Dehalorespiration with hexachlorobenzene and pentachlorobenzene by<Emphasis Type="Italic"> Dehalococcoides</Emphasis> sp. strain CBDB1

The chlororespiring anaerobe Dehalococcoides sp. strain CBDB1 used hexachlorobenzene and pentachlorobenzene as electron acceptors in an energy-conserving process with hydrogen as electron donor. Previous attempts to grow Dehalococcoides sp. strain CBDB1 with hexachlorobenzene or pentachlorobenzene as electron acceptors failed if these compounds were provided as solutions in hexadecane. However, Dehalococcoides sp. strain CBDB1 was able to grow with hexachlorobenzene or pentachlorobenzene when added in crystalline form directly to cultures. Growth of Dehalococcoides sp. strain CBDB1 by dehalorespiration resulted in a growth yield (Y) of 2.1±0.24 g protein/mol Cl^– released with hexachlorobenzene as electron acceptor; with pentachlorobenzene, the growth yield was 2.9±0.15 g/mol Cl^–. Hexachlorobenzene was reductively dechlorinated to pentachlorobenzene, which was converted to a mixture of 1,2,3,5- and 1,2,4,5-tetrachlorobenzene. Formation of 1,2,3,4-tetrachlorobenzene was not detected. The final end-products of hexachlorobenzene and pentachlorobenzene dechlorination were 1,3,5-trichlorobenzene, 1,3- and 1,4-dichlorobenzene, which were formed in a ratio of about 3:2:5. As reported previously, Dehalococcoides sp. strain CBDB1 converted 1,2,3,5-tetrachlorobenzene exclusively to 1,3,5-trichlorobenzene, and 1,2,4,5-tetrachlorobenzene exclusively to 1,2,4-trichlorobenzene. The organism therefore catalyzes two different pathways to dechlorinate highly chlorinated benzenes. In the route leading to 1,3,5-trichlorobenzene, only doubly flanked chlorine substituents were removed, while in the route leading to 1,3-and 1,4-dichlorobenzene via 1,2,4-trichlorobenzene singly flanked chlorine substituents were also removed. Reductive dehalogenase activity measurements using whole cells pregrown with different chlorobenzene congeners as electron acceptors indicated that different reductive dehalogenases might be induced by the different electron acceptors. To our knowledge, this is the first report describing reductive dechlorination of hexachlorobenzene and pentachlorobenzene via dehalorespiration by a pure bacterial culture.     

Reductive dechlorination of tetrachloroethene by a stepwise catalysis of different organohalide respiring bacteria and reductive dehalogenases

The enrichment culture SL2 dechlorinating tetrachloroethene (PCE) to ethene with strong trichloroethene (TCE) accumulation prior to cis-1,2-dichloroethene (cis-DCE) formation was analyzed for the presence of organohalide respiring bacteria and reductive dehalogenase genes (rdhA). Sulfurospirillum-affiliated bacteria were identified to be involved in PCE dechlorination to cis-DCE whereas “Dehalococcoides”-affiliated bacteria mainly dechlorinated cis-DCE to ethene. Two rdhA genes highly similar to tetrachloroethene reductive dehalogenase genes (pceA) of S. multivorans and S. halorespirans were present as well as an rdhA gene very similar to the trichloroethene reductive dehalogenase gene (tceA) of “Dehalococcoides ethenogenes” strain 195. A single strand conformation polymorphism (SSCP) method was developed allowing the simultaneous detection of the three rdhA genes and the estimation of their abundance. SSCP analysis of different SL2 cultures showed that one pceA gene was expressed during PCE dechlorination whereas the second was expressed during TCE dechlorination. The tceA gene was involved in cis-DCE dechlorination to ethene. Analysis of the internal transcribed spacer region between the 16S and 23S rRNA genes revealed two distinct sequences originating from Sulfurospirillum suggesting that two Sulfurospirillum populations were present in SL2. Whether each Sulfurospirillum population was catalyzing a different dechlorination step could however not be elucidated.     

