Hydra bioassay for the evaluation of chlordecone toxicity at environmental concentrations, alone or in complex mixtures with dechlorinated byproducts: experimental observations and modeling by experimental design.

In chlordecone (CLD)-contaminated soils of the French West Indies, if microbial remediation or a physicochemical remediation process, e.g., in situ chemical reduction, is implemented, concentrations of degradation byproducts, such as hydrochlordecones, are expected to increase in the ecosystems. To study their impact in mixtures with CLD, bioassays were carried out. They consisted in evaluating the regenerative capacity of hydra polyps, from a clone whose phylogenetic analysis confirmed that it belonged to the species Hydra vulgaris Pallas, 1766. Hydra gastric sections were exposed to CLD alone or CLD plus dechlorinated byproducts (CLD-BP) for 96 h to assess regeneration. Based on chromatographic analysis, the CLD-BP mix was composed of the 5-monohydrochlordecone isomer (CAS nomenclature), four dihydrochlordecone isomers, and one trihydrochlordecone isomer representing 50%, 47%, and 3% of the total chromatographic area, respectively. A total of 18 mixtures of CLD and CLD-BP were tested. Six environmental concentrations of CLD (2.10-4 µM to 4.10-2 µM) and a similar range of CLD-BP were used. Results from exposures to CLD alone showed the following: (i) a significant decrease in the regenerative capacity of hydra, except at the lowest concentration (2.10-4 µM); (ii) a concentration-independent deleterious effect. The regeneration scores obtained after the exposure to the addition of CLD-BP were not significantly different from those obtained after exposure to CLD alone. Using an experimental design, a modeling of the regeneration scores of hydra exposed to mixtures is proposed. Interpreted carefully, since they are limited to only one type of bioassay, the present results suggest that the situation in the aquatic environments should not become worse in terms of toxicity, if soil remediation programs resulting in the formation of hydrochlordecones are put in place.

Strategy to identify the causes and to solve a sludge granulation problem in methanogenic reactors: application to a full-scale plant treating cheese wastewater.

Granulation of biomass is at the basis of the operation of the most successful anaerobic systems (UASB, EGSB and IC reactors) applied worldwide for wastewater treatment. Despite of decades of studies of the biomass granulation process, it is still not fully understood and controlled. "Degranulation/lack of granulation" is a problem that occurs sometimes in anaerobic systems resulting often in heavy loss of biomass and poor treatment efficiencies or even complete reactor failure. Such a problem occurred in Mexico in two full-scale UASB reactors treating cheese wastewater. A close follow-up of the plant was performed to try to identify the factors responsible for the phenomenon. Basically, the list of possible causes to a granulation problem that were investigated can be classified amongst nutritional, i.e. related to wastewater composition (e.g. deficiency or excess of macronutrients or micronutrients, too high COD proportion due to proteins or volatile fatty acids, high ammonium, sulphate or fat concentrations), operational (excessive loading rate, sub- or over-optimal water upflow velocity) and structural (poor hydraulic design of the plant). Despite of an intensive search, the causes of the granulation problems could not be identified. The present case remains however an example of the strategy that must be followed to identify these causes and could be used as a guide for plant operators or consultants who are confronted with a similar situation independently of the type of wastewater. According to a large literature based on successful experiments at lab scale, an attempt to artificially granulate the industrial reactor biomass through the dosage of a cationic polymer was also tested but equally failed. Instead of promoting granulation, the dosage caused a heavy sludge flotation. This shows that the scaling of such a procedure from lab to real scale cannot be advised right away unless its operability at such a scale can be demonstrated.

Transfer of Pseudomonas pictorum Gray and Thornton 1928 to genus Stenotrophomonas as Stenotrophomonas pictorum comb. nov., and emended description of the genus Stenotrophomonas.

