Search PubMed⌕ Search

Biomedical subjects

R R Colwell

Publications and source records attributed to R R Colwell.

At least 127 records · Page 7Linked to original sources
PubMed1990

Production of Escherichia coli-specific hybridomas by using gnotobiotic mice.

Monoclonal antibodies provide a rapid and specific means of direct detection of microorganisms in water and food samples. However, monoclonal antibodies specific for some bacteria are difficult to obtain; a good example of such a bacterium is Escherichia coli. Gnotobiotic BALB/c mice immunized with whole-cell preparations of heat-treated strains of E. coli and subjected to high-frequency antigen injection showed a significant increase in the number of specific hybridomas produced. Fusions obtained by using regular BALB/c mice immunized by using standard immunization protocols produced nonspecific hybridomas. Twenty-one stable hybridomas that did not cross-react with Klebsiella pneumoniae ATCC 13883 or Citrobacter freundii 1604770 were obtained from gnotobiotic mice. The bacterial strains were selected for the specificity tests because of their high cross-reactivity, which has been detected in previous fusion experiments. The method of immunization described here offers the potential of improving the production of highly specific hybridomas for bacteria which have been difficult to obtain.

Animals↗
PubMed1990

Particulate DNA in smoker fluids: evidence for existence of microbial populations in hot hydrothermal systems.

As part of an interdisciplinary study of hydrothermal vents on the Endeavour Segment of the Juan de Fuca Ridge, we used the submersible ALVIN to collect 57 fluid samples in titanium syringes and Go Flo Niskin bottles from 17 different hot vents (smokers and flanges) and their environs for the purpose of extracting particulate DNA. The relative purity of the vent fluids collected was determined by Mg content as an indicator of seawater entrainment. Particulate material concentrated from these samples was lysed enzymatically (enz) and by a combination of enzyme and French press treatment (fp). Concentrations of partially purified DNA recovered from these lysates were determined spectrofluorometrically by using the dye Hoechst 33258. Ambient seawater surrounding the vents was found to contain low DNA concentrations, 0.18 to 0.32 ng of DNA per ml (n = 4; mean(enz) = 0.23 +/- 0.05; mean(fp) = 0.26 +/- 0.05), while low-temperature vent samples yielded significantly higher concentrations of 0.37 to 2.12 ng of DNA per ml (n = 4; mean(enz) = 0.97 +/- 0.68; mean(fp) = 1.05 +/- 0.54). Although DNA recovery values from superheated (210 to 345 degrees C) flange samples (mean(enz) = 0.14 +/- 0.10; mean(fp) = 0.12 +/- 0.14) were not significantly different from ambient seawater values, most of the superheated (174 to 357 degrees C) smoker fluid samples contained particulate DNA in concentrations too high to be attributable to entrained seawater. Detailed sampling at one smoker site demonstrated not only the existence of significant levels of particulate DNA in the superheated smoker fluids but also the presence of an elevated microbial population in the buoyant plume 20 to 100 m above the smoker. These results underscore the heterogeneity of smoker environments within a given hydrothermal vent field and indicate that microorganisms exist in some superheated fluids.

Journal Article↗
PubMed1990

Hydrocarbon mineralization in sediments and plasmid incidence in sediment bacteria from the campeche bank.

Rates of degradation of radiolabeled hydrocarbons and incidence of bacterial plasmid DNA were investigated in sediment samples collected from the Campeche Bank, Gulf of Mexico, site of an offshore oil field containing several petroleum platforms. Overall rates of mineralization of [C]hexadecane and [C]phenanthrene measured for sediments were negligible; <1% of the substrate was converted to CO(2) in all cases. Low mineralization rates are ascribed to nutrient limitations and to lack of adaptation by microbial communities to hydrocarbon contaminants. Plasmid frequency data for sediment bacteria similarly showed no correlation with proximity to the oil field, but, instead, showed correlation with water column depth at each sampling site. Significant differences between sites were observed for proportion of isolates carrying single or multiple plasmids and mean number of plasmids per isolate, each of which increased as a function of depth.

Journal Article↗
PubMed1990

Phenotypic study of bacteria associated with the caribbean sclerosponge, Ceratoporella nicholsoni.