Complete Detoxification of Vinyl Chloride by an Anaerobic Enrichment Culture and Identification of the Reductively Dechlorinating Population as a Dehalococcoides Species

A major obstacle in the implementation of the reductive dechlorination process at chloroethene-contaminated sites is the accumulation of the intermediate vinyl chloride (VC), a proven human carcinogen. To shed light on the microbiology involved in the final critical dechlorination step, a sediment-free, nonmethanogenic, VC-dechlorinating enrichment culture was derived from tetrachloroethene (PCE)-to-ethene-dechlorinating microcosms established with material from the chloroethene-contaminated Bachman Road site aquifer in Oscoda, Mich. After 40 consecutive transfers in defined, reduced mineral salts medium amended with VC, the culture lost the ability to use PCE and trichloroethene (TCE) as metabolic electron acceptors. PCE and TCE dechlorination occurred in the presence of VC, presumably in a cometabolic process. Enrichment cultures supplied with lactate or pyruvate as electron donor dechlorinated VC to ethene at rates up to 54 μmol liter⁻¹day⁻¹, and dichloroethenes (DCEs) were dechlorinated at about 50% of this rate. The half-saturation constant (K_S) for VC was 5.8 μM, which was about one-third lower than the concentrations determined for cis-DCE and trans-DCE. Similar VC dechlorination rates were observed at temperatures between 22 and 30°C, and negligible dechlorination occurred at 4 and 35°C. Reductive dechlorination in medium amended with ampicillin was strictly dependent on H₂ as electron donor. VC-dechlorinating cultures consumed H₂ to threshold concentrations of 0.12 ppm by volume. 16S rRNA gene-based tools identified a Dehalococcoides population, and Dehalococcoides-targeted quantitative real-time PCR confirmed VC-dependent growth of this population. These findings demonstrate that Dehalococcoides populations exist that use DCEs and VC but not PCE or TCE as metabolic electron acceptors.     

Quantitative PCR Confirms Purity of Strain GT, a Novel Trichloroethene-to-Ethene-Respiring Dehalococcoides Isolate

A novel Dehalococcoides isolate capable of metabolic trichloroethene (TCE)-to-ethene reductive dechlorination was obtained from contaminated aquifer material. Growth studies and 16S rRNA gene-targeted analyses suggested culture purity; however, the careful quantitative analysis of Dehalococcoides 16S rRNA gene and chloroethene reductive dehalogenase gene (i.e., vcrA, tceA, and bvcA) copy numbers revealed that the culture consisted of multiple, distinct Dehalococcoides organisms. Subsequent transfers, along with quantitative PCR monitoring, yielded isolate GT, possessing only vcrA. These findings suggest that commonly used qualitative 16S rRNA gene-based procedures are insufficient to verify purity of Dehalococcoides cultures. Phylogenetic analysis revealed that strain GT is affiliated with the Pinellas group of the Dehalococcoides cluster and shares 100% 16S rRNA gene sequence identity with two other Dehalococcoides isolates, strain FL2 and strain CBDB1. The new isolate is distinct, as it respires the priority pollutants TCE, cis-1,2-dichloroethene (cis-DCE), 1,1-dichloroethene (1,1-DCE), and vinyl chloride (VC), thereby producing innocuous ethene and inorganic chloride. Strain GT dechlorinated TCE, cis-DCE, 1,1-DCE, and VC to ethene at rates up to 40, 41, 62, and 127 μmol liter⁻¹ day⁻¹, respectively, but failed to dechlorinate PCE. Hydrogen was the required electron donor, which was depleted to a consumption threshold concentration of 0.76 ± 0.13 nM with VC as the electron acceptor. In contrast to the known TCE dechlorinating isolates, strain GT dechlorinated TCE to ethene with very little formation of chlorinated intermediates, suggesting that this type of organism avoids the commonly observed accumulation of cis-DCE and VC during TCE-to-ethene dechlorination.     

