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Vibrio vulnificus was enumerated in seawater and shellfish from two Florida estuaries at selected seasonal intervals. There were significant fluctuations in the presence and numbers of V. vulnificus. Relatively high seawater temperature and salinity favored the presence of V. vulnificus in both seawater and shellfish samples.
Production of dissolved macromolecules by ambient autotrophic and heterotrophic microbial populations was measured in a eutrophic Florida reservoir by in situ labeling with various radioactive substrates. When [3H]thymidine was used as the precursor, production of labeled dissolved DNA, RNA, and protein was observed. The rate of production of labeled dissolved macromolecules was 3.1% the rate of cellular incorporation of [3H]thymidine, and the production of dissolved DNA represented 2.3% the rate of cellular DNA incorporation. Microautotrophic populations labeled with NaH[14C]CO3 produced dissolved RNA and protein at rates of 0.24 and 0.11 micrograms of C/liter per h, respectively, or 1.8% the total rate of carbon fixation, with no measurable dissolved DNA production. In an attempt to specifically label phytoplankton DNA, samples were incubated with [3H]adenine or 32Pi in the presence and absence of the photosynthetic inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). Although DCMU inhibited 14C fixation by approximately 99%, this antimetabolite had only a slight effect on [3H]adenine incorporation and no effect on 32P incorporation into cellular macromolecules. Significant amounts of dissolved DNA were produced in both [3H]adenine and 32Pi incubations, but again DCMU had no effect on the production rates. These results indicate that actively growing populations of both phytoplankton and bacterioplankton produced dissolved RNA and protein, while only active bacterioplankton produced measurable quantities of dissolved DNA. Dead or senescent phytoplankton may have produced dissolved DNA, but would not be measured in the relatively short incubations used. These findings also indicate that [3H]adenine and 32Pi primarily labeled heterotrophic bacterioplankton and not phytoplankton in this environment.
Sewage waste disposal facilities in the Florida Keys include septic tanks and individual package plants in place of municipal collection facilities in most locations. In Key Largo, both facilities discharge into the extremely porous Key Largo limestone. To determine whether there was potential contamination of the subsurface aquifer and nearby coastal surface waters by such waste disposal practices, we examined the presence of microbial indicators commonly found in sewage (fecal coliforms, Clostridium perfringens, and enterococci) and aquatic microbial parameters (viral direct counts, bacterial direct counts, chlorophyll a, and marine vibriophage) in injection well effluent, monitoring wells that followed a transect from onshore to offshore, and surface waters above these wells in two separate locations in Key Largo in August 1993 and March 1994. Effluent and waters from onshore shallow monitoring wells (1.8- to 3.7-m depth) contained two or all three of the fecal indicators in all three samples taken, whereas deeper wells (10.7- to 12.2-m depth) at these same sites contained few or none. The presence of fecal indicators was found in two of five nearshore wells (i.e., those that were < or = 1.8 miles [< or = 2.9 km] from shore), whereas offshore wells (> or = 2.1 to 5.7 miles [< or = 3.4 to 9.2 km] from shore) showed little sign of contamination. Indicators were also found in surface waters in a canal in Key Largo and in offshore surface waters in March but not in August. Collectively, these results suggest that fecal contamination of the shallow onshore aquifer, parts of the nearshore aquifer, and certain surface waters has occurred.(ABSTRACT TRUNCATED AT 250 WORDS)
As a result of agricultural activities in regions adjacent to the northern boundary of the Florida Everglades, a nutrient gradient developed that resulted in physicochemical and ecological changes from the original system. Sulfate input from agricultural runoff and groundwater is present in soils of the Northern Everglades, and sulfate-reducing prokaryotes (SRP) may play an important role in biogeochemical processes such as carbon cycling. The goal of this project was to utilize culture-based and non-culture-based approaches to study differences between the composition of assemblages of SRP in eutrophic and pristine areas of the Everglades. Sulfate reduction rates and most-probable-number enumerations revealed SRP populations and activities to be greater in eutrophic zones than in more pristine soils. In eutrophic regions, methanogenesis rates were higher, the addition of acetate stimulated methanogenesis, and SRP able to utilize acetate competed to a limited degree with acetoclastic methanogens. A surprising amount of diversity within clone libraries of PCR-amplified dissimilatory sulfite reductase (DSR) genes was observed, and the majority of DSR sequences were associated with gram-positive spore-forming Desulfotomaculum and uncultured microorganisms. Sequences associated with Desulfotomaculum fall into two categories: in the eutrophic regions, 94.7% of the sequences related to Desulfotomaculum were associated with those able to completely oxidize substrates, and in samples from pristine regions, all Desulfotomaculum-like sequences were related to incomplete oxidizers. This metabolic selection may be linked to the types of substrates that Desulfotomaculum spp. utilize; it may be that complete oxidizers are more versatile and likelier to proliferate in nutrient-rich zones of the Everglades. Desulfotomaculum incomplete oxidizers may outcompete complete oxidizers for substrates such as hydrogen in pristine zones where diverse carbon sources are less available.