A polyphasic taxonomic approach including analysis of phenotypic, physiological and genotypic characteristics, 16S rRNA gene sequence and DNA-DNA hybridization analysis was used to determine the most consistent affiliation of Pseudomonas pictorum. Pseudomonas pictorum ATCC 23328T exhibited phenotypic traits of members of the genus Stenotrophomonas including cellular fatty acid composition, quinone and limited range of substrates that could be used. Antibiotic susceptibility and physiological characteristics were determined. The DNA G+C content was 65.7 mol%. Phylogenetic analysis revealed that the type strains of Stenotrophomonas terrae, Stenotrophomonashumi, Stenotrophomonasnitritireducens and Stenotrophomonasacidaminiphila were the nearest relatives (16S rRNA gene sequence similarity of 98.0 to 98.8 %). All the other type strains of species of the genus Stenotrophomonas showed high 16S rRNA gene sequence similarities (96.8 to 97.2 %). DNA-DNA hybridizations revealed 31.0, 32.0, 43.3 and 43.6 % reassociation between Pseudomonas pictorum ATCC 23328T and the type strains of S. terrae, S. humi, S. nitritireducens and S. acidaminiphila, respectively. Our overall results indicate that Pseudomonas pictorum should be transferred to the genus Stenotrophomonas as a novel species of this genus, Stenotrophomonas pictorum comb. nov. Since the original description of the genus Stenotrophomonaswas made with only one species (Stenotrophomonasmaltophilia), an emendation of the genus description is proposed in order to match better with the characteristics of the eleven novel species assigned to this genus since then.

Natural transformation of chlordecone into 5b-hydrochlordecone in French West Indies soils: statistical evidence for investigating long-term persistence of organic pollutants.

Chlordecone (CLD) was an organochlorine insecticide whose previous use resulted in an extensive pollution of the environment with severe health effects and social consequences. A closely related compound, 5b-hydrochlordecone (5b-hydroCLD), has been searched for and often detected in environmental matrices from the geographical area where CLD was applied. The current consensus considered that its presence was not the result of a biotic or abiotic dechlorination of CLD in these matrices but rather the consequence of its presence as impurity (synthesis by-product) in the CLD released into the environment. The aim of the present study was to determine if and to what extent degradation of CLD into 5b-hydroCLD occurred in the field. To test this hypothesis, the ratios of 5b-hydroCLD and CLD concentrations in a dataset of 810 soils collected between 2006 and 2012 in Martinique were compared to the ratios measured in 3 samples of the CLD dust commercial formulations applied in the banana fields of French West Indies (FWI) and 1 sample of the technical-grade CLD corresponding to the active ingredient used in such formulations. Soil data were processed with a hierarchical Bayesian model to account for random measurement errors and data censoring. Any pathway of CLD transformation into 5b-hydroCLD occurring over the long term in FWI soils would indeed change the ratio of 5b-hydroCLD/CLD compared to what it was in the initially applied formulations. Results showed a significant increase of the 5b-hydroCLD/CLD ratio in the soils-25 times greater in soil than in commercial formulations-which suggested that natural CLD transformation into 5b-hydroCLD over the long term occurred in these soils. Results from this study may impact future decisions for the remediation of the polluted areas.

Chlordecone retention in the fractal structure of volcanic clay.

Chlordecone (CHLD), a soil and foodstuff pollutant, as well as an environmentally persistent organochlorine insecticide, was used intensively in banana fields. The chlordecone uptake of three crops was measured for two types of polluted soils: allophanic and non-allophanic. The uptake is lower for allophanic soils even if their chlordecone content is higher than with non-allophanic soils. The fractal structure of the allophane aggregates was characterized at the nanoscale by small angle X-rays scattering, pore size distribution and transmission electron microscopy. We showed that clay microstructures should be an important physico-chemical factor governing the fate of chlordecone in the environment. Allophanic clays result in two counterintuitive findings: higher contaminant trappings yet lower contaminant availability. We propose that this specific, tortuous structure, along with its associated low accessibility, partly explains the low availability of chlordecone confined in allophanic soils. Capsule The fractal and tortuous microstructure of allophane clay favours the chlordecone retention in soils and disfavours the crop uptake.

