Distribution of Bdellovibrio bacteriovorus in sewage works, river water, and sediments.

Bdellovibrio was found in all liquid phases of the sewage works examined. The predator was also found in all the river sediments and sewage-polluted river waters examined but could not be found in some unpolluted river waters. Bdellovibrio was able to multiply on the high numbers of bacteria present in the aerobic percolating filter film but could not survive in anaerobic sludge. Similarly, the predator was present in the aerobic surface layers of river sediments but not in the anaerobic bottom layers. The major source of Bdellovibrio in the polluted rivers examined were sewage works effluents, and numbers in both river water and sediment were correlated with river water quality. It was unlikely that Bdellovibrio was important in reducing numbers of other bacteria in either sewage or river sediment.     

Human origin of Bacteroides fragilis bacteriophages present in the environment

Bacteroides fragilis HSP40 phages have been detected in waters with various levels of fecal contamination of human origin. The average numbers of B. fragilis phages present in sewage water reached 5.3 x 10(3) per 100 ml of water. We found a number 1,000 times lower in a river contaminated with domestic sewage only, in which the levels of fecal coliforms and fecal streptococci were 10,000 times lower than those found in raw sewage. In addition, B. fragilis phages were not found in significant numbers in slaughterhouse wastewaters. They were not present in fecal-polluted waters containing fecal contamination from wildlife only. Although the number of B. fragilis phages present in contaminated waters was lower than the number of coliphages, their presence indicated human fecal contamination. It is also shown that Bacteroides phages are only able to multiply under anaerobic conditions in the presence of nutrients, and they cannot multiply in natural waters and sediments.     

Human origin of Bacteroides fragilis bacteriophages present in the environment.

Bacteroides fragilis HSP40 phages have been detected in waters with various levels of fecal contamination of human origin. The average numbers of B. fragilis phages present in sewage water reached 5.3 x 10(3) per 100 ml of water. We found a number 1,000 times lower in a river contaminated with domestic sewage only, in which the levels of fecal coliforms and fecal streptococci were 10,000 times lower than those found in raw sewage. In addition, B. fragilis phages were not found in significant numbers in slaughterhouse wastewaters. They were not present in fecal-polluted waters containing fecal contamination from wildlife only. Although the number of B. fragilis phages present in contaminated waters was lower than the number of coliphages, their presence indicated human fecal contamination. It is also shown that Bacteroides phages are only able to multiply under anaerobic conditions in the presence of nutrients, and they cannot multiply in natural waters and sediments.     

Correlation of viable cell counts,metabolic activity of sulphur-oxidizing bacteria and chemical parameters of marine sediments

Viable counts of aerobic and anaerobic chemotrophic sulphur-oxidizers as well as phototrophic sulphur bacteria were determined in sediment samples taken from two different areas along the Baltic Sea shore which were known to regularly develop sulphidic conditions. Depth profiles of bacterial cell counts were correlated with concentration profiles of chloride, sulphate, sulphide, nitrate and phosphate in the pore water of these sediments and with potential activities of nitrate reduction, thiosulphate transformation and sulphate formation. The data revealed a complex multilayered structure within the sediments. Sulphide was released into the water from sediments of both sampling areas, but it was found that light and the availability of oxygen significantly reduced this amount. In the highly reduced sediment at Hiddensee, the highest numbers of phototrophic and chemotrophic sulphur-oxidizers were found near the sediment surface. Therefore, it was concluded that the combined action of both groups of bacteria most efficiently oxidizes reduced sulphur compounds in the top layers of the sediments. Nitrate may replace oxygen as final electron acceptor and will support oxidation of sulphide, in particular when oxygen and light are limiting.     

Detection of Legionella species in sewage and ocean water by polymerase chain reaction, direct fluorescent-antibody, and plate culture methods.