A seasonal study of the distribution of lysogenic bacteria in Tampa Bay, Florida, was conducted over a 13-month period. Biweekly water samples were collected and either were left unaltered or had the viral population reduced by filtration (pore size, 0.2 micro m) and resuspension in filtered (pore size, 0.2 micro m) water. Virus-reduced and unaltered samples were then treated by adding mitomycin C (0.5 micro g ml(-1)) to induce prophage or were left untreated. In order to test the hypothesis that prophage induction was phosphate limited, additional induction experiments were performed in the presence and absence of phosphate. Induction was assessed as an increase in viral direct counts, relative to those obtained in controls, as detected by epifluorescence microscopy. Induction of prophage was observed in 5 of 25 (20%) unaltered samples which were obtained during or after the month of February, paralleling the results from a previous seasonal study. Induction of prophage was observed in 9 of 25 (36%) of the virus-reduced samples, primarily those obtained in the winter months, which was not observed in a prior seasonal study (P. K. Cochran and J. H. Paul, Appl. Environ. Microbiol. 64:2308-2312, 1998). Induction was noted in the months of lowest bacterial and primary production, suggesting that lysogeny was favored under conditions of poor host growth. Phosphate addition enabled prophage induction in two of nine (22%) experiments. These results indicate that prophage induction may occasionally be phosphate limited or respond to increases in phosphate concentration, suggesting that phosphate concentration may modulate the lysogenic response of natural populations.
Waters impacted by fecal pollution can exact high risks to human health and can result in financial losses due to closures of water systems used for recreation and for harvesting seafood. Identifying the sources of fecal pollution in water is paramount in assessing the potential human health risks involved as well as in assessing necessary remedial action. Recently, various researchers have used the ribotyping method to identify sources of bacterial indicators (Escherichia coli and enterococci) in environmental waters. While these studies have identified genotypic differences between human- and animal-derived indicators that are capable of differentiating organisms isolated from humans and various animal hosts, most have focused on organisms collected from a confined geographic area and have not addressed the question of whether these ribotype profiles are watershed specific or if they can be applied universally to organisms from other geographic locations. In this study, E. coli isolates were obtained from humans, beef cattle, dairy cattle, swine, and poultry from locations in northern, central, and southern Florida and were subjected to ribotyping analysis. The intent was to determine (i) if ribotype profiles are capable of discriminating the source of E. coli at the host species level and (ii) if the resulting fingerprints are uniform over an extended geographic area or if they can be applied only to a specific watershed. Our research indicated that, using a single restriction enzyme (HindIII), the ribotyping procedure is not capable of differentiating E. coli isolates from the different animal species sampled in this study. Results indicate, however, that this procedure can still be used effectively to differentiate E. coli as being either human or animal derived when applied to organisms isolated from a large geographic region.
