylogenetic analysis of DNA fragments recovered from DGGE bands revealed that most of the predominant actinomycetal strains were uncultured and were quite different from well known culturable strains, such as the Streptomyces, Micromonospora, Microbispora, Salinispora, and Actinoplanes groups. These results suggest that the marine environment is an attractive target for discovering new actinomycetal populations producing bioactive compounds and that sampling depth and season are important considerations for isolating various populations effectively.     

Diverse and dynamic populations of cyanobacterial podoviruses in the Chesapeake Bay unveiled through DNA polymerase gene sequences

Many podoviruses have been isolated which infect marine picocyanobacteria, and they may play a potentially important role in regulating the biomass and population composition of picocyanobacteria. However, little is known about the diversity and population dynamics of autochthonous cyanopodoviruses in marine environments. Using a set of newly designed PCR primers which specifically amplify the DNA pol from cyanopodoviruses, a total of 221 DNA pol sequences were retrieved from eight Chesapeake Bay virioplankton communities collected at different times and locations. All DNA pol sequences clustered with the eight known podoviruses that infect different marine picocyanobacteria, and could be divided into at least 10 different subclusters (I-X). The presence of these cyanopodovirus genotypes based on PCR-amplification of DNA pol gene sequences was supported by the existence of similar DNA pol genotypes with metagenome libraries of Chesapeake Bay virioplankton assemblages. The composition of cyanopodoviruses in the Bay also exhibited distinct winter and summer patterns which were likely related to corresponding seasonal changes in the composition of cyanobacterial populations. Our study suggests that diverse and dynamic populations of cyanopodoviruses are present in the estuarine environment. The PCR method developed in this study provides a specific and sensitive tool to explore the abundance, distribution and phylogenetic diversity of cyanopodoviruses in aquatic environments. Linking the dynamics of host and viral populations in the natural environment is critical to broader characterization of the ecological role of virioplankton within microbial communities.     

Actinorhodopsins: proteorhodopsin-like gene sequences found predominantly in non-marine environments

Proteorhodopsins are light-energy-harvesting transmembrane proteins encoded by genes recently discovered in the surface waters of the world's oceans. Metagenomic data from the Global Ocean Sampling expedition (GOS) recovered 2674 proteorhodopsin-related sequences from 51 aquatic samples. Four of these samples were from non-marine environments, specifically, Lake Gatun within the Panama Canal, Delaware Bay and Chesapeake Bay and the Punta Cormorant Lagoon in Ecuador. Rhodopsins related to but phylogenetically distinct from most sequences designated proteorhodopsins were present at all four of these non-marine sites and comprised three different clades that were almost completely absent from marine samples. Phylogenomic analyses of genes adjacent to those encoding these novel rhodopsins suggest affiliation to the Actinobacteria , and hence we propose to name these divergent, non-marine rhodopsins 'actinorhodopsins'. Actinorhodopsins conserve the acidic amino acid residues critical for proton pumping and their genes lack genomic association with those encoding photo-sensory transducer proteins, thus supporting a putative ion pumping function. The ratio of rec A and rad A to rhodopsin genes in the different environment types sampled within the GOS indicates that rhodopsins of one type or another are abundant in microbial communities in freshwater, estuarine and lagoon ecosystems, supporting an important role for these photosystems in all aquatic environments influenced by sunlight.     

Diversity and Distribution of Sediment <Emphasis Type="Italic">NirS</Emphasis>-Encoding Bacterial Assemblages in Response to Environmental Gradients in the Eutrophied Jiaozhou Bay,China

A gene-clone-library-based molecular approach was used to study the nirS-encoding bacteria–environment relationship in the sediments of the eutrophic Jiaozhou Bay. Diverse nirS sequences were recovered and most of them were related to the marine cluster I group, ubiquitous in estuarine, coastal, and marine environments. Some NirS sequences were unique to the Jiaozhou Bay, such as the marine subcluster VIIg sequences. Most of the Jiaozhou Bay NirS sequences had their closest matches originally detected in estuarine and marine sediments, especially from the Chesapeake Bay, indicating similarity of the denitrifying bacterial communities in similar coastal environments in spite of geographical distance. Multivariate statistical analyses indicated that the spatial distribution of the nirS-encoding bacterial assemblages is highly correlated with environmental factors, such as sediment silt content, NH₄ ⁺ concentration, and OrgC/OrgN. The nirS-encoding bacterial assemblages in the most hypernutrified stations could be easily distinguished from that of the least eutrophic station. For the first time, the sedimentological condition was found to influence the structure and distribution of the sediment denitrifying bacterial community. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Isolation of Vibrio parahaemolyticus from the Processed Meat of Chesapeake Bay Blue Crabs