Legionella spp. are ubiquitous in most environmental water sources; however, sewage treatment plants have not been examined as potential environmental reservoirs for these bacteria. This study used polymerase chain reaction, direct fluorescent-antibody staining, and culture methods to examine raw and treated sewage, ocean-receiving waters, and nearshore coastal environments for the presence of Legionella pneumophila and other Legionella spp. The study concluded that Legionella spp. are present in all phases of sewage treatment and that population numbers do not significantly decline through the treatment process. Ocean-receiving waters located 5 miles offshore, where the treated sewage is discharged, were found to contain Legionella spp., but ocean water between the discharge site and coastal bathing beaches was negative. This suggests that the Legionella spp. from the ocean discharge site were not reaching the nearshore beach waters. A flood control channel and river that entered the ocean were found to contain Legionella spp., and a nearby beach swimming area was also found to be positive, suggesting that land runoff from the flood control channel and river were the source of the Legionella spp. in the beach water samples that tested positive.     

Rapid detection of sulfide-producing bacteria from sulfate and thiosulfate

An original liquid medium and a field broth bottle method for the rapid detection of the most probable number of sulfide-producing bacteria (SPB) from sulfate and thiosulfate are described. The broth bottle method enables after inoculation with a sample (water, sediment) the growth of ubiquitous aerobic bacteria, causing oxygen depletion, required for the growth of the present various anaerobic and facultative anaerobic SPB. The medium regularly gives higher numbers of the SPB than the used control medium (Postgate's E) for detection of sulfate-reducing bacteria and the final results are obtained just 36 h after the medium inoculation. The method is simple and suitable for the estimation of the physiological group of SPB in fresh waters, saline waters, sediments and industrial waters.     

Relationships Between Environmental Factors, Bacterial Indicators, and the Occurrence of Enteric Viruses in Estuarine Sediments

Current standards for evaluation of the public health safety of recreational and shellfish-harvesting waters are based upon bacteriological analysis, but do not include an evaluation of the number of viruses. The objective of this study was to determine the occurrence of enteric viruses in estuarine sediments and to find a relationship, if any, between the presence of viruses in seawater or sediment or both and various biological and physicochemical characteristics of the environment. Viruses were found in greater numbers in sediment than in overlying seawater on a volume basis. Several types of enteroviruses were isolated: coxsackievirus types A16, B1, and B5, echovirus type 1, and poliovirus type 2. On several occasions, viruses were isolated from sediments when overlying seawaters met bacteriological water quality standards for recreational use. Statistical analysis of the relationship between viruses in seawater or in sediment and other variables measured yielded only one significant association: the number of viruses in sediment was found to be positively correlated with the number of fecal coliforms in sediment. No other physical, chemical, or biological characteristic of seawater or sediment that was measured showed statistically significant association with viral numbers. No correlation was found between bacterial indicators and virus in the overlying waters. The data indicated that evaluation of the presence of bacteria and viruses in sediment may provide additional insight into long-term water quality conditions and that indicator bacteria in water are not reflective of the concentration of enteric viruses in marine waters.     

Relationships between environmental factors, bacterial indicators, and the occurrence of enteric viruses in estuarine sediments

Current standards for evaluation of the public health safety of recreational and shellfish-harvesting waters are based upon bacteriological analysis, but do not include an evaluation of the number of viruses. The objective of this study was to determine the occurrence of enteric viruses in estuarine sediments and to find a relationship, if any, between the presence of viruses in seawater or sediment or both and various biological and physicochemical characteristics of the environment. Viruses were found in greater numbers in sediment than in overlying seawater on a volume basis. Several types of enteroviruses were isolated: coxsackievirus types A16, B1, and B5, echovirus type 1, and poliovirus type 2. On several occasions, viruses were isolated from sediments when overlying seawaters met bacteriological water quality standards for recreational use. Statistical analysis of the relationship between viruses in seawater or in sediment and other variables measured yielded only one significant association: the number of viruses in sediment was found to be positively correlated with the number of fecal coliforms in sediment. No other physical, chemical, or biological characteristic of seawater or sediment that was measured showed statistically significant association with viral numbers. No correlation was found between bacterial indicators and virus in the overlying waters. The data indicated that evaluation of the presence of bacteria and viruses in sediment may provide additional insight into long-term water quality conditions and that indicator bacteria in water are not reflective of the concentration of enteric viruses in marine waters.     