Agricultural activities have produced well-documented changes in the Florida Everglades, including establishment of a gradient in phosphorus concentrations in Water Conservation Area 2A (WCA-2A) of the northern Everglades. An effect of increased phosphorus concentrations is increased methanogenesis in the eutrophic regions compared to the oligotrophic regions of WCA-2A. The goal of this study was to identify relationships between eutrophication and composition and activity of methanogenic assemblages in WCA-2A soils. Distributions of two genes associated with methanogens were characterized in soils taken from WCA-2A: the archaeal 16S rRNA gene and the methyl coenzyme M reductase gene. The richness of methanogen phylotypes was greater in eutrophic than in oligotrophic sites, and sequences related to previously cultivated and uncultivated methanogens were found. A preferential selection for the order Methanomicrobiales was observed in mcrA clone libraries, suggesting primer bias for this group. A greater diversity within the Methanomicrobiales was observed in mcrA clone libraries than in 16S rRNA gene libraries. 16S rRNA phylogenetic analyses revealed a dominance of clones related to Methanosaeta spp., an acetoclastic methanogen dominant in environments with low acetate concentrations. A significant number of clones were related to Methanomicrobiales, an order characterized by species utilizing hydrogen and formate as methanogenic substrates. No representatives of the orders Methanobacteriales and Methanococcales were found in any 16S rRNA clone library, although some Methanobacteriales were found in mcrA libraries. Hydrogenotrophs are the dominant methanogens in WCA-2A, and acetoclastic methanogen genotypes that proliferate in low acetate concentrations outnumber those that typically dominate in higher acetate concentrations.
Nutrient runoff from the Everglades Agricultural Area resulted in a well-documented gradient of phosphorus concentrations in soil and water, with concomitant ecosystem-level changes, in the northern Florida Everglades. It was recently reported that sulfate-reducing prokaryote assemblage composition, numbers, and activities are dependent on position along the gradient (H. Castro, K. R. Reddy, and A. Ogram, Appl. Environ. Microbiol. 68:6129-6137, 2002). The present study utilized a combination of culture- and non-culture-based approaches to study differences in composition of assemblages of syntrophic and methanogenic microbial communities in eutrophic, transition, and oligotrophic areas along the phosphorus gradient. Methanogenesis rates were much higher in eutrophic and transition regions, and sequence analysis of 16S rRNA gene clone libraries constructed from samples taken from these regions revealed differences in composition and activities of syntroph-methanogen consortia. Methanogens from eutrophic and transition regions were almost exclusively composed of hydrogenotrophic methanogens, with approximately 10,000-fold-greater most probable numbers of hydrogenotrophs than of acetotrophs. Most cultivable strains from eutrophic and transition regions clustered within novel lineages. In non-culture-based studies to enrich syntrophs, most bacterial and archaeal clones were either members of novel lineages or closely related to uncultivated environmental clones. Novel cultivable Methanosaeta sp. and fatty acid-oxidizing bacteria related to the genera Syntrophomonas and Syntrophobacter were observed in microcosms containing soil from eutrophic regions, and different lines of evidence indicated the existence of novel syntrophic association in eutrophic regions.
The Florida Everglades is one of the largest freshwater marshes in North America and has been subject to eutrophication for decades. A gradient in P concentrations extends for several kilometers into the interior of the northern regions of the marsh, and the structure and function of soil microbial communities vary along the gradient. In this study, stable isotope probing was employed to investigate the fate of carbon from the fermentation products propionate and butyrate in soils from three sites along the nutrient gradient. For propionate microcosms, 16S rRNA gene clone libraries from eutrophic and transition sites were dominated by sequences related to previously described propionate oxidizers, such as Pelotomaculum spp. and Syntrophobacter spp. Significant representation was also observed for sequences related to Smithella propionica, which dismutates propionate to butyrate. Sequences of dominant phylotypes from oligotrophic samples did not cluster with known syntrophs but with sulfate-reducing prokaryotes (SRP) and Pelobacter spp. In butyrate microcosms, sequences clustering with Syntrophospora spp. and Syntrophomonas spp. dominated eutrophic microcosms, and sequences related to Pelospora dominated the transition microcosm. Sequences related to Pelospora spp. and SRP dominated clone libraries from oligotrophic microcosms. Sequences from diverse bacterial phyla and primary fermenters were also present in most libraries. Archaeal sequences from eutrophic microcosms included sequences characteristic of Methanomicrobiaceae, Methanospirillaceae, and Methanosaetaceae. Oligotrophic microcosms were dominated by acetotrophs, including sequences related to Methanosarcina, suggesting accumulation of acetate.