Heterotrophic bacteria associated with the Caribbean sclerosponge, Ceratoporella nicholsoni (Hickson), were found to occur extracellularly and were confined to the mesohyl regions of the sponge tissue. Physiological, metabolic, and morphological attributes of the culturable bacteria associated with the sponge were recorded by using numerical taxonomy methods for the analysis of 158 phenotypic attributes. Morphometric methods were used to determine the proportion of the total sponge-associated bacteria that were culturable by the methods employed, with the results ranging from 3 to 11% of the total bacteria inhabiting the sponge. Approximately 78% of the culturable bacteria clustered into four groups or phena, representing two previously undescribed Vibrio spp., an Aeromonas sp., and a coryneform- or actinomycete-like sp. Most of the bacteria were facultative anaerobes, fermenting sucrose and fucose but unusual in an inability to ferment glucose. This study was the first comprehensive study of heterotrophic bacteria associated with a sponge from the Caribbean basin, a region reputed to contain the most prolific sponge populations, with respect to biomass and diversity. The possible significance of these associations is discussed.

Animals↗
PubMed1990

Attachment of Vibrio cholerae serogroup O1 to zooplankton and phytoplankton of Bangladesh waters.

Vibrio cholerae serogroup O1, the causative agent of cholera, is capable of surviving in aquatic environments for extended periods and is considered an autochthonous species in estuarine and brackish waters. These environments contain numerous elements that may affect its ecology. The studies reported here examined physical interactions between V. cholerae O1 and natural plankton populations of a geographical region in Bangladesh where cholera is an endemic disease. Results showed that four of five clinical V. cholerae O1 strains and endogenous bacterial flora were attached preferentially to zooplankton molts (exuviae) rather than to whole specimens. One strain attached in approximately equal numbers to both exuviae and whole specimens. V. cholerae O1 also attached to several phytoplankton species. The results show that V. cholerae O1 can bind to diverse plankton species collected from an area where cholera is an endemic disease, with potentially significant effects on its ecology.

Animals↗
PubMed1990

Maintenance of plasmids pBR322 and pUC8 in nonculturable Escherichia coli in the marine environment.

Maintenance of plasmids pBR322 and pUC8 in Escherichia coli that was nonculturable after exposure to seawater was studied. E. coli JM83 and JM101, which contained plasmids pBR322 and pUC8, respectively, were placed in sterile artificial seawater for 21 days. Culturability was determined by plating on both nonselective and selective agar, and plasmid maintenance was monitored by direct isolation of plasmid nucleic acid from bacteria collected on Sterivex filters. E. coli JM83 became nonculturable after incubation for 6 days in seawater yet maintained plasmid pBR322 for the entire period of the study, i.e., 21 days. E. coli JM101 was nonculturable after incubation in seawater for 21 days and also maintained plasmid pUC8 throughout the duration of the microcosm experiment. Direct counts of bacterial cells did not change significantly during exposure to seawater, even though plate counts yielded no viable (i.e., platable) cells. We concluded that E. coli cells are capable of maintaining high-copy-number plasmids, even when no longer culturable, after exposure to the estuarine or marine environment.

DNA, Bacterial↗
PubMed1990

Detection of Vibrio cholerae O1 in the aquatic environment by fluorescent-monoclonal antibody and culture methods.

Vibrio cholerae O1 in plankton samples collected from ponds and rivers between February 1987 and January 1990 in Matlab, Bangladesh, was detected by the fluorescent-monoclonal antibody (FA) technique. Samples were collected at sites which were monitored fortnightly (fixed sites) as well as at sites that were part of a case-control study. FA results were compared with those obtained by conventional culture methods (CM). A total of 876 samples were collected; V. cholerae O1 was detected in 563 samples (64.27%) by the FA method and in 3 samples (0.34%) by CM. Of the fixed-site plankton samples, 439 (63.62%) were positive by FA and none were positive by CM. Of the 93 case sites sampled on the day after the occurrence of a case of cholera, 73 (78.49%) were positive for V. cholerae O1 by FA and 3 (3.2%) were positive by CM. In comparison, of the 93 first-day sample collections at control sites at the time a case of cholera occurred, only 51 (54.83%) were positive by FA and none were positive by CM. From the data, it is concluded that V. cholerae O1 is present throughout the year in the ponds and rivers of Bangladesh that were examined in this study and that V. cholerae can be detected by FA but not always by CM. The FA procedure was found to be very useful in detecting V. cholerae in plankton, with which it was associated and often occurred in large numbers in the nonculturable stage. Thus, studies investigating the significance of the role of environmental factors in the epidemiology of cholera can be performed effectively by using FA. Such studies are in progress.

Animals↗
PubMed1990

Microbial degradation of hydrocarbons in the environment.

