Amperometric estimation of BOD by using living immobilized yeasts

Summary A microbial electrode consisting of immobilized living whole cells of yeasts, porous membrane and an oxygen electrode was prepared for continuous estimation of biochemical oxygen demand (BOD). Immobilized Trichosporon cutaneum was employed for the microbial electrode sensor for BOD. When a sample solution containing the equivalent amount of glucose and glutamic acid was injected into the sensor system, the current of the electrode decreased markedly with time until steady state was reached. The response time was within 18 min. A linear relationship was observed between the current decrease and the concentration below 41 mg l ^– of glucose and 41 mg l ^– glutamic acid (5-day BOD 60 mg l ^–). The current decrease was reproducible within ± 6% of the relative error when a sample solution containing 27 mg l ^– of glucose and 27 mg l ^– of glutamic acid (5-day BOD 40 mg l ^–) was employed. The microbial electrode sensor was applied to untreated waste waters from a fermentation factory. Good comparative results were obtained between BOD estimated by the microbial electrode and that determined by the conventional 5-day method (regression coefficient was 1.2). Furthermore, the effect of various compounds on BOD estimation was also examined. The current output of the microbial electrode sensor was almost constant for 17 d and 400 tests.     

Analyses of soil bacterial diversity of the Schirmacher Oasis,Antarctica

Schirmacher Oasis, Antarctica, is a region with relatively large exposed area and consisted of many freshwater lakes. Nevertheless, only a few studies were done on the bacterial diversity of this region. Hence, this project was undertaken to determine the bacterial community in soil samples collected from the Schirmacher Oasis using the denaturing gradient gel electrophoresis (DGGE) of amplified 16S rDNA fragments. A total of 79 partial 16S rDNA sequences were obtained from the excised DGGE bands, which corresponded to 63 different operational taxonomic units (OTUs) representing bacteria from seven different phyla. The most dominant phyla in descending order were Acidobacteria, Proteobacteria, Bacteroidetes, and Actinobacteria, Planctomycetes, Cyanobacteria and BRC1. There were 5.4 % of unclassified bacteria which cannot be grouped into any of the existing phyla. Eighty-seven percent of the OTUs had highest similarity with the uncultured bacteria from the NCBI GenBank database. Thirty-two percent of the OTUs were similar to bacteria reported in other parts of the Antarctica, while the others were related to bacteria found elsewhere outside the Antarctic.     

The Occurrence of Lysogenic Bacteria and Microbial Aggregates in the Lakes of the McMurdo Dry Valleys,Antarctica

The McMurdo Dry Valleys of Antarctica form the coldest and driest ecosystem on Earth. Within this region there are a number of perennially ice-covered (3–6 m thick) lakes that support active microbial assemblages and have a paucity of metazoans. These lakes receive limited allochthonous input of carbon and nutrients, and primary productivity is limited to only 6 months per year owing to an absence of sunlight during the austral winters. In an effort to establish the role that bacteria and their associated viruses play in carbon and nutrient cycling in these lakes, indigenous bacteria, free bacteriophage, and lysogen abundances were determined. Total bacterial abundances (TDC) ranged from 3.80 × 10⁴ to 2.58 × 10⁷ cells mL^–1 and virus-like particle (VLP) abundances ranged from 2.26 × 10⁵ to 5.56 × 10⁷ VLP mL^–1. VLP abundances were significantly correlated (P < 0.05) with TDC, bacterial productivity (TdR), chlorophyll a (Chl a), and soluble reactive phosphorus (SRP). Lysogenic bacteria, determined by induction with mitomycin C, made up between 2.0% and 62.5% of the total population of bacteria when using significant decreases and increases in TDC and VLP abundances, respectively, and 89.5% when using increases in VLP abundances as the sole criterion for a successful induction event. The contribution of viruses released from induced lysogens contributed <0.015% to the total viral production rate. Carbohydrate and protein based organic aggregates were abundant within the water column of the lakes and were heavily colonized by bacteria and VLPs. Alkaline phosphatase activity was detected within the matrix of the aggregates, implying phosphorus deficiency and consortial nutrient exchanges among microorganisms.     