Vibrio parahaemolyticus was isolated from various locations in two salt springs of Florida and appears to be a normal inhabitant of these artesian waters.
A rickettsia-like organism causing high mortality and gross pathology has been found infecting over 40% of a population of the freshwater crustacean, Crangonyx (near) floridanus Bousfield, in Florida. This is the first report of a rickettsia-like organism from a strictly aquatic animal. Morphological and developmental stages of the organisms range in size and shape from small, dense bacilliform cells measuring 150 nm in diameter by 395 nm in length, to large sausage-shaped cells 0.5 by 3 mum.
Four recent cases of Yersinia enterocolitica infections from Florida are discussed. Two of the cases presented as acute mesenteric lymphadenitis, and the other two presented as septicemia. Three of the isolates were an uncommon serotype, O:2,3, and the fourth was serotype O:5. The increasing occurrence of Y. enterocolitica in semitropical areas of the United States is emphasized.
A total of 202 methicillin-resistant Staphylococcus aureus (MRSA) single-patient isolates recovered between January and June 1998 in two hospitals in Miami, Florida, were characterized by a combination of several molecular typing techniques: multilocus sequence typing, spaA typing, pulsed-field gel electrophoresis, and determination of the structure of the SCCmec element. The overwhelming majority of the isolates-187 of 202, or 93%-belonged to one of three internationally spread epidemic clones which were identified on the basis of their multilocus sequence type (ST) as E-MRSA-16 (ST36), the New York clone V (ST8), and the New York/Japan clone (ST5; SCCmec II) and its single- and double-locus variants. The rest of the isolates (15 of 202, or 7%) were more genetically diverse and were each recovered from a few patients only. Of the 23 MRSA strains isolated from confirmed human immunodeficiency virus-positive patients, as many as 17 (or 70%) belonged to a single ST8 clone carrying SCCmec type IV. The data provide further evidence for the conclusion of earlier studies that most MRSA disease in hospitals is caused by relatively few pandemic clones.
We examined 299 methicillin-resistant, community-associated Staphylococcus aureus isolates from Florida and Washington State for the presence of the USA300 epidemic clone. Pulsed-field gel electrophoresis demonstrated the epidemic clone in 43% of our S. aureus strains and in isolates from both states. The majority of the USA300 isolates (88%) were from wound infections.
STUDY OBJECTIVE: To investigate a community wide outbreak of hepatitis A (HA). DESIGN: Description of the outbreak, with a case-control study to assess transmission. SETTING: A Florida county, USA, 1988-9. SUBJECTS: A total of 311 cases of HA. MEASUREMENTS AND MAIN RESULTS: A 13 month outbreak of HA is described. Most of the 311 cases (95%) were residents of a large metropolitan area (attack rate per 10,000 population (AR) = 3.7) and two smaller cities (AR = 61.5 and AR = 6.4). The ARs were greater for males than females and for residents aged 25-34 years (9.7) and < 5 years (8.3). Altogether 37% of cases were linked to day care centres, independent of the city of residence. A household case-control study showed an increased risk of HA in households in which a child attended a day care centre (p = 0.02), and centres that could take more than 50 children had an increased risk of HA introduction than smaller ones (p = 0.05). CONCLUSIONS: Day care centres were an important source of HA in the community, and the need for timely surveillance and immunoglobin prophylaxis is emphasised. Homosexual transmission may have played an important role in this outbreak.
