Biomedical subjects

M P Shiaris

Publications and source records attributed to M P Shiaris.

13 recordsLinked to original sources

PubMed1998

Spatial and temporal variation of phenanthrene-degrading bacteria in intertidal sediments.

Phenanthrene-degrading bacteria were isolated from a 1-m2 intertidal sediment site in Boston Harbor. Samples were taken six times over 2 years. A total of 432 bacteria were isolated and characterized by biochemical testing. When clustered on the basis of phenotypic characteristics, the isolates could be separated into 68 groups at a similarity level of approximately 70%. Several groups (a total of 200 isolates) corresponded to well-characterized species belonging the genera Vibrio and Pseudomonas. Only 51 of the 437 isolates (< 11.7% of the total) hybridized to a DNA probe that encodes the upper pathway of naphthalene and phenanthrene degradation in Pseudomonas putida NCIB 9816. A cluster analysis indicated that the species composition of the phenanthrene-degrading community changed significantly from sampling date to sampling date. At one sampling time, 12 6-mm-diameter core subsamples were taken within the 1-m2 site to determine the spatial variability of the degrading communities. An analysis of molecular variance, performed with the phenotypic characteristics, indicated that only 6% of the variation occurred among the 12 subsamples, suggesting that the subsamples were almost identical in composition. We concluded that the communities of phenanthrene-degrading bacteria in the sediments are very diverse, that the community structure undergoes significant change with time but does not vary significantly on a spatial scale of centimeters, and that the predominant genes that encode phenanthrene degradation in the communities are not well-characterized.

Geologic Sediments↗

PubMed1994

Relative role of eukaryotic and prokaryotic microorganisms in phenanthrene transformation in coastal sediments.

THE RELATIVE ROLE OF EUKARYOTIC VERSUS PROKARYOTIC MICROORGANISMS IN PHENANTHRENE TRANSFORMATION WAS MEASURED IN SLURRIES OF COASTAL SEDIMENT BY TWO DIFFERENT APPROACHES: detection of marker metabolites and use of selective inhibitors on phenanthrene biotransformation. Phenanthrene biotransformation was measured by polar metabolite formation and CO(2) evolution from [9-C]phenanthrene. Radiolabeled metabolites were tentatively identified by high-performance liquid chromatography (HPLC) separation combined with UV/visible spectral analysis of HPLC peaks and comparison to authentic standards. Both yeasts and bacteria transformed phenanthrene in slurries of coastal sediment. Two products of phenanthrene oxidation by fungi, phenanthrene trans-3,4-dihydrodiol and 3-phenanthrol, were produced in yeast-inoculated sterile sediment. However, only products of phenanthrene oxidation typical of bacterial transformation, 1-hydroxy-2-naphthoic acid and phenanthrene cis-3,4-dihydrodiol, were isolated from slurries of coastal sediment with natural microbial populations. Phenanthrene trans-dihydrodiols or other products of fungal oxidation of phenanthrene were not detected in the slurry containing a natural microbial population. A predominant role for bacterial transformation of phenanthrene was also suggested from selective inhibitor experiments. Addition of streptomycin to slurries, at a concentration which suppressed bacterial viable counts and rates of [methyl-H]thymidine uptake, completely inhibited phenanthrene transformation. Treatment with colchicine, at a concentration which suppressed yeast viable counts, depressed phenanthrene transformation by 40%, and this was likely due to nontarget inhibition of bacterial activity. The relative contribution of eukaryotic microorganisms to phenanthrene transformation in inoculated sterile sediment was estimated to be less than 3% of the total activity. We conclude that the predominant degraders of phenanthrene in muddy coastal sediments are bacteria and not eukaryotic microorganisms.

Journal Article↗

PubMed1994

Metabolism of naphthalene, fluorene, and phenanthrene: preliminary characterization of a cloned gene cluster from Pseudomonas putida NCIB 9816.