Tetrachloroethene and 3-chlorobenzoate dechlorination activities are co-induced inDesulfomonile tiedjei DCB-1

Desulfomonile tiedjei, a strict anaerobe capable of reductively dechlorinating 3-chlorobenzoate, also dechlorinates tetrachloroethene and trichloroethene. It is not known, however, if the aryl and aliphatic dechlorination activities are catalyzed by the same enzymatic system. Cultures induced for 3-chlorobenzoate activity dechlorinated tetrachloroethene and trichloroethene to lower chlorinated products while uninduced parallel cultures did not dechlorinate either substrate. The observed rate of PCE dechlorination in induced cultures was 22 µmol h^–1 g protein^–1, which is considerably faster than previous rates obtained with defined cultures of this organism. These results show that both dechlorination activities are co-induced and therefore, that the dechlorination mechanisms may share at least some components.Abbreviations PCE tetrachloroethene - TCE trichloroethene - cis-DCE cis-dichloroethene - trans-DCE trans-dichloroethene - 3FBz 3-fluorobenzoate - 3ClBz 3-chlorobenzoate     

Discrimination of Multiple Dehalococcoides Strains in a Trichloroethene Enrichment by Quantification of Their Reductive Dehalogenase Genes

While many anaerobic microbial communities are capable of reductively dechlorinating tetrachloroethene (PCE) and trichloroethene (TCE) to dichloroethene (DCE), vinyl chloride (VC), and finally ethene, the accumulation of the highly toxic intermediates, cis-DCE (cDCE) and VC, presents a challenge for bioremediation processes. Members of the genus Dehalococcoides are apparently solely responsible for dechlorination beyond DCE, but isolates of Dehalococcoides each metabolize only a subset of PCE dechlorination intermediates and the interactions among distinct Dehalococcoides strains that result in complete dechlorination are not well understood. Here we apply quantitative PCR to 16S rRNA and reductase gene sequences to discriminate and track Dehalococcoides strains in a TCE enrichment derived from soil taken from the Alameda Naval Air Station (ANAS) using a four-gene plasmid standard. This standard increased experimental accuracy such that 16S rRNA and summed reductase gene copy numbers matched to within 10%. The ANAS culture was found to contain only a single Dehalococcoides 16S rRNA gene sequence, matching that of D. ethenogenes 195, but both the vcrA and tceA reductive dehalogenase genes. Quantities of these two genes in the enrichment summed to the quantity of the Dehalococcoides 16S rRNA gene. Further, between ANAS subcultures enriched on TCE, cDCE, or VC, the relative copy number of the two dehalogenases shifted 14-fold, indicating that the genes are present in two different Dehalococcoides strains. Comparison of cell yields in VC-, cDCE-, and TCE-enriched subcultures suggests that the tceA-containing strain is responsible for nearly all of the TCE and cDCE metabolism in ANAS, whereas the vcrA-containing strain is responsible for all of the VC metabolism.     

Complete degradation of tetrachloroethene by combining anaerobic dechlorinating and aerobic methanotrophic enrichment cultures

 Degradation of tetrachloroethene (perchloroethylene, PCE) was investigated by combining the metabolic abilities of anaerobic bacteria, capable of reductive dechlorination of PCE, with those of aerobic methanotrophic bacteria, capable of co-metabolic degradation of the less-chlorinated ethenes formed by reductive dechlorination of PCE. Anaerobic communities reductively dechlorinating PCE, trichloroethene (TCE) and dichloroethenes were enriched from various sources. The maximum rates of dechlorination observed for various chloroethenes in these batch enrichments were: PCE to TCE (341 μmol l^-1 day^-1), TCE to cis-dichloroethene (159 μmol l^-1 day^-1), cis-dichloroethene to chloroethene (99 μmol l^-1 day^-1) and trans-dichloroethene to chloroethene (22 μmol l^-1 day^-1). A mixture of these enrichments was inoculated into an anoxic fixed-bed upflow column. In this column PCE was converted mainly into cis-1, 2-dichloroethene, small amounts of TCE and chloroethene, and chloride. Enrichments of aerobic methanotrophic bacteria were grown in an oxic fixed-bed downflow column. Less-chlorinated ethenes, formed in the anoxic column, were further metabolized in this oxic methanotrophic column. On the basis of analysis of chloride production and the disappearance of chlorinated ethenes it was demonstrated that complete degradation of PCE was possible by combining these two columns. Operation of the two-column system under various process conditions indicated that the sensitivity of the methanotrophic bacteria to chlorinated intermediates represented the bottle-neck in the sequential anoxic/oxic degradation process of PCE. Received: 24 October 1994 / Received revision: 20 January 1995 / Accepted: 23 January 1995     