OBJECTIVES: Hepatitis C virus (HCV) is the most common chronic blood borne viral infection in the United States. We assessed the HCV prevalence, risk factors, and sensitivity of the Centers for Disease Control and Prevention's (CDC) routine screening criteria among clients of a large urban sexually transmitted disease (STD) clinic. METHODS: Participants were recruited from a public STD clinic in Miami, Florida, and were interviewed regarding known and potential risk factors. The survey assessed CDC screening criteria, as well as other risk factors (for example, intranasal drug use, history of incarceration, exchanging sex for money, number of lifetime sex partners, and history of an STD). Testing was done by enzyme immunoassay (EIA) and confirmed by recombinant immunoblot assay (RIBA). RESULTS: The prevalence of anti-HCV positivity was 4.7%. Four variables were significantly associated with being anti-HCV positive, independent of confounding factors. These included injection drug use (odds ratio (OR) = 31.6; 95% confidence intervals (CI) 11.0 to 90.5); history of incarceration (OR = 3.0; 95% CI 1.1 to 8.1); sexual contact with an HCV positive person (OR 12.7; 95% CI 2.5 to 64.7); and older age (OR 1.4; 95% CI 1.2, 1.6). The sensitivity of CDC's routine screening criteria was 69% and specificity was 91%. CONCLUSIONS: The prevalence of anti-HCV in this clinic was similar to that determined in studies of comparable populations. Having sexual contact with an HCV positive person and history of incarceration were independently associated with being anti-HCV positive. CDC's screening criteria identified approximately two thirds of the anti-HCV positive participants.
Earlier virologic studies established that human T-cell lymphotropic virus type II (HTLV-II) is the predominant retrovirus type found among Seminole Indians in southern Florida. We studied 46 members of the Seminole tribe living on 3 reservations to determine the risk factors for HTLV-II and to investigate disease association with the virus. The donors' plasma samples were evaluated with the enzyme-linked immunosorbent and Western blot assays. DNA extracted from their peripheral blood mononuclear cells were analyzed by type-specific polymerase chain reaction amplification and detection of the HTLV pol gene using the primer pair SK110/SK111, and the probes SK112 or SK188. One of 46 (2%) subjects was identified as HTLV-I positive and 11 (24%) were identified as HTLV-II positive. Restriction fragment length polymorphism and sequence analyses indicated that all of the HTLV-II strains were subtype b. Mitochondrial DNA analyses indicated that all of the HTLV-II-positive subjects had an Amerindian haplotype. HTLV-II was more prevalent in Indians who were >45 years of age or female, had multiple sex partners or had received a blood transfusion. However, only the latter risk factor was statistically significant. Three of the HTLV-II-positive Indians demonstrated signs and symptoms of an ataxic neuropathy. The data support that HTLV-IIb is endemic among the Seminoles and that they will be a key population for further virologic studies.
The retina and choriocapillaris of the Florida garfish, Lepisosteus platyrhincus (Ginglymodi), was examined at the light and electron microscopic levels. The inner limiting membrane is covered by an extensive system of vitreal blood vessels emanating from the hyaloid artery, which enters the eye ventrally at the proximal end of the elongated optic nerve head. Two size classes of ganglion cell soma are segregated by optic axon fascicles and Müller cell endfeet, all of which lie at the level of the ganglion cell layer. A third class of 'displaced' ganglion cells lies at the border of the inner plexiform and inner nuclear layers amidst tightly packed amacrine, bipolar and Müller cell soma. Two layers of horizontal cells lie vitread of a synaptic zone consisting of a complex arrangement of horizontal and bipolar dendrites invaginating rod spherules and cone pedicles to form single and multiple (three to six) synaptic ribbon connections, respectively. Immediately vitread of the photoreceptor nuclei lie a population of 'displaced' bipolars. Three types of photoreceptors are characterised: unequal double cones, single cones (large and small) and rods. These show retinomotor movements where the rods elongate in the light and are masked by the pigment epithelium and contract in the dark as the pigment migrates sclerad. Ultrastructurally, 4 types of dark-staining (osmophilic) granules are described: (1) Small glycogen granules (0.033 microns) aggregated at the bases of the photoreceptor nuclei and larger similar granules (0.078 microns and termed paraboloids) vitread to the ellipsoid; (2) tapetal granules (0.32 microns) distributed throughout the dorsal four-fifths of the retinal pigment epithelium (RPE); (3) pigment granules (0.5-2.0 microns) in the RPE, concentrated in ventral retina; (4) granules or melanosomes (0.813 microns) of the choriocapillaris. The second class of granules constitute a tapetum lucidum eliciting a yellow eyeshine when viewed in the dark. Two other tapeta also exist, a guanine tapetum (irregular guanine crystals) and a tapetum fibrosum (stacks of collagen fibrils). Functional correlations are made, and the putative ancestral (primitive) condition of particular visual characters is established for the ray-finned fishes by out-group comparisons.