Can Dietary Fatty Acids Affect the COVID-19 Infection Outcome in Vulnerable Populations?

There is high mortality in coronavirus disease 2019 (COVID-19)-infected individuals with chronic inflammatory diseases, like obesity, diabetes, and hypertension. A cytokine storm in some patients after infection contributes to this mortality. In addition to lungs, the intestine is targeted during COVID-19 infection. The intestinal membrane serves as a barrier to prevent leakage of microorganisms and their products into the bloodstream; however, dietary fats can affect the gut microbiome and may increase intestinal permeability. In obese or diabetic individuals, there is an increase in the abundance of either Gram-negative bacteria in the gut or their product, endotoxin, in systemic circulation. We speculate that when the COVID-19 infection localizes in the intestine and when the permeability properties of the intestinal membrane are compromised, an inflammatory response is generated when proinflammatory endotoxin, produced by resident Gram-negative bacteria, leaks into the systemic circulation. This review discusses conditions contributing to inflammation that are triggered by microbially derived factors from the gut.

Host genotype and exercise exhibit species-level selection for members of the gut bacterial communities in the mouse digestive system.

The mammalian gut microbiome can potentially impact host health and disease state. It is known that the mouse-genome, eating-behavior, and exercise-status promotes higher taxonomic rank-level alterations (e.g. family to phyla-level) of the gut microbiota. Here, host genotype or activity status was investigated to determine if selection of individual bacterial species or strains could be discerned within the murine digestive system. For this study, the fecal bacterial community of adenylyl cyclase 5 knock-out (AC5KO, n = 7) mice or their wild-type (WT, n = 10) littermates under exercise or sedentary conditions were profiled by sequencing rRNA operons. AC5KO mice were chosen since this genotype displays enhanced longevity/exercise capacity and protects against cardiovascular/metabolic disease. Profiling of rRNA operons using the Oxford MinION yielded 65,706 2-D sequences (after size selection of 3.7-5.7 kb) which were screened against an NCBI 16S rRNA gene database. These sequences were binned into 1,566 different best BLAST hits (BBHs) and counted for each mouse sample. Non-metric multidimensional scaling (NMDS) of the gut microbial community demonstrated clustering by physical activity (p = 0.001) but not by host genotype. Additionally, sequence similarity and phylogenetic analysis demonstrated that different bacterial species (closely related to Muribaculum intestinale and Parasutterella excrementihominis) inhabit AC5KO or WT mice depending on activity status. Other bacterial species of the gut microbiota did not follow such patterning (e.g. Turicibacter sanguinis and Turicimonas muris). Our results support the need of improved taxonomic resolution for better characterization of bacterial communities to deepen our understanding of the role of the gut microbiome on host health.

Effect of temperature on the reductive dechlorination of 1,2,3,4-tetrachlorodibenzofuran in anaerobic PCDD/F-contaminated sediments.

The effect of temperature on the reductive dechlorination in sediments of the PCDD/F-contaminated Kymijoki River, Finland was assessed with 1,2,3,4-tetrachlorodibenzofuran (1,2,3,4-TeCDF) at various temperatures and with co-amendment of 2,3,4,6-tetrachlorophenol (2,3,4,6-TeCP) in laboratory microcosms. The dechlorination rate of 1,2,3,4-TeCDF increased with incubation temperature, with TeCDF half-lives of 2.1 y at 21°C, 3.9 y at 15°C, and 19.0 y at 4°C. Co-amendment with 2,3,4,6-TeCP reduced the TeCDF half-life to 1.8 y at 21°C. 1,2,3,4-TeCDF was dechlorinated mainly in the lateral position to 1,3,4-TrCDF and then to 1,3-DiCDF over 29 months, but incubation temperature affected the relative molar ratios of the dechlorination products. The abundance of the Dehalococcoides-like Chloroflexi community did not substantially change in microcosms over 24 months incubation at the different temperatures. The dechlorination activity of 1,2,3,4-TeCDF was significantly limited at lower temperatures, which should be considered in predicting the environmental fate of aged PCDD/Fs in sediments of the Kymijoki River.

Anaerobic reductive dechlorination of 1,2,3,4-tetrachlorodibenzofuran in polychlorinated dibenzo-p-dioxin- and dibenzofuran-contaminated sediments of the Kymijoki River, Finland.