Detection and Quantification of Dehalococcoides-Related Bacteria in a Chlorinated Ethene-Contaminated Aquifer Undergoing Natural Attenuation

Detection and quantification of bacteria related to Dehalococcoides is essential for the development of effective remediation strategies for tetrachloroethene (PCE)-contaminated sites. In this study, the authors applied three methods for quantifying Dehalococcoides-like bacteria in a PCE-contaminated aquifer undergoing natural attenuation in Grenchen, Switzerland: a catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) protocol, a competitive nested polymerase chain reaction (PCR) approach, and a direct PCR end point quantification with external standards. For the investigated aquifer, multiple lines of evidence indicated that reductive dechlorination (and likely dehalorespiration) was an active process. Both PCR-based quantification methods indicated that low numbers of mostly sediment-bound Dehalococcoides were present in the contaminated zone of the Grenchen aquifer. Estimates based on the quantitative PCR methods ranged from 2.1 × 10⁷ to 1.5 × 10⁸ sediment-bound Dehalococcoides 16S rRNA gene copies per liter of aquifer volume. In contrast, the liquid phase only contained between 8 and 80 copies per liter aquifer volume. CARD-FISH was not sensitive enough for the quantification of Dehalococcoides cell numbers in this aquifer. Cloning and sequencing of the PCR products revealed the presence of sequences closely related to Dehalococcoides isolates such as D. ethenogenes and Dehalococcoides sp. BAV1. An apparently abundant group (termed “Grenchen Cluster”) of sequences more distantly related to Dehalococcoides was also identified, so far without cultured representatives.     

Molecular characterization of microbial populations at two sites with differing reductive dechlorination abilities

This study compares three molecular techniques, including terminal restriction fragment length polymorphism (T-RFLP), RFLP analysis with clone sequencing, and quantitative PCR (Q-PCR) for surveying differences in microbial communities at two contaminated field sites that exhibit dissimilar chlorinated solvent degradation activities. At the Idaho National Engineering and Environmental Laboratory (INEEL), trichloroethene (TCE) was completely converted to ethene during biostimulation with lactate. At Seal Beach, California, perchloroethene (PCE) was degraded only to cis-dichloroethene (cDCE) during biostimulation but was degraded to ethene after bioaugmentation with a dechlorinating culture containing Dehalococcoides strains. T-RFLP analysis showed that microbial community composition differed significantly between the two sites, but was similar within each site among wells that had low or no electron donor exposure. Analysis of INEEL clone libraries by RFLP with clone sequencing revealed a complex microbial population but did not identify any Dehalococcoides strains. Q-PCR targeting the 16S rRNA gene of Dehalococcoides strains – known for their unique capability to dechlorinate solvents completely to ethene – revealed a significant population at INEEL, but no detectable population at Seal Beach prior to bioaugmentation. Detection of Dehalococcoides by Q-PCR correlated with observed dechlorination activity and ethene production at both sites. Q-PCR showed that Dehalococcoides was present in even the pristine well at INEEL, suggesting that the difference in dechlorination ability at the two sites was due to the initial absence of this genus at Seal Beach. Of the techniques tested, Q-PCR quantification of specific dechlorinating species provided the most effective and direct prediction of community dechlorinating potential.     