A method for the recovery of Vibrio parahaemolyticus from seafoods is described. By this procedure, a total of 56 biochemically positive cultures of V. parahaemolyticus were recovered from market samples of Chesapeake Bay processed blue crab (cooked, picked, packed, and refrigerated meat). All of the isolates were tested serologically, and 22 strains were serotyped according to the schema of Sakazaki as follows: K3, K5, K28, K31, K36, K37, K39, K43, and K44. These results indicate the broad distribution of these specific serotypes in a seafood harvested from the Chesapeake Bay.     

Molecular genetic variation within and among isolates of QPX (Thraustochytridae), a parasite of the hard clam Mercenaria mercenaria

The thraustochytrid known as QPX (Quahog Parasite Unknown) has sporadically caused disease in the hard clam Mercenaria mercenaria along the east coast of North America since the 1960s. We hypothesized that genetically distinct QPX strains might be responsible for outbreaks of QPX disease in different areas and tested this hypothesis by comparing several QPX isolates recovered from the recent outbreak in Raritan Bay, New York with QPX strains isolated from 2 outbreaks in Massachusetts, USA. There was no variation in small subunit rDNA (SSU rDNA), 5.8S rDNA, or 4 mitochondrial gene sequences. In contrast, both of the ribosomal ribonucleic acid (rRNA) operon intergenic spacers, internal transcribed spacers 1 and 2 (ITS1 and ITS2), revealed substantial sequence variation. However, strain-specific sequences were not detected because the ITS sequence variation within QPX isolates was comparable to the variation between isolates. ITS1 sequences recovered from an infected clam by amplification with a QPX ITS2-specific primer were identical to those recovered from the QPX isolates.     

Multiple strains of the parasitic dinoflagellate Amoebophrya exist in Chesapeake Bay

Small subunit rRNA sequences were amplified from Amoebophrya strains infecting Karlodinium micrum, Gymnodinium instriatum and an unidentified Scrippsiella species in Chesapeake Bay. The alignable parts of the sequences differed from each other and from the previously reported rRNA sequence of the Amoebophrya strain infecting Akashiwo sanguinea in Chesapeake Bay by 4 to 10%. This is a greater degree of difference than sometimes found between sequences from separate genera of free-living dinoflagellates. These sequence differences indicate that the Amoebophrya strains parasitizing dinoflagellates in Chesapeake Bay do not all belong to the same species. In spite of their relative dissimilarity, the sequences do group together into a single clade with high bootstrap support in phylogenetic trees constructed from the sequences.     

Genetic diversity of Vibrio cholerae in Chesapeake Bay determined by amplified fragment length polymorphism fingerprinting

Vibrio cholerae is indigenous to the aquatic environment, and serotype non-O1 strains are readily isolated from coastal waters. However, in comparison with intensive studies of the O1 group, relatively little effort has been made to analyze the population structure and molecular evolution of non-O1 V. cholerae. In this study, high-resolution genomic DNA fingerprinting, amplified fragment length polymorphism (AFLP), was used to characterize the temporal and spatial genetic diversity of 67 V. cholerae strains isolated from Chesapeake Bay during April through July 1998, at four different sampling sites. Isolation of V. cholerae during the winter months (January through March) was unsuccessful, as observed in earlier studies (J. H. L. Kaper, R. R. Colwell, and S. W. Joseph, Appl. Environ. Microbiol. 37:91-103, 1979). AFLP fingerprints subjected to similarity analysis yielded a grouping of isolates into three large clusters, reflecting time of the year when the strains were isolated. April and May isolates were closely related, while July isolates were genetically diverse and did not cluster with the isolates obtained earlier in the year. The results suggest that the population structure of V. cholerae undergoes a shift in genotype that is linked to changes in environmental conditions. From January to July, the water temperature increased from 3 degrees C to 27.5 degrees C, bacterial direct counts increased nearly an order of magnitude, and the chlorophyll a concentration tripled (or even quadrupled at some sites). No correlation was observed between genetic similarity among isolates and geographical source of isolation, since isolates found at a single sampling site were genetically diverse and genetically identical isolates were found at several of the sampling sites. Thus, V. cholerae populations may be transported by surface currents throughout the entire Bay, or, more likely, similar environmental conditions may be selected for a specific genotype. The dynamic nature of the population structure of this bacterial species in Chesapeake Bay provides new insight into the ecology and molecular evolution of V. cholerae in the natural environment.     