Antimycobacterial activity in vitro of pigments isolated from Antarctic bacteria

In this study, we describe the antimycobacterial activity of two pigments, violacein, a purple violet pigment from Janthinobacterium sp. Ant5-2 (J-PVP), and flexirubin, a yellow-orange pigment from Flavobacterium sp. Ant342 (F-YOP). These pigments were isolated from bacterial strains found in the land-locked freshwater lakes of Schirmacher Oasis, East Antarctica. The minimum inhibitory concentrations (MICs) of these pigments for avirulent and virulent mycobacteria were determined by the microplate Alamar Blue Assay (MABA) and Nitrate Reductase Assay (NRA). Results indicated that the MICs of J-PVP and F-YOP were 8.6 and 3.6 μg/ml for avirulent Mycobacterium smegmatis mc²155; 5 and 2.6 μg/ml for avirulent Mycobacterium tuberculosis mc²6230; and 34.4 and 10.8 μg/ml for virulent M. tuberculosis H₃₇Rv, respectively. J-PVP exhibited a ~15 times lower MIC for Mycobacterium sp. than previously reported for violacein pigment from Chromobacterium violaceum, while the antimycobacterial effect of F-YOP remains undocumented. Our results indicate these pigments isolated from Antarctic bacteria might be valuable lead compounds for new antimycobacterial drugs used for chemotherapy of tuberculosis.     

Observations of Barophilic Microbial Activity in Samples of Sediment and Intercepted Particulates from the Demerara Abyssal Plain

To better understand the ecological significance of pressure effects on bacteria in the abyssobenthic boundary layer, experimental suspensions of sediments and sinking particulates were prepared from samples collected in boxcore and bottom-moored sediment traps at two stations (depth, 4,470 and 4,850m) in the Demerara abyssal plain off the coast of Brazil. Replicate samples were incubated shipboard at 3°C and at both atmospheric and deep-sea pressures (440 or 480 atm [4.46 × 10⁴ or 4.86 × 10⁴ kPa]) following the addition of [¹⁴C]glutamic acid (<10 μg liter⁻¹) or yeast extract (0.025%) and the antibiotic nalidixic acid (0.002%). In seven of the eight samples supplemented with isotope, a barophilic microbial response was detected, i.e., substrate incorporation and respiration were greater under in situ pressure than at 1 atm (101.3 kPa). In the remaining sample, prepared from a sediment trap warmed to 24°C before recovery, pressure was observed to inhibit substrate utilization. Total bacterial counts by epifluorescence microscopy decreased with depth in each sediment core, as did utilization of glutamic acid. Significant percentages of the total bacterial populations in cold sediment trap samples (but not the prewarmed one or any boxcore sample) were abnormally enlarged and orange fluorescing after incubation with yeast extract and nalidixic acid under deep-sea conditions. Results indicated that in the deep sea, barophilic bacteria play a predominant role in the turnover of naturally low levels of glutamic acid, and the potential for intense microbial activity upon nutrient enrichment is more likely to occur in association with recently settled particulates, especially fecal pellets, than in buried sediments.     

Microbiological Survey of Adirondack Lakes with Various pH Values

Nine high-altitude oligotrophic Adirondack lakes in upstate New York having water of pH 4.3 to 7.0 were surveyed for total bacterial numbers and possible adaptation of the microbial communities to environmental pH. The number of heterotrophic bacteria from water samples recoverable on standard plate count agar were low (10¹ to 10³ per ml) for most of the lakes. Acridine orange direct counts were approximately two orders of magnitude higher than plate counts for each lake. Sediment aerobic heterotrophs recovered on standard plate count agar ranged from 1.4 × 10⁴ to 1.3 × 10⁶ per g of sediment. Direct epifluorescence counts of bacteria in sediment samples ranged from 3.0 × 10⁶ to 1.4 × 10⁷ per g. Low density values were consistent with the oligotrophic nature of all the lakes surveyed. There were no apparent differences in numbers of bacteria originally isolated at pH 5.0 and pH 7.0 between circumneutral lakes (pH > 6.0) and acidic lakes (pH < 5.0). Approximately 1,200 isolates were recultured over a range of pH from 3.0 to 7.0. Regardless of the original isolation pH (pH 5.0 or pH 7.0), less than 10% of the isolates grew at pH < 5.0. Those originally isolated at pH 5.0 also grew at pH 6.0 and 7.0. Those originally isolated at pH 7.0 preferred pH 7.0, with 98% able to grow at pH 6.0 and 44% able to grow at pH 5.0. A chi-square contingency test clearly showed (P < 0.005) that two distinct heterotrophic populations had been originally isolated at pH 5.0 and pH 7.0, although there is undoubtedly some overlap between the two populations.     