A modified cloning procedure was used to obtain large DNA insertions (20 to 30 kb) from Pseudomonas putida NCIB 9816 that expressed polycyclic aromatic hydrocarbon (PAH) transformation activity in Escherichia coli HB101. Four subclones (16 [in both orientations], 12, and 8.5 kb in size) were constructed from the initial clones. Naphthalene, fluorene, and phenanthrene transformations were investigated in these eight NCIB 9816 clones by a simple agar plate assay method, which was developed to detect and identify potential PAH metabolites. Results indicated that the necessary genes encoding the initial ring fission of the three PAHs in E. coli cells are located in an 8.5-kb EcoRI-XhoI portion, but the lower-pathway genes are not present in a 38-kb neighborhood region. These NCIB 9816 clones could transform naphthalene and phenanthrene to salicylic acid and 1-hydroxy-2-naphthoic acid, respectively. With the same clones, fluorene was degraded to 9-hydroxyfluorene, 9-fluorenone, and two unidentified compounds. Genetic similarity between the NAH7 upper-pathway genes and the cloned NCIB 9816 genes was confirmed by Southern blot DNA-DNA hybridization. In spite of this genetic similarity, the abilities of the two clusters to transform multiple PAHs were different. Under our experimental conditions, only the metabolites from naphthalene transformation by the NAH7 clone (pE317) were detected, whereas the NCIB 9816 clones produced metabolites from all three PAHs.

Biotransformation↗

PubMed1993

Biotransformation of polycyclic aromatic hydrocarbons by yeasts isolated from coastal sediments.

Yeast abundance in the sediments of 13 coastal sites in Massachusetts was quantified, and the potential of yeast isolates to biotransform polycyclic aromatic hydrocarbons (PAHs) was determined. Plate counts of yeasts varied between 10(2) to 10(7) CFU g (dry weight) of sediment-1. The most abundant genera isolated and identified included Candida, Cryptococcus, Rhodotorula, Torulopsis, and Trichosporon. More than 50% of the isolates from heavily contaminated sites transformed phenanthrene, as determined by spray-plate screening. The plate counts of phenanthrene-transforming yeasts correlated significantly to the sediment concentrations of phenanthrene. Transformation of [9-14C]phenanthrene and [12-14C]benz[a]anthracene by individual isolates varied greatly, ranging from 0.15 to 8.15 mumol of PAH g-1 in 120-h incubations. Of the isolated yeasts, Trichosporon penicillatum exhibited the greatest capacity for phenanthrene transformation. The ability to transform PAHs appears to be widespread among yeasts in coastal sediments.

Benz(a)Anthracenes↗

PubMed1992

Rates of microbenthic and meiobenthic bacterivory in a temperate muddy tidal flat community.

Rates of bacterivory in micro- and meiobenthic species were determined by an improved technique in a muddy tidal flat community in Boston Harbor, Mass. The predominant grazers of bacteria were identified, and their rates of grazing were measured in the top 1 cm of the sediment. Grazing rates were measured by a fluorescence-labeled bacteria (FLB) technique. A mixture of two Enterococcus spp. isolates and two isolates of Escherichia coli were prepared as FLB, and they were added to intact sediment cores by replacing the pore water in the upper centimeter of the core. A standard FLB procedure was modified by filtering sediment dilutions onto cellulose membrane filters and processing the filters to render them optically transparent while preserving the physical integrity of the micro- and meiobenthic organisms. Thus, it was possible, on the same microscopic field, to switch from light microscopy for identification of grazers to epifluorescence microscopy for counting FLB present in the gut contents of the same grazers. The majority of benthic organisms present in these sediments consumed FLB, but their consumption rates varied widely. Two ciliate species, a Prorodon sp. and a Chlamidodon sp., and a nematode, a Metoncholaimus sp., consumed fluorescence-labeled coliforms at the highest rates, 126 to 169 FLB per individual per h. Other ciliates and nematodes, as well as microflagellates and harpacticoid copepods, consumed fluorescence-labeled coliforms at lower rates, 1.2 to 26 FLB per individual per h. Foraminiferans and gastrotriches did not contain FLB. Some ciliate grazers discriminated between enterococci and coliforms, consuming the rod-shaped fluorescence-labeled coliforms at 74- to 155-fold-higher rates than did the coccus-shaped fluorescence-labeled enterococci. Other ciliates did not select between fluorescence-labeled enterococci and fluorescence-labeled coliforms. The high rates of bacterivory by some ciliates and nematodes indicated intensive grazing. However, at their low extant densities, the grazers consumed only a small portion of the bacterial standing stock. Major bacterial grazers, e.g., microflagellates, ciliates, and nematodes, could potentially consume, per day, only 0.2, 0.1, and 0.03%, respectively, of the bacterial standing stock (7.5 x 10 bacteria per cm).

Journal Article↗

PubMed1989

Seasonal Biotransformation of Naphthalene, Phenanthrene, and Benzo[a]pyrene in Surficial Estuarine Sediments.