Using molecular and ecological data to diagnose endangered populations of the puritan tiger beetle Cicindela puritana

Populations of the puritan tiger beetle Cicindela puritana in the eastern United States were found to be highly threatened at the Connecticut River, whereas several large populations on the western shore and newly discovered populations on the eastern shore of the Chesapeake Bay appeared to be less endangered. We assessed if the disjunct C. puritana subgroups are genetically distinct and therefore should be treated as separate units for conservation purposes. A total of 13 individuals from the Connecticut River and 27 individuals from the Chesapeake Bay were each analysed by sequencing of up to 837 base pairs of mitochondrial DNA per individual. Five different haplotypes could be distinguished. In a phylogenetic analysis of these DNA sequences that included four related Cicindela species as out-groups, haplotypes from the Chesapeake Bay represent a distinct clade. The conservation status of these populations was evaluated using a phylogenetic approach based on cladistic analysis and the framework of the phylogenetic species concept. According to this analysis, beetles from the Connecticut River and the Chesapeake Bay have to be considered as independent units. Populations from the eastern and western shore of Chesapeake Bay are not split in more than one unit using the same criteria, although they exhibited some degree of genetic subdivision. The results from the mtDNA analysis were corroborated by ecological parameters in that the Chesapeake Bay populations can be distinguished from all congeners by their different tat association.     

Analysis of a marine picoplankton community by 16S rRNA gene cloning and sequencing.

The phylogenetic diversity of an oligotrophic marine picoplankton community was examined by analyzing the sequences of cloned ribosomal genes. This strategy does not rely on cultivation of the resident microorganisms. Bulk genomic DNA was isolated from picoplankton collected in the north central Pacific Ocean by tangential flow filtration. The mixed-population DNA was fragmented, size fractionated, and cloned into bacteriophage lambda. Thirty-eight clones containing 16S rRNA genes were identified in a screen of 3.2 x 10(4) recombinant phage, and portions of the rRNA gene were amplified by polymerase chain reaction and sequenced. The resulting sequences were used to establish the identities of the picoplankton by comparison with an established data base of rRNA sequences. Fifteen unique eubacterial sequences were obtained, including four from cyanobacteria and eleven from proteobacteria. A single eucaryote related to dinoflagellates was identified; no archaebacterial sequences were detected. The cyanobacterial sequences are all closely related to sequences from cultivated marine Synechococcus strains and with cyanobacterial sequences obtained from the Atlantic Ocean (Sargasso Sea). Several sequences were related to common marine isolates of the gamma subdivision of proteobacteria. In addition to sequences closely related to those of described bacteria, sequences were obtained from two phylogenetic groups of organisms that are not closely related to any known rRNA sequences from cultivated organisms. Both of these novel phylogenetic clusters are proteobacteria, one group within the alpha subdivision and the other distinct from known proteobacterial subdivisions. The rRNA sequences of the alpha-related group are nearly identical to those of some Sargasso Sea picoplankton, suggesting a global distribution of these organisms.     