Isolates of Arthrobacter from the soils of Schirmacher Oasis,Antarctica

Summary Thirteen isolates of bacteria from the soils of Schirmacher Oasis, Antarctica have been identified as members of the genus Arthrobacter. All the isolates exhibited a rod-coccus cycle during growth; were gram positive, catalase positive, non-motile and non-fermentative; did not form endospores; and contained MK-8 (H₂) as the major menaquinone. The mole %G + C in DNA of the isolates ranged from 58% to 72%. Based on their morphology, physiology, nutritional requirements, biochemical characteristics, and mole %G + C of their DNA, the isolates were identified as A. globiformis, A. pascens and A. protophormiae. However, unlike the mesophilic isolates the Antarctic Arthrobacter could be considered to be unique as they were psychrotrophic, contained glucose and lysine in the cell wall, and did not hydrolyze starch.     

Microbial biomass and activity in subsurface sediments from Vejen,Denmark

Subsurface sediment samples were collected from 4 to 31 m below landsurface in glacio-fluvial sediments from the Quaternary period. The samples were described in terms of pH, electrical conductivity, chloride concentration, organic matter content, and grain size distribution. Viable counts of bacteria varied from 0.5 to 1,203 x 103 colony forming units/g dry weight (gdw); total numbers of bacteria acridine orange direct counts (AODC) varied from 1.7 to 147 × 10⁷ cells/gdw; growth rates (incorporation of [³H]-thymidine) varied from 1.4 to 60.7 × 10⁴ cells/(gdw · day); and rate constants for mineralization of ¹⁴C-labelled compounds varied from 0.2 to 2.3 × 10^–3 ml/(dpm · day) for acetate, and from 0 to 2.0 × 10^–3 ml/(dpm · day) for phenol. Sediment texture influenced the total number of bacteria and potential for mineralization; with increasing content of clay and silt and decreasing content of sand, AODC increased and the mineralization rate declined. Intrinsic permeability calculated from grain size correlated positively with mineralization rate for acetate. Statistical correlation analysis showed high correlations between some of the abiotic parameters, but it was not possible to point out a single abiotic parameter that could explain the variation of size and activity of the microbial population. The microbial data obtained in these geologically young sediments were compared to literature data from older sediments, and this comparison showed that age and type of geological formation might be important for the size and activity of the microbial populations.Offprint requests to: H.-J. Albrechtsen.     

The microbial plankton of freshwater lakes in the Vestfold Hills,Antarctica

Summary The plankton of twelve freshwater and slightly saline lakes in the Vestfold Hills, Antarctica was sampled in February 1991. All of the lakes are oligotrophic. The chlorophyll a concentrations in the lakes ranged from 0.10–2.69 g · 1^–1. The majority of the phytoplankton were flagellates or picoplanktonic cyanobacteria with the species composition varying between the lakes. Cyanobacteria were found in five of the lakes. Five to 6 species of ciliated protozoa occurred, among them oligotrichs, including the mixotrophic species Strombidium viride. The concentrations of protists and bacteria were an order to several orders of magnitude lower than reported from lower latitude oligotrophic lakes. Low species diversity and low numbers in the plankton characterise these eastern Antarctica lakes which reflects their low nutrient status and isolation.     

Kinetic analysis of ethanol production by an acetate-resistant strain of recombinant Zymomonas mobilis

Zymomonas mobilis ZM4/Ac^R (pZB5), a mutant recombinant strain with increased acetate resistance, has been isolated following electroporation of Z. mobilis ZM4/Ac^R. This mutant strain showed enhanced kinetic characteristics in the presence of 12 g sodium acetate l^–1 at pH 5 in batch culture on 40 g glucose, 40 g xylose l^–1 medium when compared to ZM4 (pZB5). In continuous culture, there was evidence of increased maintenance energy requirements/uncoupling of metabolism for ZM4/Ac^R (pZB5) in the presence of sodium acetate; a result confirmed by analysis of the effect of acetate on other strains of Z. mobilis. Nomenclature m Cell maintenance energy coefficient (g g^–1 h^–1)Maximum overall specific growth rate (1 h^–1)Maximum specific ethanol production rate (g g^–1 h^–1)Maximum specific total sugar utilization rate (g g^–1 h^–1)Biomass yield per mole of ATP (g mole^–1 Ethanol yield on total sugars (g g^–1)Biomass yield on total sugars (g g^–1)True biomass yield on total sugars (g g^–1)     