Anaerobic metabolic processes in the deep terrestrial subsurface

Anaerobic microorganisms were enumerated and metabolic activities measured in deep Coastal Plain sediments sampled from three water‐bearing formations at depths down to 300 m. Aseptically obtained sediment cores harbored the potential for anaerobic biodegradation of various substrates in almost all samples. Although the sediments were not predominantly anaerobic, viable methanogens and sulfate‐reducing bacteria (SRB) were present almost throughout the depth profile. Coliform organisms were also found at various locations, but were not recoverable from drilling muds or water used to slurry the muds. The anaerobic metabolism of lactate and formate was easily detected in most samples. However, acetate and benzoate were degraded only in portions of the subsurface that harbored methanogens. The water‐saturated transmissive zones harbored the highest numbers of SRB and the potential for the widest variety of anaerobic metabolic activities. Small or negligible anaerobic microbial activity was associated with thick clay layers. The accumulation of acetate and the production of methane in samples not amended with exogenous organic matter demonstrated that some strata contained reserves of fermentable carbon and suggested that environmental factors or nutrients other than carbon were potentially limiting in situ microbial activity.     

Autecological Study of the Chemoautotroph Nitrobacter by Immunofluorescence

Fluorescent antibodies (FA) prepared for Nitrobacter agilis and N. winogradskyi were highly reactive in homologous staining. Low-level cross-reactions between the two species were removed by adsorption. All 15 pure-culture isolates of Nitrobacter tested reacted strongly with either N. agilis FA or N. winogradskyi FA. All pure-culture isolates from soils were determined to be N. winogradskyi; those from Mammoth Cave sediments and a cattle waste oxidation ditch were N. agilis. No cross-reaction was found in extensive tests that included five isolates of Nitrosomonas europaea and 668 heterotrophic aerobic and anaerobic bacteria isolated from soil, sewage, and cave sites. The FA preparations were used to detect Nitrobacter species in Mammoth Cave sediments, in a cattle waste oxidation ditch, and in surface waters and sediments of a river and to observe that N. winogradskyi can outgrow N. agilis in enrichment culture.     

Dehalogenation in marine sediments containing natural sources of halophenols

Halophenols such as 2,4-dibromophenol (DBP) occur naturally in some marine sediments, as a consequence of various animal and algal activities. In an earlier study, DBP was observed in the burrow microenvironment of the hemichordate Saccoglossus kowalewskii. At the concentrations found in the burrow lining, aerobic respiration appeared to be inhibited significantly relative to anaerobic catabolism. This effect, as well as factors contributing to the degradation of DBP, has been documented further here. Results from the addition of radiolabeled DBP to oxic and anoxic sediment slurries and growth experiments with aerobic and anaerobic enrichments suggested that aerobes did not significantly metabolize DBP and that concentrations likely to be encountered on the inner surfaces of the burrow wall were inhibitory. In contrast, only minimal inhibition of growth occurred for anaerobes exposed to 1 mM DBP; in addition, DBP was substantially degraded in both enrichments and sediments under anaerobic conditions. Dehalogenation with the consequent production of phenol appeared to initiate anaerobic degradation. Sulfate-reducing bacteria did not dehalogenate DBP but appeared to degrade phenol. Decreased bacterial numbers and marked differences in the concentration and chemical speciation of iron in sediments from S. kowalewskii burrows may be attributed to toxic effects of DBP on aerobic bacteria.     