The ecology of hydrocarbon degradation by microbial populations in the natural environment is reviewed, emphasizing the physical, chemical, and biological factors that contribute to the biodegradation of petroleum and individual hydrocarbons. Rates of biodegradation depend greatly on the composition, state, and concentration of the oil or hydrocarbons, with dispersion and emulsification enhancing rates in aquatic systems and absorption by soil particulates being the key feature of terrestrial ecosystems. Temperature and oxygen and nutrient concentrations are important variables in both types of environments. Salinity and pressure may also affect biodegradation rates in some aquatic environments, and moisture and pH may limit biodegradation in soils. Hydrocarbons are degraded primarily by bacteria and fungi. Adaptation by prior exposure of microbial communities to hydrocarbons increases hydrocarbon degradation rates. Adaptation is brought about by selective enrichment of hydrocarbon-utilizing microorganisms and amplification of the pool of hydrocarbon-catabolizing genes. The latter phenomenon can now be monitored through the use of DNA probes. Increases in plasmid frequency may also be associated with genetic adaptation. Seeding to accelerate rates of biodegradation has been shown to be effective in some cases, particularly when used under controlled conditions, such as in fermentors or chemostats.

Bacteria↗
PubMed1990

Variation in epitopes of the B subunit of Vibrio cholerae non-O1 and Vibrio mimicus cholera toxins.

Monoclonal antibodies reacting with the B subunit of Vibrio cholerae O1 strain 569B cholera toxin (CT-B) were used to identify unique and common epitopes of V. cholerae non-O1 and Vibrio mimicus CT-B. Vibrio cholerae non-O1 strains produced CT-B showing three monoclonal antibody reaction patterns (epitypes), which corresponded with epitypes described previously for V. cholerae O1 classical biotype CT-B (CT1), El Tor biotype CT-B (CT2), and a unique V. cholerae non-O1 CT-B (CT3), which lacked an epitope located in or near the GM1 ganglioside binding site of 569B CT-B. Vibrio mimicus CT-B was immunologically indistinguishable from 569B CT-B. These and previous results define six epitopes on 569B CT-B, and a fourth epitope in or near the GM1 ganglioside binding site.

Antibodies, Monoclonal↗
PubMed1989

Variation in epitopes of the B subunit of El Tor and classical biotype Vibrio cholerae O1 cholera toxin.

Variation in epitopes of the B subunit of cholera toxin (CT-B) produced by strains of El Tor and classical biotype Vibrio cholerae O1 was examined using monoclonal antibodies prepared to V. cholerae 569B CT. CT-B epitopes were markedly conserved for V. cholerae classical biotypes. In contrast, epitope variation was observed for El Tor biotypes, which produced both a classical-like CT-B and a unique CT-B lacking at least one epitope common to 569B CT-B. The missing epitope was located outside the GM1 ganglioside-binding site. From results of the study reported here, genetic divergence is exhibited in the El Tor biotype CT-B versus classical CT-B. Furthermore, at least five unique epitopes of V. cholerae 569B CT-B can be defined.

Antibodies, Bacterial↗
PubMed1989

Simple, rapid method for direct isolation of nucleic acids from aquatic environments.

Direct isolation of nucleic acids from the environment may be useful in several respects, including the estimation of total biomass, detection of specific organisms and genes, estimations of species diversity, and cloning applications. We have developed a method that facilitates the concentration of microorganisms from aquatic samples and the extraction of their nucleic acids. Natural water samples of 350 to greater than 1,000 ml are concentrated on a single cylindrical filter membrane (type SVGS01015; Millipore Corp., Bedford, Mass.), and cell lysis and proteolysis are carried out within the filter housing. Crude, high-molecular-weight nucleic acid solutions are then drawn off the filter. These solutions can be immediately analyzed, concentrated, or purified, depending on the intended application. The method is simple, rapid, and economical and provides high-molecular-weight chromosomal DNA, plasmid DNA, and speciated RNAs which comigrate with 5S, 16S, and 23S rRNAs. The methods presented here should prove useful in studying both the ecology and the phylogeny of microbes that resist classical culture methods.

Acetates↗
PubMed1988

A tissue culture assay for tetrodotoxin, saxitoxin and related toxins.

In the presence of ouabain, veratridine enhances sodium influx in the mouse neuroblastoma cell line Neuro-2A (ATCC, CCL131), causing cellular swelling and subsequent death. Tetrodotoxin (puffer fish toxin) or saxitoxin (paralytic shellfish poison), both of which block the sodium channel of excitable membranes, antagonize this effect, enabling cell growth to continue. This phenomenon was used as the basis of a new assay for these toxins. It is also possible to estimate the quantity of TTX from the relationship between TTX concentration and percentage of living cells. This new method is simple, inexpensive, and sensitive, and may replace the conventional mouse bioassay.