Microbial population dynamics on seeds during drum and steeping priming

In the UK, seeds are primed commercially via drum or steeping priming processes to improve seed germination and seedling establishment but the microbial population dynamics occurring during these processes are unknown. Consequently, changes in culturable bacterial and fungal populations that occurred during laboratory and commercial scale drum priming of carrot, leek and parsnip seed and during laboratory scale and commercial scale steeping priming of carrot and sugar beet seed were investigated. Populations of bacteria and pseudomonads increased by 2 and 4 log₁₀ cfu g^–1 seed during priming to reach between 7 and 10 log₁₀ cfu g^–1 seed with less than 1 log₁₀ cfu g^–1 seed decrease after redrying, irrespective of seed type or priming process. Pseudomonads represented approximately 10% of the culturable bacterial population. Fungal populations also increased by between 1 and 3 log₁₀ cfu g^–1 seed during priming of carrot, leek and parsnip seed and were little affected by redrying. Addition of a tefluthrin coating (a standard commercial insecticide treatment) during the drying stage of commercial scale drum priming had little effect on microbial populations. During steeping priming, populations of bacteria and pseudomonads on sugar beet also increased by 2 and 4 log₁₀ cfu g^–1 seed with less than 1 log₁₀ cfu g^–1 seed decrease after redrying. However, fungi did not increase in the same manner on sugar beet, suggesting that the sugar beet seed itself may have an inhibitory effect on fungal growth. The presence of thiram in the steeping liquid (a standard commercial fungicide treatment) inhibited fungal proliferation on carrot and sugar beet seed. Spore forming bacteria generally occurred at low levels on all seeds (between 2 and 5 log₁₀ cfu g^–1seed) and did not exhibit any characteristic population changes during priming or redrying. Clearly some culturable populations of microorganisms can increase reproducibly during these priming processes and survive the redrying procedure, thereby demonstrating the potential for a novel method of introducing microbial inoculants onto seed.     

Effects of Inorganic Particles on Metabolism by a Periphytic Marine Bacterium

Measurements were made of adsorption of a periphytic marine bacterium, glucose, and glutamic acid to inorganic particles in seawater and defined bacterial growth medium. Measurements of the metabolism of bacteria were made in the presence and absence of particles by microcalorimetry and radiorespirometry. It was found that hydroxyapatite adsorbs glutamic acid, but not glucose, from the experimental medium. It was also found that hydroxyapatite adsorbs essentially all of the bacteria from the medium when the bacterial concentration is approximately 6 × 10⁵ bacteria per ml. If the bacterial concentration is approximately 6 × 10⁷, then only a small fraction of cells become attached. It was therefore possible to select bacterial concentrations and organic nutrients so that bacterial attachment, organic nutrient adsorption, or both would occur in different experiments. In this experimental system the metabolism by attached and nonattached bacteria of adsorbing and nonadsorbing organic nutrients was measured. The results show that bacterial activity in this model system was not enhanced by the particles, regardless of whether the bacteria, the organic nutrient, or both were associated with the surface. In fact, the respiratory activity of the attached bacteria was diminished in comparison with that of free bacteria.     

Production and decomposition processes in a saline meromictic lake

Bacterial and phytoplankton cell number and productivity were measured in the mixolimnion and chemocline of saline meromictic Mahoney Lake during the spring (Apr.–May) and fall (Oct.) between 1982 and 1987. High levels of bacterial productivity (methyl ³H-thymidine incorporation), cell numbers, and heterotrophic assimilation of ¹⁴C-glucose and ¹⁴C-acetate in the mixolimnion shifted from near surface (1.5 m), at a secondary chemocline, to deeper water (4–7 m) as this zone of microstratification gradually weakened during a several year drying trend in the watershed. In the mixolimnion, bacterial carbon (13–261 µgC 1^–1) was often similar to phytoplankton carbon (44–300 µgC 1^–1) and represented between 14–57% of the total microbial (phytoplankton + bacteria) carbon depending on the depth interval. Phototrophic purple sulphur bacteria were stratified at the permanent primary chemocline (7.5–8.3 m) in a dense layer (POC 250 mg 1^–1, bacteriochlorophyll a 1500–70001µ 1^–1), where H₂S changed from 0.1 to 2.5 mM over a 0.2 m depth interval. This phototrophic bacterial layer contributed between 17–66% of the total primary production (115–476 mgC m^–2 d^–1) in the vertical water column. Microorganisms in the phototrophic bacterial layer showed a higher uptake rate for acetate (0.5–3.7 µC 1^–1 h^–1) than for glucose (0.3–1.4 µgC 1^–1 h^–1) and this heterotrophic activity as well as bacterial productivity were 1 to 2 orders of magnitude higher in the dense plate than in the mixolimnetic waters above. Primary phytoplanktonic production in the mixolimnion was limited by phosphorus while light penetration appeared to regulate phototrophic productivity of the purple sulphur bacteria.     