Gibbs free energy of formation of chlordecone and potential degradation products: implications for remediation strategies and environmental fate.

Chlordecone (C(10)Cl(10)O; CAS number 143-50-0) has been used extensively as an organochlorine insecticide but is nowadays banned under The Stockholm Convention on Persistent Organic Pollutants (POPs). A search for chlordecone-respiring organisms and choosing between reductive versus oxidative remediation tools and strategies to clean up chlordecone-polluted environments would benefit from the availability of Gibbs free energy data of chlordecone and its potential dechlorination products. Presently such data are not available. Polycyclic "cage" molecules of which chlordecone is an example contain considerable strain energy. It is not a priori clear how this affects the thermodynamic properties of the chlorinated members of this unique class of compounds and to what extent redox potentials for the halogenated congeners are different from those of other aliphatic and aromatic organohalogens. We performed ab initio quantum chemical calculations to estimate Δ(f)H(m)° and Δ(f)G(m)° values of chlordecone and selected dechlorination products and used these data to calculate their Gibbs free energy and redox potential. With redox potentials in the range of 336-413 mV chlordecone has an E(o)' value similar to that of other organochlorines. The results indicate that there are no thermodynamic reasons why chlordecone-respiring or -fermenting organisms should not exist.

Multiple syntrophic interactions in a terephthalate-degrading methanogenic consortium.

Terephthalate (TA) is one of the top 50 chemicals produced worldwide. Its production results in a TA-containing wastewater that is treated by anaerobic processes through a poorly understood methanogenic syntrophy. Using metagenomics, we characterized the methanogenic consortium inside a hyper-mesophilic (that is, between mesophilic and thermophilic), TA-degrading bioreactor. We identified genes belonging to dominant Pelotomaculum species presumably involved in TA degradation through decarboxylation, dearomatization, and modified ß-oxidation to H(2)/CO(2) and acetate. These intermediates are converted to CH(4)/CO(2) by three novel hyper-mesophilic methanogens. Additional secondary syntrophic interactions were predicted in Thermotogae, Syntrophus and candidate phyla OP5 and WWE1 populations. The OP5 encodes genes capable of anaerobic autotrophic butyrate production and Thermotogae, Syntrophus and WWE1 have the genetic potential to oxidize butyrate to CO(2)/H(2) and acetate. These observations suggest that the TA-degrading consortium consists of additional syntrophic interactions beyond the standard H(2)-producing syntroph-methanogen partnership that may serve to improve community stability.

Pseudoxanthomonas mexicana sp. nov. and Pseudoxanthomonas japonensis sp. nov., isolated from diverse environments, and emended descriptions of the genus Pseudoxanthomonas Finkmann et al. 2000 and of its type species.