Animals↗
PubMed1988

Enzymatic characterization of three aeromonas species using API Peptidase, API "Osidase," and API Esterase test kits.

An enzymatic characterization of 16 strains of Aeromonas species including A. hydrophila (7), A. sobria (5), and A. caviae (4) was carried out using API Peptidase (strips numbered 1, 2, 3, 4, 5, and 6); API Esterase and API "Osidase" test strips. A total of 89 substrates was used in the assay and included 59 arylamides (aminopeptides), 10 esters, and 20 carbohydrates. All three species were remarkably uniform in their reactivities. Nineteen (32%) of the arylamide substrates used were hydrolyzed by all three species. Very strong arylamidase activity was displayed by all three species for L-lysine, L-hydroxyproline, L-arginine, L-alanine, L-proline, and L-leucyl-L-alanine. Esterase activity was strongest against caproate (C6), caprylate (C8), nonanoate (C9), and caprate (C10) substrates. Only a limited number of carbohydrate substrates were hydrolyzed; strong N-acetyl-beta-D-glucosaminidase activity was given by all strains. Both A. hydrophila and A. caviae gave strong beta-D-glucosidase reactivities, while A. sobria appeared to be negative for this enzyme. The results of our preliminary study show that some of the enzymes examined may be useful in the identification and differentiation of these species. The API enzyme assays yielded rapid (4 hr) results. The assays were easy to perform, relatively inexpensive and reproducible. The importance of replicate testing and the inclusion of uninoculated (buffer only) controls as part of the assay is emphasized.

Aeromonas↗
PubMed1988

Comparative analysis of the chemical composition of mixed and pure cultures of green algae and their decomposed residues by C nuclear magnetic resonance spectroscopy.

It is known that macromolecular organic matter in aquatic environments, i.e., humic substances, is highly aliphatic. These aliphatic macromolecules, predominantly paraffinic in structure, are prevalent in marine and lacustrine sediments and are believed to originate from algae or bacteria. A comparative study of mixed and pure cultures of green algae and their decomposed residues was performed by using solid-state C nuclear magnetic resonance spectroscopy as the primary analytical method. Results obtained in this study confirm the presence of components that are chemically refractory and that are defined as alghumin and hydrolyzed alghumin. These were detected in heterogeneous, homogeneous, and axenic biomasses composed of several genera of Chlorophyta. Although the chemical composition of algal biomass varied with culture conditions, the chemical structure of the alghumin and hydrolyzed alghumin, demonstrated by C nuclear magnetic resonance spectroscopy appeared to be constant for members of the Chlorophyta examined in this study. The alghumin was dominated by carbohydrate-carbon, with minor amounts of amide or carboxyl carbon and paraffinic carbon, the latter surviving strong hydrolysis by 6 N HCI (hydrolyzed alghumin). Bacterial decomposition of heterogeneous algal biomass labeled with C was conducted under both aerobic and anaerobic conditions to determine chemical structure and stability of the refractory material. The refractory fraction ranged from 33% in aerobic to 44% in anaerobic cultures. The refractory fraction recovered from either aerobic or anaerobic degradation comprised 40% alghumin, which represented an enrichment by 10% relative to the proportion of alghumin derived from whole cells of algae. The paraffinic component in the hydrolyzed alghumin of whole algal cells was found to be 1.8% and increased to 5.1 and 6.9% after aerobic and anaerobic bacterial degradation, respectively. It is concluded that members of the Chlorophyta contain a common insoluble structure composed of paraffinic carbon that is resistant to chemical and bacterial degradation under conditions used in this study. The paraffinic structure is identical to those constituting humin of aquatic origin. Thus, alga-derived macromolecular compounds deposited in aquatic environments (alghumin) probably contribute to sedimentary humic substances.

Journal Article↗
PubMed1988

Frequency and characteristics of plasmids in bacteria isolated from deep-sea amphipods.

Bacterial strains isolated from deep-sea amphipods were identified, classified, and screened for plasmid content. Plasmids were common, with 11 of 16 isolates carrying one or more plasmids; these ranged in size from 2.9 to 63 megadaltons. Several of the strains demonstrated distinctly different phenotypic traits yet contained plasmids of the same molecular weight. Results of agarose gel electrophoresis, DNA hybridization, and restriction analysis indicate that the plasmids detected in these deep-sea isolates are identical, suggesting that transmission may occur in the deep-sea environment and that plasmids are common in some deep-sea habitats.

Journal Article↗