Sediments of the Kymijoki River are highly contaminated with polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs). These persistent PCDD/Fs resist biotic degradation and therefore the potential for microbial reductive dechlorination was assessed to determine how microbes impact the fate of these compounds. Anaerobic sediment microcosms of five different sites in the river were spiked with 1,2,3,4-tetrachlorodibenzofuran (1,2,3,4-TeCDF) as a model compound to determine the dechlorination potential in the sediments. Dechlorinating bacteria were active in all the study sites of the river. The extent of dechlorination over 10 and 29 months corresponded to the levels of aged PCDD/Fs, with sediments of the most contaminated site at Kuusankoski being the most active for reductive dechlorination. The dechlorination activity and levels of aged PCDD/Fs were correlated within the sediment cores at the all sites. The pathway of 1,2,3,4-TeCDF dechlorination was mainly via 1,3,4-trichlorodibenzofuran (TrCDF) to 1,3-dichlorodibenzofuran (DiCDF). Dechlorination via 1,2,4-TrCDF to further dechlorination products was also detected. Lateral reductive dechlorination would decrease the toxicity of 2,3,7,8-substituted PCDD/Fs. Our data suggest that sediments of the Kymijoki River contain indigenous microorganisms that are responsible for dechlorination of PCDD/Fs, especially at the most contaminated site.

Impact of temperature, CO2 fixation and nitrate reduction on selenium reduction, by a paddy soil Clostridium strain.

AIMS: To elucidate the impact of CO(2) fixation, nitrate reduction and temperature on selenium reduction by a newly identified acetogenic bacterium, Clostridium sp. BXM. METHODS AND RESULTS: A series of culture experiments were designed to evaluate the impact of temperature, CO(2) fixation and nitrate reduction on the rate and extent of selenium reduction by strain BXM. The products of selenium reduction, CO(2) fixation and nitrate reduction were determined. Molecular analysis was performed to identify the functional genes involved in the selenium reduction process. CO(2) may have enhanced the activity of hydrogenase I and/or the level of cytochrome b, thus increasing selenium reduction. Nitrate may inhibit selenium reduction due to its higher reduction potential and/or by decreasing selenite/selenate reductase activity. The suitable temperature was 37 and 30 °C for selenite reduction under anaerobic and aerobic conditions, respectively. The optimum temperature was 30 °C for selenate reduction under both anaerobic and aerobic conditions. CO(2) fixation and nitrate reduction by Clostridium sp. BXM stimulated each other. CONCLUSIONS: Clostridium sp. BXM was capable of reducing up to 36-94% of 1 mmol l(-1) selenate and selenite under anaerobic or aerobic conditions over 15 days. The strain might be used for the precipitation of Se from highly selenium-contaminated water or sediments. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings contribute to the current understanding about the role that micro-organisms play in the detoxification of toxic selenium compounds in paddy soils. Micro-organisms in paddy soils can influence selenium accumulation in rice grain and hence human selenium intake.

Characterization of the indigenous PAH-degrading bacteria of Spartina dominated salt marshes in the New York/New Jersey Harbor.

The aerobic polyaromatic hydrocarbon (PAH) degrading microbial communities of two petroleum-impacted Spartina-dominated salt marshes in the New York/New Jersey Harbor were examined using a combination of microbiological, molecular and chemical techniques. Microbial isolation studies resulted in the identification of 48 aromatic hydrocarbon-degrading bacterial strains from both vegetated and non-vegetated marsh sediments. The majority of the isolates were from the genera Paenibacillus and Pseudomonas. Radiotracer studies using (14)C-phenanthrene and (14)C-pyrene were used to measure the PAH-mineralization activity in salt marsh sediments. The results suggested a trend towards increased PAH mineralization in vegetated sediments relative to non-vegetated sediments. This trend was supported by the enumeration of PAH-degrading bacteria in non-vegetated and vegetated sediment using a Most Probable Numbers (MPN) technique, which demonstrated that PAH-degrading bacteria existed in non-vegetated and vegetated sediments at levels ranging from 10(2 )to 10(5 )cells/g sediment respectively. No difference between microbial communities present in vegetated versus non-vegetated sediments was found using terminal restriction fragment length polymorphism (of the 16S rRNA gene) or phospholipid fatty acid analysis. These studies provide information on the specific members and activity of the PAH-degrading aerobic bacterial communities present in Spartina-dominated salt marshes in the New York/New Jersey Harbor estuary.