Characterization of microbial community structure and population dynamics of tetrachloroethene-dechlorinating tidal mudflat communities

Tetrachloroethene (PCE) and trichloroethene (TCE) are common groundwater contaminants that also impact tidal flats, especially near urban and industrial areas. However, very little is known about dechlorinating microbial communities in tidal flats. Titanium pyrosequencing, 16S rRNA gene clone libraries, and dechlorinator-targeted quantitative real-time PCR (qPCR) characterized reductive dechlorinating activities and populations in tidal flat sediments collected from South Korea’s central west coast near Kangwha. In microcosms established with surface sediments, PCE dechlorination to TCE began within 10 days and 100% of the initial amount of PCE was converted to TCE after 37 days. cis-1,2-Dichloroethene (cis-DCE) was observed as dechlorination end product in microcosms containing sediments collected from deeper zones (i.e., 35–40 cm below ground surface). Pyrosequencing of bacterial 16S rRNA genes and 16S rRNA gene-targeted qPCR results revealed Desulfuromonas michiganensis-like populations predominanted in both TCE and cis-DCE producing microcosms. Other abundant groups included Desulfuromonas thiophila and Pelobacter acidigallici-like populations in the surface sediment microcosms, and Desulfovibrio dechloracetivorans and Fusibacter paucivorans-like populations in the deeper sediment microcosms. Dehalococcoides spp. populations were not detected in these sediments before and after incubation with PCE. The results suggest that tidal flats harbor novel, salt-tolerant dechlorinating populations and that titanium pyrosequencing provides more detailed insight into community structure dynamics of the dechlorinating microcosms than conventional 16S rRNA gene sequencing or fingerprinting methods.     

Effect of chloroethene concentrations and granular activated carbon on reductive dechlorination rates and growth of Dehalococcoides spp

This study focused on the investigation of (i) the tetrachloroethene (PCE) toxicity threshold of a reductively dechlorinating mixed culture containing Dehalococcoides spp., (ii) the adsorption of PCE on different types of granular activated carbon (GAC), and (iii) the bioavailability and reductive dechlorination in the presence of GAC. The abundance of Dehalococcoides spp. detected by quantitative real-time polymerase chain reaction (qPCR) was found to increase by 2-4 orders of magnitude during degradation of PCE. No degradation occurred at dissolved concentrations beyond 420 μM (70 mg/L). Different adsorption isotherms were determined for thermally and chemically activated carbons. The addition of GAC to biological assays reduced the dissolved PCE concentration below the toxicity threshold. The combination of microbial reductive dechlorination with GAC adsorption proved to be a promising method for remediation of groundwater contaminated by high concentrations of chloroethenes.     

Isolation and characterization of Dehalospirillum multivorans gen. nov., sp. nov., a tetrachloroethene-utilizing,strictly anaerobic bacterium

A strictly anaerobic bacterium dechlorinating tetrachloroethene (perchloroethylene, PCE) via trichloroethene (TCE) to cis-1,2-dichloroethene (DCE) was isolated from activated sludge with pyruvate plus PCE as energy substrates. The organism, called Dehalospirillum multivorans, is a gram-negative spirillum that does not form spores. The G+C content of the DNA was 41.5 mol%. According to 16S rRNA gene sequence analysis, D. multivorans represents a new genus and a new species belonging to the epsilon subdivision of Proteobacteria. Quinones, cytochromes b and c, and corrinoids were extracted from the cells. D. multivorans grew in defined medium with PCE and H₂ as sole energy sources and acetate as carbon source; the growth yield under these conditions was 1.4g of cell protein per mol chloride released. Alternatively to PCE, fumarate and nitrate could serve as electron acceptors; sulfate could not replace fumarate, nitrate, or PCE in this respect. In addition to H₂, the organism utilized a variety of electron donors for dechlorination (pyruvate, lactate, ethanol, formate, glycerol). Upon growth on pyruvate plus PCE, the main fermentation products formed were acetatc, lactate, DCE, and H₂. At optimal pH (7.3–7.6) and temperature (30°C), and in the presence of pyruvate (20mM) and PCE (160M), a dechlorination rate of about 50 nmol min^-1 (mg cell protein)^-1 and a doubling time of about 2.5h were obtained with growing cultures. The ability to reduce PCE to DCE appears to be constitutive under the experimental conditions applied since cultures growing in the absence of PCE for several generations immediately started dechlorination when transferred to a medium containing PCE. The organism may be useful for bioremediation of environments polluted with tetrachloroethene.Abbreviations PCE Perchloroethylene, tetrachloroethene - TCE Trichloroethene - DCE cis-1,2-Dichloroethene - CHC Chlorinated hydrocarbon     