Laboratory Study of Chemical Speciation of Mercury in Lake Sediment and Water under Aerobic and Anaerobic Conditions

Chemical speciation and partitioning of radiolabeled HgCl₂ were studied in model aquatic systems consisting of undisturbed eutrophic lake sediment and water in plastic cylinders. The cylinders were either gradually made anaerobic by a gentle flow of N₂-CO₂ or kept aerobic by air flow. The proportion of methylated ²⁰³Hg was significantly higher, in both water and sediment, in the anaerobic systems than in the aerobic systems. The composition and total concentration of fatty acids originating from bacterial phospholipids, as well as the concentration of vitamin B₁₂, including related cobalamins, were similar in sediments from the anaerobic and aerobic systems. Bacterial cell numbers were, on average, 3.6 times higher in the anaerobic water columns than in the aerobic ones. Volatilization of ²⁰³Hg occurred in all systems except in an autoclaved control and was of similar magnitudes in the anaerobic and aerobic systems. Incorporation of ²⁰³Hg into the sediment was significantly faster in the aerobic systems than in the anaerobic systems. These results suggest that episodes of anoxia in bottom waters and sediment cause an increase in net mercury methylation and, hence, an increase in bioavailable mercury.     

Dehalogenation in marine sediments containing natural sources of halophenols.

Halophenols such as 2,4-dibromophenol (DBP) occur naturally in some marine sediments, as a consequence of various animal and algal activities. In an earlier study, DBP was observed in the burrow microenvironment of the hemichordate Saccoglossus kowalewskii. At the concentrations found in the burrow lining, aerobic respiration appeared to be inhibited significantly relative to anaerobic catabolism. This effect, as well as factors contributing to the degradation of DBP, has been documented further here. Results from the addition of radiolabeled DBP to oxic and anoxic sediment slurries and growth experiments with aerobic and anaerobic enrichments suggested that aerobes did not significantly metabolize DBP and that concentrations likely to be encountered on the inner surfaces of the burrow wall were inhibitory. In contrast, only minimal inhibition of growth occurred for anaerobes exposed to 1 mM DBP; in addition, DBP was substantially degraded in both enrichments and sediments under anaerobic conditions. Dehalogenation with the consequent production of phenol appeared to initiate anaerobic degradation. Sulfate-reducing bacteria did not dehalogenate DBP but appeared to degrade phenol. Decreased bacterial numbers and marked differences in the concentration and chemical speciation of iron in sediments from S. kowalewskii burrows may be attributed to toxic effects of DBP on aerobic bacteria.     

Survival and Activity of Bacteria in a Deep,Aged Lake Sediment (Lake Constance)

Abstract Viable counts and potential activities of different bacteria were determined as a function of depth in the deep profundal sediment of Lake Constance, Germany. The sediment layer at the bottom of the lake had a total depth of about 7 m and was deposited in the time after the last ice age, i.e., over the past 13,000 years. The high clay content of the sediment prevents seepage. Below 25 cm all of the viable heterotrophic bacteria were present as heat-resistant spores. Numbers of viable spores of both aerobic and anaerobic heterotrophic bacteria decreased exponentially with sediment depth and were below the detection limit (5–55 cells ml⁻¹) at 4–6 m, i.e., in about 8,900-year-old sediment. Absence of viable heterotrophic bacteria in deeper sediment layers demonstrated that aseptic sampling conditions were achieved. The decrease of viable spores with depth may be interpreted as time-dependent death of spores resulting in a death rate of about 0.0013–0.0025 year⁻¹. Viable units of specific metabolic groups of bacteria were detected only in the upper sediment layers (0–50 cm). Nitrifying bacteria could not be detected below 30 cm. Methane-oxidizing bacteria were present in the sediment down to >30 cm, but were in a dormant state. Nitrate reduction activity decreased by a factor of 6 within the upper 25 cm of the sediment, but was still detected at 50 cm. Sulfate reduction, on the other hand, could not be detected at depths of 20 cm and below. By contrast, methanogenesis and methanogenic bacteria could be detected down to 50 cm. These observations indicate that bacteria eventually become nonviable in aged sediments. Received: 5 March 1996; Accepted: 12 March 1996     

Microbial communities associated with phosphogenic sediments and phosphoclast‐associated DNA of the Benguela upwelling system