Transformation rates of naphthalene, phenanthrene, and benzo[a]pyrene in oxidized surficial sediments of a polluted urban estuary, Boston Harbor, Mass., were determined over a period of 15 months. Three sites characterized by muddy sediments were selected to represent a >300-fold range of ambient polycyclic aromatic hydrocarbon (PAH) concentration. Transformation rates were determined by a trace-level radiolabel PAH assay which accounted for PAH mineralization, the formation of polar metabolites, residue, and recovered parental PAHs in sediment slurries. Transformation rates of the model PAHs increased with increasing ambient PAH concentrations. However, turnover times for a given PAH were similar at all sites. The turnover times were as follows: naphthalene, 13.2 to 20.1 days; phenanthrene, 7.9 to 19.8 days, and benzo[a]pyrene, 53.7 to 82.3 days. At specific sites, rates were significantly affected by salinity, occasionally affected by temperature, but not affected by pH over the course of the study. Seasonal patterns of mineralization were observed for each of the PAHs at all sites. The timing of seasonal maxima of PAH mineralization varied from site to site. Seasonal potential heterotrophic activities as measured by acetate and glutamate mineralization rates did not always coincide with PAH mineralization maxima and minima, suggesting that the two processes are uncoupled in estuarine sediments.

Journal Article↗

PubMed1987

Comparative survival of antibiotic-resistant and -sensitive fecal indicator bacteria in estuarine water.

The survival of antibiotic-resistant and -sensitive strains of Escherichia coli, Enterococcus faecalis, Enterococcus faecium, Streptococcus equinus, and two environmental isolates, AP17 and AQ62, was examined in estuarine water. Each strain was rendered resistant to a combination of two antibiotics by serial passage in increasing concentrations of antibiotics. Cultures were incubated in filter-sterilized estuarine water for up to 7 days. Recovery was assessed by examining colony-forming ability on media with and without antibiotics. None of the antibiotic-resistant forms survived longer than its antibiotic-sensitive counterpart in estuarine water. Three of the resistant strains died off more rapidly than the antibiotic-sensitive wild type. Survival of the test bacteria in estuarine water was as follows: sensitive and resistant AQ62, resistant Escherichia coli less than sensitive Escherichia coli less than resistant AP17 less than resistant Enterococcus faecium less than sensitive AP17, sensitive and resistant S. equinus less than sensitive and resistant Enterococcus faecalis, sensitive Enterococcus faecium. The results supported the suggestion that fecal entercocci may serve as better indicators of fecal pollution than Escherichia coli in marine ecosystems. Moreover, the results indicated that the use of antibiotic-resistant mutants to follow the fate of bacteria in the environment is inappropriate without adequate studies to ensure that resistant and wild-type strains react similarly to environmental stressors.

Anti-Bacterial Agents↗

PubMed1987

Distribution of indicator bacteria and Vibrio parahaemolyticus in sewage-polluted intertidal sediments.

The impact of a sewage point source on the bacterial densities in an intertidal mud flat in Boston Harbor, Mass., was investigated. The area, Savin Hill Cove, acts as a receiving basin for a combined storm and sewage outlet (CSO). Preliminary examination of sediments and overlying water at high tide demonstrated that fecal coliforms were present in sediments at abundances 2 to 4 orders of magnitude higher than in the overlying water column. The following bacterial counts were determined from sediments along a sampling transect extending 460 m from the CSO: total bacteria by epifluorescent microscopy, heterotrophic bacteria by plate counts on nutrient-rich and nutrient-poor media, fecal coliforms and enterococci by membrane filtration, and Vibrio parahaemolyticus by a most-probable-number technique with a resuscitation step. Median sediment grain size, average tidal exposure, carbon/nitrogen ratio, and total organic carbon were also measured. All bacterial indices, except for V. parahaemolyticus, declined significantly with distance from the outfall. Multiple regression analysis indicated that tidal exposure (low tides) may affect densities of total bacteria. Fecal coliforms and enterococci were still present in appreciable numbers in sediments as far as 460 m away from the CSO. In contrast, V. parahaemolyticus densities did not correlate with the other bacterial counts nor with any of the environmental parameters examined. These results indicate that intertidal sediments which adjoin point sources of pollution are severely contaminated and should be considered as potentially hazardous reservoirs of sewage-borne diseases.

Enterobacteriaceae↗

PubMed1983

Replica plating method for estimating phenanthrene-utilizing and phenanthrene-cometabolizing microorganisms.