Effects of Polychlorinated Biphenyl Congener Concentration and Sediment Supplementation on Rates of Methanogenesis and 2,3,6-Trichlorobiphenyl Dechlorination in an Anaerobic Enrichment

We have employed a method of enrichment that allows us to significantly increase the rate of reductive polychlorinated biphenyl (PCB) dechlorination. This method shortens the time required to investigate the effects that culture conditions have on dechlorination and provides an estimate of the potential activity of the PCB-dechlorinating anaerobes. The periodic supplementation of sterile sediment and PCB produced an enhanced, measurable, and sustained rate of dechlorination. We observed volumetric rates of the dechlorination of 2,3,6-trichlorobiphenyl (2,3,6-CB) to 2,6-dichlorobiphenyl (2,6-CB) of more than 300 μmol liter^-1 day^-1 when the cultures were supplemented daily. A calculation of this activity that is based on an estimate of the number of dechlorinating anaerobes present indicates that 1.13 pmol of 2,3,6-CB was dechlorinated to 2,6-CB day^-1 bacterial cell^-1. This rate is similar to that of the reductive dechlorination of 3-chlorobenzoate by Desulfomonile tiedjei. Methanogenesis declined from 585.3 to 125.9 μmol of CH₄ liter^-1 day^-1, while dechlorination increased from 8.2 to 346.0 μmol of 2,3,6-CB dechlorinated to 2,6-CB liter^-1 day^-1.     

Energy requirements of the red kangaroo (<Emphasis Type="Italic">Macropus rufus</Emphasis>): impacts of age,growth and body size in a large desert-dwelling herbivore.

Generally, young growing mammals have resting metabolic rates (RMRs) that are proportionally greater than those of adult animals. This is seen in the red kangaroo (Macropus rufus), a large (>20 kg) herbivorous marsupial common to arid and semi-arid inland Australia. Juvenile red kangaroos have RMRs 1.5–1.6 times those expected for adult marsupials of an equivalent body mass. When fed high-quality chopped lucerne hay, young-at-foot (YAF) kangaroos, which have permanently left the mother's pouch but are still sucking, and recently weaned red kangaroos had digestible energy intakes of 641±27 kJ kg^–0.75 day^–1 and 677±26 kJ kg^–0.75 day^–1, respectively, significantly higher than the 385±37 kJ kg^–0.75 day^–1 ingested by mature, non-lactating females. However, YAF and weaned red kangaroos had maintenance energy requirements (MERs) that were not significantly higher than those of mature, non-lactating females, the values ranging between 384 kJ kg^–0.75 day^–1 and 390 kJ kg^–0.75 day^–1 digestible energy. Importantly, the MER of mature female red kangaroos was 84% of that previously reported for similarly sized, but still growing, male red kangaroos. Growth was the main factor affecting the proportionally higher energy requirements of the juvenile red kangaroos relative to non-reproductive mature females. On a good quality diet, juvenile red kangaroos from permanent pouch exit until shortly after weaning (ca. 220–400 days) had average growth rates of 55 g body mass day^–1. At this level of growth, juveniles had total daily digestible energy requirements (i.e. MER plus growth energy requirements) that were 1.7–1.8 times the MER of mature, non-reproductive females. Our data suggest that the proportionally higher RMR of juvenile red kangaroos is largely explained by the additional energy needed for growth. Energy contents of the tissue gained by the YAF and weaned red kangaroos during growth were estimated to be 5.3 kJ g^–1, within the range found for most young growing mammals.Abbreviations BMR basal metabolic rate - DEI digestible energy intake - MER maintenance energy requirement - MER_g maintenance plus growth energy requirement - PPE permanent pouch exit - RMR resting metabolic rate - YAF young-at-foot Communicated by I.D. Hume     

Field metabolic rate and water turnover of the numbat (<Emphasis Type="Italic">Myrmecobius fasciatus</Emphasis>)