The processes that lead to the precipitation of authigenic calcium phosphate minerals in certain marine pore waters remain poorly understood. Phosphogenesis occurs in sediments beneath some oceanic upwelling zones that harbor polyphosphate‐accumulating bacteria. These bacteria are believed to concentrate phosphate in sediment pore waters, creating supersaturated conditions with respect to apatite precursors. However, the relationship between microbes and phosphorite formation is not fully resolved. To further study this association, we examined microbial community data generated from two sources: sediment cores recovered from the shelf of the Benguela upwelling region where phosphorites are currently forming, and DNA preserved within phosphoclasts recovered from a phosphorite deposit along the Benguela shelf. iTag and clone library sequencing of the 16S rRNA gene showed that many of our sediment‐hosted communities shared large numbers of phylotypes with one another, and that the same metabolic guilds were represented at localities across the shelf. Sulfate‐reducing bacteria and sulfur‐oxidizing bacteria were particularly abundant in our datasets, as were phylotypes that are known to carry out nitrification and the anaerobic oxidation of ammonium. The DNA extracted from phosphoclasts contained the signature of a distinct microbial community from those observed in the modern sediments. While some aspects of the modern and phosphoclast communities were similar, we observed both an enrichment of certain common microbial classes found in the modern phosphogenic sediments and a relative depletion of others. The phosphoclast‐associated DNA could represent a relict signature of one or more microbial assemblages that were present when the apatite or its precursors precipitated. While these taxa may or may not have contributed to the precipitation of the apatite that now hosts their genetic remains, several groups represented in the phosphoclast extract dataset have the genetic potential to metabolize polyphosphate, and perhaps modulate phosphate concentrations in pore waters where carbonate fluorapatite (or its precursors) are known to be precipitating.     

The mineralization of chloroform in river sediments

The formation of¹⁴CO₂ from 3 μg l⁻¹ labelled chloroform was studied in anaerobic Dutch river sediments. All incubations were performed under anaerobic conditions. The observed first order mineralization kinetics showed half-lives of 2–37 days at 20°C in 12 muddy sediments. In contrast most of the sandy sediment samples did not show a mineralization of chloroform. Most probable number analysis revealed about 3.10⁴ chloroform mineralizing bacteria per g of dry sediment in a muddy sediment and 1–2.10³ chloroform mineralizing bacteria per g of dry sediment in a sandy sediment. Therefore the persistence of chloroform in sandy sediments is not caused by the absence of chloroform mineralizing bacteria but by the inactivity of these bacteria. This inactivity of the sandy sediments might allow chloroform from infiltrating river water to reach the groundwater. Mud samples from a relatively unpolluted site showed a similar chloroform mineralization rate compared with the polluted sediments from the rivers Rhine and Meuse. The data indicate that the reductive dechlorination of aliphatic compounds is not influenced at the polluted sites.     

Occurrence of Cryptosporidium oocysts in sewage effluents and selected surface waters

An existing method for the detection of Cryptosporidium oocysts in water was modified to investigate oocyst prevalence in large volumes of water. Surface waters and sewage effluents were filtered, eluted from the filter, and concentrated using centrifugation. The resultant pellet was then homogenized, sonicated, and placed on a sucrose gradient to separate oocysts from the sediment. The uppermost gradient layer was then examined by immunofluorescence using a labeled monoclonal antibody. Using this technique, average numbers of oocysts detected in raw and treated sewage were 5.18 X 10(3) and 1.30 X 10(3)/L, respectively. Filtered sewage effluents had significantly lower numbers of oocysts (10.0/L). These data show that sand filtration may reduce the concentrations of this parasite in waste waters. Highly variable oocyst numbers were encountered in surface waters. Since Cryptosporidium oocysts are frequently present in environmental waters, they could be responsible for waterborne outbreaks of disease.     