Biodegradation of methyl tert-butyl ether by cold-adapted mixed and pure bacterial cultures.

An aerobic mixed bacterial culture (CL-EMC-1) capable of utilizing methyl tert-butyl ether (MTBE) as the sole source of carbon and energy with a growth temperature range of 3 to 30 degrees C and optimum of 18 to 22 degrees C was enriched from activated sludge. Transient accumulation of tert-butanol (TBA) occurred during utilization of MTBE at temperatures from 3 degrees C to 14 degrees C, but TBA did not accumulate above 18 degrees C. The culture utilized MTBE at a concentration of up to 1.5 g l(-1) and TBA of up to 7 g l(-1). The culture grew on MTBE at a pH range of 5 to 9, with an optimum pH of 6.5 to 7.1. The specific growth rate of the CL-EMC-1 culture on 0.1 g l(-1) of MTBE at 22 degrees C and pH 7.1 was 0.012 h(-1), and the growth yield was 0.64 g (dry weight) g(-1). A new MTBE-utilizing bacterium, Variovorax paradoxus strain CL-8, isolated from the mixed culture utilized MTBE, TBA, 2-hydroxy isobutyrate, lactate, methacrylate, and acetate as sole sources of carbon and energy but not 2-propanol, acetone, methanol, formaldehyde, or formate. Two other isolates, Hyphomicrobium facilis strain CL-2 and Methylobacterium extorquens strain CL-4, isolated from the mixed culture were able to grow on C(1) compounds. The combined consortium could thus utilize all of the carbon of MTBE.

Atmospheric oxygen and other conditions affecting the production of cereulide by Bacillus cereus in food.

Factors influencing the production of cereulide, the emetic toxin of Bacillus cereus in food and laboratory media were investigated, using liquid chromatography-ion trap mass spectrometry and sperm motility inhibition bioassay for detection and quantitation. Oxygen was essential for production of the emetic toxin by B. cereus. When beans, rice or tryptic soy broth were inoculated with cereulide producing strains B203, B116 (recent food isolates) or the strain F-4810/72, high amounts (2 to 7 microg ml(-1) or g(-1) wet wt) of cereulide accumulated during 4-day storage at room temperature. In parallel cultures and foods, stored under nitrogen atmosphere (> 99.5% N2), less than 0.05 microg of cereulide ml(-1) or g(-1) wet wt accumulated. The outcome of the bioassay matched that of the chemical assay, with no indication of interference by substances in the rice or beans. Boiling for 20 to 30 min did not inactivate cereulide or cereulide producing strains in rice or the beans. Adding l-leucine and l-valine (0.3 g l(-1)) stimulated cereulide production 10- to 20-fold in R2A and in rice water agar. When the B. cereus strains were grown on agar media under permissive conditions (air, room temperature), cereulide was produced overnight with little or no increase when the incubation was extended to 4 days. In broth culture, the production of cereulide started later than 16-24 h. Anoxic storage prevented cereulide production also when the amino acids had been supplied. Packaging with modified atmosphere low in oxygen may thus be used to reduce the risk of cereulide formation during storage of food.

Effects of fungal food quality and starvation on the fatty acid composition of Protaphorura fimata (Collembola).

The lipid pattern of animals is influenced by species, life stage, environmental conditions and diet. We investigated the effects of food quality and starvation on the phospholipid (PLFA) and neutral lipid (NLFA) fatty acid pattern of the collembolan Protaphorura fimata. Collembolans were fed with two common soil fungi, Agrocybe gibberosa and Chaetomium globosum, of which the cellular lipid composition was analysed. A. gibberosa was grown on agar with different nitrogen contents, resulting in altered fatty acid patterns and C:N ratios, i.e. fungi of different food quality. Collembolans did not mirror the lipid composition of the fungal diet as the pattern of major NLFAs in P. fimata was vice versa. Presumably, altered food quality of fungi caused compensatory responses by the collembolans, thereby diminishing the fungal signal. In a further experiment P. fimata (previously maintained with C. globosum) was kept without food for up to 4 weeks. Starvation resulted in a decline in the total amount of NLFAs; however, it did not affect the fatty acid pattern, indicating that NLFAs were degraded indiscriminately. Generally, the PLFA profile of the collembolans changed only slightly due to variations in diet quality or starvation.