The numbat (Myrmecobius fasciatus) is a diurnal and exclusively termitivorous marsupial. This study examines interrelationships between diet, metabolic rate and water turnover for wild, free-living numbats. The numbats (488±20.8 g) remained in mass balance during the study. Their basal metabolic rate (BMR) was 3.6 l CO₂ day^–1, while their field metabolic rate (FMR) was 10.8±1.22 l CO₂ day^–1 (269±30.5 kJ day^–1). The ratio FMR/BMR was 3±0.3 for numbats. We suggest that the most accurate way to predict the FMR of marsupials is from the regression log FMR=0.852 log BMR+0.767; (r²=0.97). The FMR of the numbat was lower than, but not significantly different from, that of a generalised marsupial, both before (76%) and after (62–69%) correction for the significant effect of phylogeny on FMR. However the numbat's FMR is more comparable with that of other arid-habitat Australia marsupials (98–135%), for which the regression relating mass and FMR is significantly lower than for nonarid-habitat marsupials, independent of phylogeny. The field water turnover rate (FWTR) of free-living numbats (84.1 ml H₂O day^–1) was highly correlated with FMR, and was typical (89–98%) of that for an arid-habitat marsupial after phylogenetic correction. The higher than expected water economy index for the numbat (FWTR/FMR=0.3±0.03) suggests that either the numbats were drinking during the study, the water content of their diet was high, or the digestibility of their termite diet was low. Habitat and phylogenetic influences on BMR and FMR appear to have pre-adapted the numbat to a low-energy termitivorous niche.Abbreviations BMR basal metabolic rate - FMR field metabolic rate - EWL evaporative water loss - FWTR field water turnover rate - MR metabolic rate - PVR phylogenetic vector regression - RER respiratory exchange ratio - T_a ambient temperature - T_b body temperature - TBW total body water - CO₂ rate of carbon dioxide production - O₂ rate of oxygen consumption - WEI water economy index - WER water efflux rate - WIR water influx rateCommunicated by I.D. Hume     

Effect of prey stage on life-table attributes of a genetically manipulated strain ofMetaseiulus occidentalis (Acari: Phytoseiidae)

Metaseiulus occidentalis females from the carbaryl-organophosphate-sulfur resistant strain (COS) lived longer (25.3 days versus 19.7 days), had a higher total fecundity (43.8 eggs female^–1 versus 33.6 eggs female^–1) and a higher daily fecundity rate (2.4 eggs female^–1 day^–1 versus 2.0 eggs female^–1 day^–1), and exhibited a higher intrinsic rate of increase (0.243 individuals female^–1 day^–1 versus 0.182 individuals female^–1 day^–1) and shorter generation time (13.9 days versus 17.0 days), at 24–28°C, 47–56%rh under continuous fluorescent light, when reared on a diet of 0–48-h-old eggs rather than a diet of mixed actives ofTetranychus pacificus McGregor on bean leaf disks. The sexratio of the progeny was female-biased for both diets, 2.1 females to 1 male forM. occidentalis reared on eggs and 2.0:1 : forM. occidentalis reared on mixed actives, suggesting that diet influences sex-ratio in some unknown way.There was no significant difference in oviposition rates for repeatedly-mated and once-matedM. occidentalis females reared on a diet of younger (0–24-h old) eggs compared to a diet of older (72–96-h old) eggs ofT. pacificus.The COS strain ofM. occidentalis exhibited life-table parameters comparable to the other strains reported in the literature, suggesting that the reproductive attributes of this acarine predator were not reduced as a result of artificial laboratory selection. Diet, a biotic factor, produced substantial differences in life-table parameters, suggesting that this factor can influence conclusions regarding the potential efficacy of biological control agents.     

Sequential kinetic parameter estimation of anaerobic fluidized bed bioreactor using an optimization technique

Mathematical model parameters for the methanogenic degradation of propylene glycol were estimated in a sequential manner by means of an optimization technique. Model parameters determined from an initial experimental data set using one bioreactor were then verified with the results from a second bioreactor. The proposed methodology is a useful tool to obtain model parameters for continuous flow reactors with completely mixed regime. Abbrevations: S – substrate concentration (mg COD l^–1); S _in – influent substrate concentration (mg COD l^–1); D _L – dilution rate (day^–1); – stoichiometric coefficients (ND); nx – number of microbial species (ND); X _S – fixed biomass concentration (mg biomass l^–1); X _L – suspended biomass concentration of (mg biomass l^–1); k _d – decay rate of biomass (day^–1); b _S – specific detachment rate of biofilm (day^–1); – specific growth rate of biomass (day^–1); _m – maximum specific growth rate of biomass (day^–1); K _S – half saturation constant (mg COD l^–1); K _I – inhibition constant (mg COD l^–1).     