Three mesophilic bacteria (strains AMX 26B(T), UR374_02 and 12-3(T)) isolated respectively from an anaerobic digester, human urine and urban riverside soil were characterized. Cells were Gram-negative, motile, non-sporulating, straight to curved rods with one polar flagellum and had a strictly respiratory metabolism with O(2) as the preferential terminal electron acceptor. Phylogenetic analysis based on 16S rRNA gene sequences revealed that all strains clustered within the Xanthomonadaceae branch of the Proteobacteria. Isolates AMX 26B(T) and UR374_02 exhibited 100 % 16S rRNA gene sequence similarity and both were related to strain 12-3(T) (99.6 % similarity). The closest relative of all the isolates was Pseudoxanthomonas broegbernensis DSM 12573(T) (similarity 97.1-97.5 %), and they were equidistantly related to Xanthomonas species (95.4-96.6 %), Stenotrophomonas species (95.3-96.1 %) and Pseudoxanthomonas taiwanensis ATCC BAA-4040(T) (95.3-95.4 %). Chemotaxonomic and biochemical data (branched-chain cellular fatty acid pattern without C(13 : 0) iso 3-OH, ubiquinone with eight isoprenoid units, limited range of substrates used, ability to reduce nitrite but not nitrate with the production of N(2)O) supported their affiliation to the genus Pseudoxanthomonas. The results of DNA-DNA hybridization and/or phenotypic analysis allowed them to be differentiated from the two Pseudoxanthomonas species with validly published names and showed that strain 12-3(T) was genomically and phenotypically distinct from the other two isolates. On the basis of these results, two novel species of the genus Pseudoxanthomonas are proposed: Pseudoxanthomonas mexicana sp. nov., consisting of strains AMX 26B(T) (=ATCC 700993(T)=CIP 106674(T)=JCM 11524(T)) (type strain) and UR374_02 (=DSM 15133), and Pseudoxanthomonas japonensis sp. nov., consisting of strain 12-3(T) (=CCUG 48231(T)=CIP 107388(T)=JCM 11525(T)). The report of these two novel species leads to the emendation of the description of the genus Pseudoxanthomonas and the re-evaluation of the phenotype of P. broegbernensis DSM 12573(T) necessitates the emendation of its description.

Microbial community structure in a thermophilic anaerobic hybrid reactor degrading terephthalate.

A thermophilic terephthalate-degrading methanogenic consortium was successfully enriched for 272 days in an anaerobic hybrid reactor, and the microbial structure was characterized using terminal RFLPs, clone libraries and fluorescence in-situ hybridization with rRNA-targeted oligonucleotide probes. All the results suggested that Methanothrix thermophila-related methanogens, Desulfotomaculum-related bacterial populations in the Gram-positive low-G + C group, and OP5-related populations were the key members responsible for terephthalate degradation under thermophilic methanogenic conditions except during periods when the reactor experienced heat shock and pump failure. These perturbations caused a significant shift in bacterial population structure in sludge samples taken from the sludge bed but not from the surface of the packing materials. After system recovery, many other bacterial populations emerged, which belonged mainly to the Gram-positive low-G + C group and Cytophaga-Flexibacter-Bacteroides, as well as beta-Proteobacteria, Planctomycetes and Nitrospira. These newly emerged populations were probably also capable of degrading terephthalate in the hybrid system, but were out-competed by those bacterial populations before perturbations.

Bosea minatitlanensis sp. nov., a strictly aerobic bacterium isolated from an anaerobic digester.

A strictly aerobic, mesophilic bacterium, strain AMX 51(T), was isolated from anaerobic digester sludge. Cells were Gram-negative, motile, non-sporulating, straight to curved rods with one polar flagellum. The isolate had phenotypic traits of the genus Bosea, including cellular fatty acid and substrate utilization profiles. Physiological characteristics and antibiotic susceptibility were determined. Phylogenetic analysis revealed that strain AMX 51(T) was a member of the alpha-Proteobacteria, most closely related to Bosea thiooxidans DSM 9653(T) (similarity of 98.88 %). Methylobacterium organophilum JCM 2833(T), Methylobacterium mesophilicum JCM 2829(T), Afipia clevelandensis DSM 7315(T), Afipia felis DSM 7326(T), Afipia broomeae DSM 7327(T), Blastobacter denitrificans LMG 8443(T) and Bradyrhizobium japonicum DSM 30131(T) showed significant 16S rRNA gene sequence similarities to strain AMX 51(T). The DNA G+C composition of strain AMX 51(T) was 68.5 mol%. DNA-DNA hybridization analysis revealed 44.2 and 15.1 % relatedness between strain AMX 51(T) and the respective type strains of Bosea thiooxidans and A. felis. Overall results suggest that strain AMX 51(T) (=DSM 13099(T)=ATCC 700918(T)=CIP 106457(T)) represents a novel species of the genus Bosea; the name Bosea minatitlanensis sp. nov. is proposed.