PAH-degradation by Paenibacillus spp. and description of Paenibacillus naphthalenovorans sp. nov., a naphthalene-degrading bacterium from the rhizosphere of salt marsh plants.

Bacteria belonging to the genus Paenibacillus were isolated by enrichment from petroleum-hydrocarbon-contaminated sediment and salt marsh rhizosphere using either naphthalene or phenanthrene as the sole carbon source, and were characterized using phenotypic, morphological and molecular techniques. The isolates were grouped by their colony morphologies and polyaromatic hydrocarbon-degradation patterns. Phenanthrene-degrading isolates produced mottled colonies on solid media and were identified as P. validus by fatty acid methyl ester and 16S rRNA gene sequence analyses. In contrast, the naphthalene-degrading isolates with mucoid colony morphology were distantly related to Paenibacillus validus, according to fatty acid methyl ester and 16S rRNA gene sequence analyses. The predominant fatty acids of the mucoid isolates were 15:0 anteiso, 16:1omega11c, 16:0 and 17:0 anteiso, constituting, on average, 50.5, 12.0, 11.2 and 6.5% of the total, respectively. The G+C contents of their DNA ranged from 47 to 52 mol%. The 16S rDNA sequence analysis revealed the highest (< or = 94%) similarity to P. validus. In addition, phylogenetic analyses based on 16S rDNA sequences showed that the mucoid isolates formed a distinct cluster within Paenibacillus. DNA-DNA hybridization experiments showed only a 6% DNA similarity between the type strain of P. validus and mucoid strain PR-N1. On the basis of the morphological, phenotypic and molecular data, the naphthalene-degrading isolates merit classification as a new Paenibacillus species, for which the name Paenibacillus naphthalenovorans sp. nov. is proposed, with PR-N1T (= ATCC BAA-206T = DSM 14203T) as the type strain.

Isolation and characterization of polycyclic aromatic hydrocarbon-degrading bacteria associated with the rhizosphere of salt marsh plants.

Polycyclic aromatic hydrocarbon (PAH)-degrading bacteria were isolated from contaminated estuarine sediment and salt marsh rhizosphere by enrichment using either naphthalene, phenanthrene, or biphenyl as the sole source of carbon and energy. Pasteurization of samples prior to enrichment resulted in isolation of gram-positive, spore-forming bacteria. The isolates were characterized using a variety of phenotypic, morphologic, and molecular properties. Identification of the isolates based on their fatty acid profiles and partial 16S rRNA gene sequences assigned them to three main bacterial groups: gram-negative pseudomonads; gram-positive, non-spore-forming nocardioforms; and the gram-positive, spore-forming group, Paenibacillus. Genomic digest patterns of all isolates were used to determine unique isolates, and representatives from each bacterial group were chosen for further investigation. Southern hybridization was performed using genes for PAH degradation from Pseudomonas putida NCIB 9816-4, Comamonas testosteroni GZ42, Sphingomonas yanoikuyae B1, and Mycobacterium sp. strain PY01. None of the isolates from the three groups showed homology to the B1 genes, only two nocardioform isolates showed homology to the PY01 genes, and only members of the pseudomonad group showed homology to the NCIB 9816-4 or GZ42 probes. The Paenibacillus isolates showed no homology to any of the tested gene probes, indicating the possibility of novel genes for PAH degradation. Pure culture substrate utilization experiments using several selected isolates from each of the three groups showed that the phenanthrene-enriched isolates are able to utilize a greater number of PAHs than are the naphthalene-enriched isolates. Inoculating two of the gram-positive isolates to a marine sediment slurry spiked with a mixture of PAHs (naphthalene, fluorene, phenanthrene, and pyrene) and biphenyl resulted in rapid transformation of pyrene, in addition to the two- and three-ringed PAHs and biphenyl. This study indicates that the rhizosphere of salt marsh plants contains a diverse population of PAH-degrading bacteria, and the use of plant-associated microorganisms has the potential for bioremediation of contaminated sediments.

Characterization of halobenzoate-degrading, denitrifying Azoarcus and Thauera isolates and description of Thauera chlorobenzoica sp. nov.