Trophic interactions among heterotrophic microplankton,nanoplankton, and bacteria in Lake Constance

A considerable portion of the pelagic energy flow in Lake Constance (FRG) is channelled through a highly dynamic microbial food web. In-situ experiments using the lake water dilution technique according to Landry & Hasset (1982) revealed that grazing by heterotrophic nanoflagellates (HNF) smaller than 10 µm is the major loss factor of bacterial production. An average flagellate ingests 10 to 100 bacteria per hour. Nano- and micro-ciliates have been identified as the main predators of HNF. If no other food is used between 3 and 40 HNF are consumed per ciliate and hour. Other protozoans and small metazoans such as rotifers are of minor importance in controlling HNF population dynamics.Clearance rates varied between 0.2 and 122.8 nl HNF^–1 h^–1 and between 0.2 and 53.6 µl ciliate^–1 h^–1, respectively.Ingestion and clearance rates measured for HNF and ciliates are in good agreement with results obtained by other investigators from different aquatic environments and from laboratory cultures. Both the abundance of all three major microheterotrophic categories — bacteria, HNF, and ciliates — and the grazing pressure within the microbial loop show pronounced seasonal variations.     

Seasonal variations of metabolically active bacteria in a hypertrophic lake (Hartbeespoort Dam,South Africa)

The number of metabolically active bacteria was measured with nalidixic acid over two annual cycles at three depths in the epilimnion of hypertrophic Hartbeespoort Dam, South Africa. Concurrent measurements were made of water temperature, DOC, phytoplankton production of dissolved (EDOC) and particulate organic carbon, chlorophyll a and the uptake of glucose (V_max). The objective was to determine the dominant factors correlated to the number of metabolically active bacteria and the relationship between active bacterial numbers and heterotrophic activity.The number of active bacteria was usually highest at the surface and ranged between 0.70 and 6.82 x 10⁶ cells ml^–1. The dominant factors correlated to the number of bacteria at the surface were water temperature (r = 0.65, n = 54, p<0.001), primary production (r = 0.53, n = 51, p<0.001) and EDOC (r = 0.37, n = 45, p = 0.005). Surface V_max for glucose ranged between 0.11 and 4.0 µgC 1^–1 h^–1 and was positively correlated to the number of active bacteria (r = 0.61, n = 53, p<0.001). The specific activity index (10^–12 µgC cell^–1 h^–1) varied between 80 and 2290 at the surface and was most strongly correlated to EDOC (r = 0.70, n = 48, p<0.001). Relationships between active bacterial numbers, water temperature, phytoplankton activity and glucose uptake were also found at two additional depths within the epilimnion. These data suggest that bacterial populations in nutrient enriched lakes contain a large number of metabolically active cells with high individual activity as a result of enhanced phytoplankton growth.     

Bacterial diversity and activity along a salinity gradient in soda lakes of the Kulunda Steppe (Altai, Russia)

Here we describe the diversity and activity of sulfate reducing bacteria along a salinity gradient in four different soda lakes from the Kulunda Steppe (South East Siberia, Russia). For this purpose, a combination of culture-dependent and independent techniques was applied. The general bacterial and SRB diversity were analyzed by denaturing gradient gel electrophoresis (DGGE) targeting the 16S rDNA gene. DNA was used to detect the microbial populations that were present in the soda lake sediments, whereas ribosomal RNA was used as a template to obtain information on those that were active. Individual DGGE bands were sequenced and a phylogenetic analysis was performed. In addition, the overall activity of SRB was obtained by measuring the sulfate reduction rates (SRR) and their abundance was estimated by serial dilution. Our results showed the presence of minor, but highly active microbial populations, mostly represented by members of the Proteobacteria. Remarkably high SRR were measured at hypersaline conditions (200 g L⁻¹). A relatively high viable count indicated that sulfate reducing bacteria could be highly active in hypersaline soda lakes. Furthermore, the increase of sodium carbonate/bicarbonate seemed to affect the composition of the microbial community in soda lakes, but not the rate of sulfate reduction.     