A Probabilistic Ecological Risk Assessment of Tributyltin in Surface Waters of the Chesapeake Bay Watershed

The goal of this study was to conduct a probabilistic ecological risk assessment for tributyltin (TBT) in surface waters of the Chesapeake Bay watershed. Ecological risk was characterized by comparing the probability distributions of environmental exposure concentrations with the probability distributions of species response data determined from laboratory studies. The overlap of these distributions was a measure of risk to aquatic life. Tributyltin exposure data from the Chesapeake Bay watershed were available from over 3600 water column samples from 41 stations in nine basins from 1985 through 1996. Most of the stations were located in the Virginia waters of Chesapeake Bay, primarily the James, Elizabeth and York Rivers. In Maryland waters of the Bay, various marina, harbor and river systems were also sampled. As expected, the highest environmental concentrations of tributyltin (based on 90th percentiles) were reported in and near marina areas. The sources of TBT causing these high concentrations were primarily boat hulls and painting/depainting operations. Lower concentrations of TBT were reported in open water areas, such as the Potomac River, Choptank River and C and D Canal, where the density of boats was minimal. Temporal data from a ten year data base (1986-1996) from two areas in Virginia showed that TBT water column concentrations have declined since 1987 legislation prohibited the use of TBT paints on recreation boats (<25?m). Acute saltwater and freshwater TBT toxicity data were available for 43 and 23 species, respectively. Acute effects for saltwater species were reported for concentrations exceeding 420?ng/L; the lowest acute value for a freshwater species was 1110?ng/L. The acute 10th percentiles for all saltwater and freshwater species were 320 and 103?ng/L, respectively. The order of sensitivity from most to least sensitive for saltwater trophic groups and corresponding acute 10th percentiles were as follows: zooplankton (5?ng/L), phytoplankton (124?ng/L), benthos (312?ng/L) and fish (1009?ng/L). For freshwater species, the order of sensitivity from most to least sensitive trophic groups and corresponding acute 10th percentiles were: benthos (44?ng/L), zooplankton (400?ng/L), and fish (849?ng/L). Chronic data for both saltwater and freshwater species were limited to a few species in each water type. Based on these limited data, the saltwater and freshwater chronic 10th percentiles were 5 and 102?ng/L, respectively. Limited mesocosm and microcosm studies in saltwater suggested that TBT concentrations less than 50?ng/L did not impact the structure and function of biological communities. The saltwater acute (320?ng/L) and chronic (5?ng/L) 10th percentiles were used to determine ecological risk because all exposure data were from saltwater areas of the Chesapeake Bay watershed. Highest ecological risk was reported for marina areas in Maryland waters of Chesapeake Bay and for areas in Virginia such as the Elizabeth River, Hampton Creek and Sarah Creek. Low ecological risk was reported for areas such as the Potomac River, Choptank River, C and D Canal and Norfolk Harbor. Regulation of TBT on recreational watercraft in 1987 has successfully reduced water column concentrations of this organometallic compound. However, various studies have showed that TBT may remain in the sediment for years and continue to be source for water column exposures.     

Diverse and unique picocyanobacteria in Chesapeake Bay, revealed by 16S-23S rRNA internal transcribed spacer sequences

rRNA internal transcribed spacer phylogeny showed that Chesapeake Bay is populated with diverse Synechococcus strains, including members of the poorly studied marine cluster B. Marine cluster B prevailed in the upper bay, while marine cluster A was common in the lower bay. Interestingly, marine cluster B Synechococcus included phycocyanin- and phycoerythrin-rich strains.     

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.     

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.     

Incidence of Vibrio parahaemolyticus in Chesapeake Bay

A Bay-wide survey of the distribution of Vibrio parahaemolyticus was carried out in Chesapeake Bay during May 1972, to determine whether the annual cycle of V. parahaemolyticus which was observed to occur in the Rhode River subestuary of Chesapeake Bay took place in other parts of Chesapeake Bay. In an earlier study, April to early June, when the water temperature rises from 14 to 19 C, was found to be a critical period in the annual cycle of the organism in the Rhode River, since this is the time period when the annual cycle is initiated. Results of this study, however, revealed that V. parahaemolyticus could not be found in the water column during May 1972. Nevertheless, several samples of sediment and plankton yielded V. parahaemolyticus isolates. Comparison of data with those for the Rhode River area examined in the earlier studies of the annual cycle of V. parahaemolyticus suggests that the time of initiation of the annual cycle of V. parahaemolyticus in the open Bay proper may be influenced by various factors such as temperature and salinity, i.e., deeper water locations may show initiation of the V. parahaemolyticus annual cycle later than shallow areas. Confirmation of the presence of the organisms in the samples studied was accomplished using numerical taxonomy with 19 reference strains also included in the analyses.     

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 degrees 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.