The taxonomic relationships of Azoarcus and Thauera isolates in the beta-subclass of the Proteobacteria capable of degrading fluoro-, chloro- or bromobenzoate under denitrifying conditions were analysed. A detailed classification of these strains was performed using a polyphasic approach, which included studies on morphology, phenotypic characterization, fatty acid analysis, 16S rRNA gene sequence analysis, 16S rRNA gene mapping (ribotyping) and DNA-DNA hybridization. The analyses of fatty acids and 16S rRNA gene sequencing differentiated strains 2FB2, 2FB6 and 4FB10 as new members of the genus Azoarcus and strains 4FB1, 4FB2, 3CB2, 3CB3 and 3BB1 as new members of the genus Thauera. DNA-DNA hybridization experiments established that strains 2FB2, 2FB6 and 4FB10 belong to the species Azoarcus tolulyticus. Strains 3CB2 and 3CB3 were assigned to the species Thauera aromatica on the basis of DNA-DNA hybridization and ribotyping experiments. Strains 4FB1, 4FB2 and 3BB1 showed close relatedness with strain 3CB-1T, previously described as T. aromatica genomovar chlorobenzoica. This group of strains is clearly differentiated from the species T. aromatica on the basis of 16S rRNA sequence analysis, DNA homology and ribotyping analysis. Strains 3CB-1T, 4FB1, 4FB2 and 3BB1 are proposed as members of the new species Thauera chlorobenzoica sp. nov., strain 3CB-1T (= ATCC 700723T) being the type strain.

Anaerobic degradation and dehalogenation of chlorosalicylates and salicylate under four reducing conditions.

The anaerobic biodegradability and transformation of the mono-and dichlorinated salicylates (2-hydroxybenzoates) was examined under denitrifying, Fe (III) reducing, sulfate reducing and methanogenic conditions. 3,6-Dichlorosalicylate and 6-chlorosalicylate are anaerobic microbial metabolites of dicamba, a widely used herbicide. Anaerobic microcosms were established with dicamba treated soil from Wyoming, and golf course drainage stream sediments from New Jersey, which were each spiked with salicylate, 3,6-dichlorosalicylate or one of the four monochlorosalicylate isomers. Salicylate was degraded under denitrifying, sulfidogenic and methanogenic conditions. In methanogenic enrichments 5-chlorosalicylate and 3-chlorosalicylate were reductively dehalogenated to salicylate which was then utilized. Dehalogenation of monochlorinated salicylates to salicylate was also observed in denitrifying chlorosalicylate degrading cultures. The study revealed that the position of the chlorine substituent as well as the predominant electron accepting process affect the rate and extent of chlorosalicylate degradation in anoxic environments.

Anaerobic reductive dechlorination of chlorinated dioxins in estuarine sediments.

The biotransformation of 1,2,3,4-tetrachlorodibenzo-p-dioxin (1,2,3,4-tetraCDD) under anaerobic sulfate-reducing, methanogenic, and iron-reducing conditions was examined with anaerobic enrichment cultures established with sediment from an estuarine intertidal strait in the New York/New Jersey harbor. In addition, the effect of prior enrichment on 2-bromophenol or a mixture of 2-, 3-, and 4-bromophenol on dioxin dechlorination was examined. All enrichments were spiked with 1 ppm 1,2,3,4-tetraCDD and monitored by gas chromatography-mass spectrometry for up to a 3-year period. Reductive dechlorination was initially observed only under methanogenic conditions in the cultures enriched on all three bromophenol isomers. 1,2,3,4-TetraCDD was dechlorinated in the lateral position to 1,2,4-triCDD. The initial appearance of 1,2,4-triCDD was observed after 2 months, with further dechlorination to 1,3-diCDD within 17 months.

Isolation and characterization of diverse halobenzoate-degrading denitrifying bacteria from soils and sediments.

Denitrifying bacteria capable of degrading halobenzoates were isolated from various geographical and ecological sites. The strains were isolated after initial enrichment on one of the monofluoro-, monochloro-, or monobromo-benzoate isomers with nitrate as an electron acceptor, yielding a total of 33 strains isolated from the different halobenzoate-utilizing enrichment cultures. Each isolate could grow on the selected halobenzoate with nitrate as the terminal electron acceptor. The isolates obtained on 2-fluorobenzoate could use 2-fluorobenzoate under both aerobic and denitrifying conditions, but did not degrade other halobenzoates. In contrast, the 4-fluorobenzoate isolates degraded 4-fluorobenzoate under denitrifying conditions only, but utilized 2-fluorobenzoate under both aerobic and denitrifying conditions. The strains isolated on either 3-chlorobenzoate or 3-bromobenzoate could use 3-chlorobenzoate, 3-bromobenzoate, and 2- and 4-fluorobenzoates under denitrifying conditions. The isolates were identified and classified on the basis of 16S rRNA gene sequence analysis and their cellular fatty acid profiles. They were placed in nine genera belonging to either the alpha-, beta-, or gamma-branch of the Proteobacteria, namely, Acidovorax, Azoarcus, Bradyrhizobium, Ochrobactrum, Paracoccus, Pseudomonas, Mesorhizobium, Ensifer, and Thauera. These results indicate that the ability to utilize different halobenzoates under denitrifying conditions is ubiquitously distributed in the Proteobacteria and that these bacteria are widely distributed in soils and sediments.