Effects of electron donor and microbial concentration on the enhanced dechlorination of carbon tetrachloride by anaerobic consortia

 Reductive dechlorination of carbon tetra-chloride (CCl₄) by anaerobic bacterial communities from anaerobic digester sludge with the amendment of low concentrations of electron donors and microorganisms was undertaken to evaluate the influence of electron donors and microbial concentration on the rate of dechlorination of CCl₄. Humic acid, acetate, and glucose were selected to examine the feasibility of the electron donor with respect to the remediation of a contaminated subsurface. The addition of an electron donor and microorganisms significantly enhanced the dechlorination rate of carbon tetrachloride. The addition of an electron donor increased the cell numbers of anaerobic consortia, thereby increasing the rate of dechlorination. Glucose was a better electron donor than acetate and humic acid under reducing environments. The pseudo-first-order degradation rate constants of CCl₄ ranged from 0.0057 day^-1 to 0.135 day^-1, depending on the conditions of the electron donor and biomass supplemented. Furthermore, the addition of the electron donor in the batches amended with 0.56 mg volatile suspended solids (VSS)/l biomass had a higher enhanced efficiency than those with 1.7 mg VSS/l biomass. These results suggest that there is a potential for stimulating the dechlorinating capability of anaerobic consortia to remedy the chlorinated hydrocarbons in the oligotrophic environment if the conditions of the supplementing electron donor are properly selected. Received: 14 August 1995/Received last revision: 15 March 1996/Accepted: 15 April 1996     

Benzoate-driven dehalogenation of chlorinated ethenes in microbial cultures from a contaminated aquifer

Microbial dehalogenation of tetrachloroethene (PCE) and cis-dichloroethene (cis-DCE) was studied in cultures from a continuous stirred tank reactor initially inoculated with aquifer material from a PCE-contaminated site. Cultures amended with hydrogen and acetate readily dechlorinated PCE and cis-DCE; however, this transformation was incomplete and resulted in the accumulation of chlorinated intermediates and only small amounts of ethene within 60 days of incubation. Conversely, microbial PCE and cis-DCE dechlorination in cultures with benzoate and acetate resulted in the complete transformation to ethene within 30 days. Community fingerprinting by denaturing gradient gel electrophoresis (DGGE) revealed the predominance of phylotypes closely affiliated with Desulfitobacterium, Dehalococcoides, and Syntrophus species. The Dehalococcoides culture VZ, obtained from small whitish colonies in cis-DCE dechlorinating agarose cultures, revealed an irregular cell diameter between 200 and 500 nm, and a spherical or biconcave disk-shaped morphology. These organisms were identified as responsible for the dechlorination of cis-DCE to ethene in the PCE-dechlorinating consortia, operating together with the Desulfitobacterium as PCE-to-cis-DCE dehalogenating bacterium and with a Syntrophus species as potential hydrogen-producing partner in cultures with benzoate. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Ethanol production by immobilized whole cells ofZymomonas mobilis in a continuous flow expanded bed bioreactor and a continuous flow stirred tank bioreactor

Continuous ethanol fermentation by immobilized whole cells ofZymomonas mobilis was investigated in an expanded bed bioreactor and in a continuous stirred tank reactor at glucose concentrations of 100, 150 and 200 g L^–1. The effect of different dilution rates on ethanol production by immobilized whole cells ofZymomonas mobilis was studied in both reactors. The maximum ethanol productivity attained was 21 g L^–1 h^–1 at a dilution rate of 0.36 h^–1 with 150 g glucose L^–1 in the continuous expanded bed bioreactor. The conversion of glucose to ethanol was independent of the glucose concentration in both reactors.