A replica plating method was developed for detecting and enumerating phenanthrene-degrading microorganisms. The method is designed to discriminate between aquatic organisms that utilize phenanthrene as the sole carbon and energy source and organisms that cometabolize phenanthrene. The method was used to demonstrate that phenanthrene utilizers and phenanthrene cometabolizers coexist in estuarine sediments.

Journal Article↗

PubMed1982

Effects of polychlorinated biphenyls and environmental biotransformation products on aquatic nitrification.

The effects of polychlorinated biphenyls (PCBs) on nitrification were examined for pure cultures and natural reservoir samples. PCBs at concentrations greater than 10 microgram liter-1 inhibited nitrification, principally ammonium oxidation, in one of two natural reservoir environments. However, this inhibition could not be reproduced in pure high-cell-density cultures or in previously contaminated reservoir waters. A PCB environmental biotransformation product, p-chlorophenylglyoxylic acid, and p-chloromandelic acid had no effect on nitrification.

Biotransformation↗

PubMed1982

Impact of coal-coking effluent on sediment microbial communities: a multivariate approach.

The functional response to and recovery from coal-coking waste effluent was evaluated for sediment microbial communities. Twenty estimates of microbial population density, biomass, and activity were measured five times during a 15-month period. Significant effects on microbial communities were observed in response to both wastewater contamination and diversion of the wastewater. Multivariate analysis of variance and discriminant analysis indicated that accurate differentiation between uncontaminated and contaminated sediments required a minimum of nine estimates of community response. Total viable population density, ATP, alkaline phosphatase, naphthalene, and phenanthrene mineralization rates were found to be highly weighted variables in site discrimination. Lipid and glucose mineralization, nitrogen fixation, and sediment protein also contributed significantly to explaining variation among sites. Estimates of anaerobic population densities and rates of methane production contributed little to discrimination among sites in the environment examined. In general, total viable population density, ATP, and alkaline phosphatase activity were significantly depressed in contaminated sediments. However, after removal of this contamination, the previously affected sites demonstrated greater temporal variability but a closer approximation of the mean response at the control site. Naphthalene and phenanthrene mineralization did not follow the general trend and were elevated at the contaminated sites throughout the investigation. Results of the investigation supported the hypothesis that multiple functional measures of microbial community response are required to evaluate the effect of and recovery from environmental contamination. In addition, when long-term effects are evaluated, select physiological traits, i.e., polyaromatic hydrocarbon mineralization, may not reflect population and biomass estimates of community response.

Journal Article↗

PubMed1980

Tenax-GC Extraction Technique for Residual Polychlorinated Biphenyl and Polyaromatic Hydrocarbon Analysis in Biodegradation Assays.

A rapid Tenax-GC extraction technique has been evaluated for use in conjunction with aqueous biodegradation assays for polyaromatic hydrocarbons and polychlorinated biphenyls. The method was quantitatively efficient and reproducible for phenanthrene, but variable and not quantitative for Aroclor 1254 (polychlorinated biphenyls). Aqueous sample volumes and varying concentrations of organic matter influenced polychlorinated biphenyl and polyaromatic hydrocarbon extraction efficiency. Phenanthrene recovery was decreased by soil extract but unaffected by spent bacteriological culture medium. Both types of organic matter caused significant reduction of Aroclor 1254 recovery. Polyaromatic hydrocarbon and polychlorinated biphenyl biodegradation assays, performed with reservoir samples, supported the laboratory evaluation. The study demonstrated the utility of the Tenax-GC extraction technique for phenanthrene analysis in biodegradation assessment; however, Tenax-GC extraction was not appropriate for Aroclor 1254 biodegradation studies.

Journal Article↗

PubMed1979

Comparative effects of Aroclor 1254 (polychlorinated biphenyls) and phenanthrene on glucose uptake by freshwater microbial populations.

The effects of polychlorinated biphenyl (PCB) and phenanthrene stress on glucose uptake by natural microbial populations were examined by the heterotrophic potential technique. Temporal and spatial distributions in glucose uptake velocities were examined for natural samples as well as PCB- and phenanthrene-stressed samples. Statistical analysis indicated significant variability among the various samples. It was demonstrated that the environmental variables contributed significantly to the variability in uptake kinetics. Although general trends indicated a PCB-induced stimulation in uptake velocities, these trends were in part masked by sample variability. Data analysis indicated no statistically significant PCB or phenanthrene effect on either total glucose uptake velocities or the proportion of 14CO2 evolved, as compared to natural unstressed samples.

Aroclors↗