Anaerobic degradation of fluorinated aromatic compounds.

Anaerobic enrichment cultures with sediment from an intertidal strait as inoculum were established under denitrifying, sulfate-reducing, iron-reducing and methanogenic conditions to examine the biodegradation of mono-fluorophenol and mono-fluorobenzoate isomers. Both phenol and benzoate were utilized within 2-6 weeks under all electron-accepting conditions. However, no degradation of the fluorophenols was observed within 1 year under any of the anaerobic conditions tested. Under denitrifying conditions, 2-fluorobenzoate and 4-fluorobenzoate were depleted within 84 days and 28 days, respectively. No loss of 3-fluorobenzoate was observed. All three fluorobenzoate isomers were recalcitrant under sulfate-reducing, iron-reducing, and methanogenic conditions. The degradation of the fluorobenzoate isomers under denitrifying conditions was examined in more detail using soils and sediments from different geographic regions around the world. Stable enrichment cultures were obtained on 2-fluorobenzoate or 4-fluorobenzoate with inoculum from most sites. Fluoride was released stoichiometrically, and nitrate reduction corresponded to the values predicted for oxidation of fluorobenzoate to CO2 coupled to denitrification. The 2-fluorobenzoate-utilizing and 4-fluorobenzoate-utilizing cultures were specific for fluorobenzoates and did not utilize other halogenated (chloro-, bromo-, iodo-) benzoic acids. Two denitrifying strains were isolated that utilized 2-fluorobenzoate and 4-fluorobenzoate as growth substrates. Preliminary characterization indicated that the strains were closely related to Pseudomonas stutzeri.

Description of strain 3CB-1, a genomovar of Thauera aromatica, capable of degrading 3-chlorobenzoate coupled to nitrate reduction.

A Gram-negative bacterium, strain 3CB-1, isolated from a 3-chlorobenzoate enrichment culture inoculated with a sediment sample is capable of degrading various aromatic compounds and halogenated derivatives with nitrate as electron acceptor. Compounds capable of serving as carbon and energy sources include 3-chlorobenzoate, 3-bromobenzoate, 2-fluorobenzoate, 4-fluorobenzoate, benzoate, 3-hydroxybenzoate, 4-hydroxybenzoate, 3-aminobenzoate, protocatechuate, m-cresol and p-cresol. Oxygen, nitrate and nitrite were used as electron acceptors for growth. Cells are Gram-negative short rods with peritrichous flagellation. The predominant fatty acids are cis-9-hexadecenoic acid (16:1 omega 7c), hexadecanoic acid (16:0), octadecanoic acid (18:0), octadecenoic acid (18:1), 3-hydroxydecanoic acid (10:0 3OH) and dodecanoic acid (12:0). The sequence of the 16S rRNA gene, as well as the fatty acid composition, indicate that the strain is a member of the genus Thauera in the beta-subclass of the Proteobacteria and very close to Thauera aromatica. DNA-DNA hybridization and nutrient screening indicate that strain 3CB-1 is a genomovar of Thauera aromatica with the proposed name Thauera aromatica genomovar chlorobenzoica.

Anaerobic decomposition of halogenated aromatic compounds.