Glutamic acid production byArthrobacter globiformis

Two hundred and fiftyArthrobacter strains were tested in a basal salts-glucose medium for their ability to produce glutamic acid; 50 strains produced small amounts of glutamic acid and alanine, as well as traces of other amino acids. Five biotin-dependent strains produced extraordinarily large amounts of glutamic acid. One of these, which was identified asA. globiformis, was selected for further study. Glutamic acid was only produced by this organism at biotin levels suboptimal for growth; maximal production (0.45 moles of glutamic acid per mole of glucose consumed) occurred at a biotin level of 10^–5 µg/ml. Other factors which markedly influenced glutamic acid production were temperature, (NH₄)₂SO₄ concentration, and pH of the growth medium.The taxonomy of glutamic acid-producing bacteria and the correlation between biotin deficiency and glutamic acid production are discussed.     

Planktonic bacterial biomass and seasonal pattern of the heterotrophic uptake and respiration of glucose and amino acids in the shallow sandpit lake of Créteil (Paris Suburb,France)

Biomass and activity of planktonic bacteria were investigated during a one year study in a shallow sandpit lake. The shallowness of the lake helped keep the water column homogeneous regarding bacterioplankton. Small free-living bacteria (0.03 µm³ cell^–1) dominated the populations throughout the period studied. Bacterial abundances varied from 1 to 11 × 10⁶ cells ml^–1. Kinetic parameters (V _max, K + S and T) were determined with ¹⁴C labelled compounds (glucose and amino acids mixture). V _max values were high and averaged 0.056 and 0.050 µgCl^–1 h^–1 for glucose and amino acids respectively. Maximal V _max values were observed in summer at the highest temperatures, but also in early spring. T values were much greater in winter. K + S values were significantly higher for amino acids (3 µg Cl^–1) than for glucose (1 µg Cl^–1). A low percentage of mineralization (about 25% for both tracers) could be the expression of the high growth efficiency expected when bacteria are growing at the expense of low molecular weight compounds as phytoplankton exudates.     

Effects of selection and fate of substrates supplied to anaerobic bacteria involved in the corrosion of pipe-line steel

Summary The corrosion of AISI C1020 carbon steel in an anoxic, marine, sulphide-containing environment was examined as a function of bacterial physiology and consortial complexity. The carbon steel was exposed to three organism;Eubacterium limosum, Desulfovibrio sp. andDesulfobacter sp. which were provided with H₂/CO₂, butanol, glucose, and acetate as carbon and electron sources. A consortium of these bacteria utilizing hydrogen gave rise to relatively high corrosion rates (5.7×10^–4 mhos cm^–2) with respect to corrosion resulting from bacteria supplied with organic electron sources (0.6–1.6×10^–4 mhos cm^–2). Disproportionation of electrons between sulphate reduction and fermentation had a significant effect on the corrosion rate in the case ofDesulfovibrio. Surface examination using scanning electron microscopy coupled with electrochemical impedance spectroscopy supported the hypothesis that the corrosion rate was controlled by the relative intactness of a ferrous sulphide film in which the bacteria were embedded.     

In Situ Exposure to Low Herbicide Concentrations Affects Microbial Population Composition and Catabolic Gene Frequency in an Aerobic Shallow Aquifer

The aim of this study was to evaluate how the in situ exposure of a Danish subsurface aquifer to phenoxy acid herbicides at low concentrations (<40 μg l⁻¹) changes the microbial community composition. Sediment and groundwater samples were collected inside and outside the herbicide-exposed area and were analyzed for the presence of general microbial populations, Pseudomonas bacteria, and specific phenoxy acid degraders. Both culture-dependent and culture-independent methods were applied. The abundance of microbial phenoxy acid degraders (10⁰ to 10⁴ g⁻¹ sediment) was determined by most probable number assays, and their presence was only detected in herbicide-exposed sediments. Similarly, PCR analysis showed that the 2,4-dichlorophenoxyacetic acid degradation pathway genes tfdA and tfdB (10² to 10³ gene copies g⁻¹ sediment) were only detected in sediments from contaminated areas of the aquifer. PCR-restriction fragment length polymorphism measurements demonstrated the presence of different populations of tfd genes, suggesting that the in situ herbicide degradation was caused by the activity of a heterogeneous population of phenoxy acid degraders. The number of Pseudomonas bacteria measured by either PCR or plating on selective agar media was higher in sediments subjected to high levels of phenoxy acid. Furthermore, high numbers of CFU compared to direct counting of 4′,6-diamidino-2-phenylindole-stained cells in the microscope suggested an increased culturability of the indigenous microbial communities from acclimated sediments. The findings of this study demonstrate that continuous exposure to low herbicide concentrations can markedly change the bacterial community composition of a subsurface aquifer.