Abundance and Activity of Methanotrophic Bacteria in Littoral and Profundal Sediments of Lake Constance (Germany)

The abundances and activities of aerobic methane-oxidizing bacteria (MOB) were compared in depth profiles of littoral and profundal sediments of Lake Constance, Germany. Abundances were determined by quantitative PCR (qPCR) targeting the pmoA gene and by fluorescence in situ hybridization (FISH), and data were compared to methane oxidation rates calculated from high-resolution concentration profiles. qPCR using type I MOB-specific pmoA primers indicated that type I MOB represented a major proportion in both sediments at all depths. FISH indicated that in both sediments, type I MOB outnumbered type II MOB at least fourfold. Results obtained with both techniques indicated that in the littoral sediment, the highest numbers of methanotrophs were found at a depth of 2 to 3 cm, corresponding to the zone of highest methane oxidation activity, although no oxygen could be detected in this zone. In the profundal sediment, highest methane oxidation activities were found at a depth of 1 to 2 cm, while MOB abundance decreased gradually with sediment depth. In both sediments, MOB were also present at high numbers in deeper sediment layers where no methane oxidation activity could be observed.     

Functional Groups and Activities of Bacteria in a Highly Acidic Volcanic Mountain Stream and Lake in Patagonia,Argentina

Acidic volcanic waters are naturally occurring extreme habitats that are subject of worldwide geochemical research but have been little investigated with respect to their biology. To fill this gap, the microbial ecology of a volcanic acidic river (pH approximately equal to 0-1.6), Rio Agrio, and the recipient lake Caviahue in Patagonia, Argentina, was studied. Water and sediment samples were investigated for Fe(II), Fe(III), methane, bacterial abundances, biomass, and activities (oxygen consumption, iron oxidation and reduction). The extremely acidic river showed a strong gradient of microbial life with increasing values downstream and few signs of life near the source. Only sulfide-oxidizing and fermentative bacteria could be cultured from the upper part of Rio Agrio. However, in the lower part of the system, microbial biomass and oxygen penetration and consumption in the sediment were comparable to non-extreme aquatic habitats. To characterize similarities and differences of chemically similar natural and man-made acidic waters, our findings were compared to those from acidic mining lakes in Germany. In the lower part of the river and the lake, numbers of iron and sulfur bacteria and total biomass in sediments were comparable to those known from acidic mining lakes. Bacterial abundance in water samples was also very similar for both types of acidic water (around 10(5) mL(-1)). In contrast, Fe(II) oxidation and Fe(III) reduction potentials appeared to be lower despite higher biogenic oxygen consumption and higher photosynthetic activity at the sediment-water interface. Surprisingly, methanogenesis was detected in the presence of high sulfate concentrations in the profundal sediment of Lake Caviahue. In addition to supplementing microbiological knowledge on acidic volcanic waters, our study provides a new view of these extreme sites in the general context of aquatic habitats.     

Influence of soil on fecal indicator organisms in a tidally influenced subtropical environment

The potential regrowth of fecal indicator bacteria released into coastal environments in recreational water bodies has been of concern, especially in tropical and subtropical areas where the number of these bacteria can be artificially elevated beyond that from fecal impacts alone. The task of determining the factors that influence indicator bacterial regrowth was addressed though a series of field sampling and laboratory experiments using in situ densities of Escherichia coli, enterococci, and Clostridium perfringens in river water, sediment, and soil. Field sampling efforts included the collection of surface sediments along the cross section of a riverbank, a 20-cm-deep soil core, and additional surface soils from remote locations. In addition to field sampling, two types of laboratory experiments were conducted. The first experiment investigated the survival of bacteria already present in river water with the addition of sterile and unsterile sediment. The second experiment was designed to simulate the wetting and drying effects due to tidal cycles. The results from the sampling study found elevated numbers of E. coli and C. perfringens in surficial sediments along the riverbank near the edge of the water. C. perfringens was found in high numbers in the subsurface samples obtained from the soil core. Results from laboratory experiments revealed a significant amount of regrowth for enterococci and E. coli with the simulation of tides and addition of sterile sediment. Regrowth was not observed for C. perfringens. This study demonstrates the need to further evaluate the characteristics of indicator microbes within tropical and subtropical water systems where natural vegetation, soil embankments, and long-term sediment accumulation are present. In such areas, the use of traditional indicator microbes to regulate recreational uses of a water body may not be appropriate.