Halogenated compounds constitute one of the largest groups of environmental pollutants, partly as a result of their widespread use as biocides, solvents and other industrial chemicals. A critical step in degradation of organohalides is the cleavage of the carbon?halogen bond. Reductive dehalogenation is generally the initial step in metabolism under methanogenic conditions, which requires a source of reducing equivalents, with the halogenated compound serving as an electron acceptor. Dehalogenation is greatly influenced by alternate electron acceptors; e.g. sulfate frequently inhibits reductive dehalogenation. On the other hand, a number of halogenated aromatic compounds can be degraded under different electron-accepting conditions and their complete oxidation to CO(2) can be coupled to processes such as denitrification, iron(III)-reduction, sulfate reduction and methanogenesis. Reductive dehalogenation was the initial step in degradation not only under methanogenic, but also under sulfate- and iron(III)-reducing conditions. Dehalogenation rates were in general slower under sulfidogenic and iron-reducing conditions, suggesting that dehalogenation was affected by the electron acceptor. The capacity for dehalogenation appears to be widely distributed in anoxic environments; however, the different substrate specificities and activities observed for the halogenated aromatic compounds suggest that distinct dehalogenating microbial populations are enriched under the different reducing conditions. Characterization of the microbial community structure using a combination of biomolecular techniques, such as cellular fatty acid profiling, and 16 S rRNA fingerprinting/sequence analysis, was used to discern the distinct populations enriched with each substrate and under each electron-accepting condition. These combined techniques will aid in identifying the organisms responsible for dehalogenation and degradation of halogenated aromatic compounds.

Taxonomic characterization of denitrifying bacteria that degrade aromatic compounds and description of Azoarcus toluvorans sp. nov. and Azoarcus toluclasticus sp. nov.

A taxonomic characterization of twenty-one strains capable of degrading aromatic compounds under denitrifying conditions, isolated from ten different geographical locations, was performed on the basis of general morphological and physiological characteristics, cellular fatty acids, DNA base composition, small ribosomal (16S) subunit DNA sequences, whole-cell protein patterns and genomic DNA fragmentation analysis, in addition to DNA similarity estimations using hybridization methods. The collection of strains was subdivided into a number of different groups. A first group, consisting of four strains, could be assigned to the previously described species Azoarcus tolulyticus. A second group (five strains) had DNA which reannealed highly to that of strains of the first group, and it is considered to represent a genomovar of A. tolulyticus. The third and fourth groups, composed of a total of five strains, represent a new species of Azoarcus, Azoarcus toluclasticus (group 3) and a genomovar of this species (group 4), respectively. Finally, the fifth group, with two strains, corresponds to another new species of the genus Azoarcus, Azoarcus toluvorans. In addition to these five groups, the collection includes five individual strains perhaps representing as many different new species. The above classification is partially consistent with the results of approaches other than DNA-DNA hybridization (electrophoretic patterns of whole-cell proteins and of the fragments obtained after digestion of total DNA with infrequently cutting restriction enzymes). On the other hand, no correlation of these groupings was found in terms of the cellular fatty acid composition. It is also unfortunate that no simple sets of easily determinable phenotypic properties could be defined as being characteristic of each of the groups.

Anaerobic degradation of 3-halobenzoates by a denitrifying bacterium.

A denitrifying bacterium was isolated from a river sediment after enrichment on 3-chlorobenzoate under anoxic, denitrifying conditions. The bacterium, designated strain 3CB-1, degraded 3-chlorobenzoate, 3-bromobenzoate, and 3-iodobenzoate with stoichiometric release of halide under conditions supporting anaerobic growth by denitrification. The 3-halobenzoates and 3-hydroxybenzoate were used as growth substrates with nitrate as the terminal electron acceptor. The doubling time when growing on 3-halobenzoates ranged from 18 to 25 h. On agar plates with 1 mM 3-chlorobenzoate as the sole carbon source and 30 mM nitrate as the electron acceptor, strain 3CB-1 formed small colonies (1-2 mm in diameter) in 2 to 3 weeks. Anaerobic degradation of both 3-chlorobenzoate and 3-hydroxybenzoate was dependent on nitrate as an electron acceptor and resulted in nitrate reduction corresponding to the stoichiometric values for complete oxidation of the substrate to CO2. 3-Chlorobenzoate was not degraded in the presence of oxygen. 3-Bromobenzoate and 3-iodobenzoate were also degraded under denitrifying conditions with stoichiometric release of halide, but 3-fluorobenzoate was not utilized by the bacterium. Utilization of 3-chlorobenzoate was inducible, while synthesis of enzymes for 3-hydroxybenzoate degradation was constitutively low, but inducible. Degradation was specific to the positive of the halogen substituent, and strain 3CB-1 did not utilize 2- or 4-chlorobenzoate.