Clostridium thiosulfatireducens sp. nov., a proteolytic, thiosulfate- and sulfur-reducing bacterium isolated from an upflow anaerobic sludge blanket (UASB) reactor.

A strictly anaerobic, gram-positive, sporulating rod (0.5-0.6 x 2.0-4.0 microm), designated strain Lup 21T, was isolated from an upflow anaerobic sludge blanket (UASB) reactor treating cheese-factory wastewater. Strain Lup 21T was motile by means of peritrichous flagella, had a G+C content of 31.4 mol% and grew optimally at 37 degrees C, pH 7.4, in the absence of NaCl. It is a heterotrophic micro-organism, utilizing proteinaceous compounds (gelatin, peptides, Casamino acids and various single amino acids) but unable to use any of the carbohydrates tested as a carbon and energy source. It reduced thiosulfate and elemental sulfur to sulfide in the presence of Casamino acids as carbon and energy sources. Acetate, butyrate, isobutyrate, isovalerate, CO2 and sulfide were end products from oxidation of gelatin and Casamino acids in the presence of thiosulfate as an electron acceptor. In the absence of thiosulfate, serine, lysine, methionine and histidine were fermented. On the basis of 16S rRNA similarity, strain Lup 21T was related to members of the low-G+C Clostridiales group, Clostridium subterminale DSM 6970T being the closest relative (with a sequence similarity of 99.4%). DNA-DNA hybridization was 56% with this species. On the basis of phenotypic, genotypic and phylogenetic characteristics, the isolate was designated as a novel species of the genus Clostridium, Clostridium thiosulfatireducens sp. nov. The type strain is strain Lup 21T (= DSM 13105T = CIP 106908T).

Sporanaerobacter acetigenes gen. nov., sp. nov., a novel acetogenic, facultatively sulfur-reducing bacterium.

A strictly anaerobic, moderately thermophilic, sporulating rod, designated strain Lup 33T, was isolated from an upflow anaerobic sludge blanket (UASB) reactor in Mexico. Strain Lup 33T possessed a few laterally inserted flagella, had a DNA G+C content of 32.2 mol % and grew optimally at pH 7.4 and 40 degrees C. Growth was observed at temperatures of up to 50 degrees C and was inhibited in the presence of 5% NaCl. Strain Lup 33T is heterotrophic and utilized some sugars, peptides and various single amino acids. Gelatin and casein were not used as energy sources. It performed the Stickland reaction and reduced elemental sulfur to sulfide. Acetate was the only fatty acid detected from glucose fermentation, whereas acetate together with isobutyrate and isovalerate were found as end products from peptone fermentation. Phylogenetically, strain Lup 33T branched with members of cluster XII of the order Clostridiales, with Clostridium hastiforme as the closest relative (similarity of 93%). On the basis of the phenotypic, genotypic and phylogenetic characteristics of the isolate, it is proposed as a novel species of a new genus, Sporanaerobacter acetigenes gen. nov., sp. nov. The type strain is strain Lup 33T (= DSM 13106T = CIP 106730T).

Stenotrophomonas acidaminiphila sp. nov., a strictly aerobic bacterium isolated from an upflow anaerobic sludge blanket (UASB) reactor.

Two of several strictly aerobic, mesophilic bacteria isolated from a lab-scale upflow anaerobic sludge blanket (UASB) reactor treating a petrochemical wastewater, strains AMX 17 and AMX 19T, were subjected to detailed taxonomic study. Cells were gram-negative, motile, non-sporulating, straight to curved rods with a polar flagellum. The isolates exhibited phenotypic traits of members of the genus Stenotrophomonas, including cellular fatty acid composition and the limited range of substrates that could be used. Sugars and many amino acids were utilized. Antibiotic susceptibility and physiological characteristics were determined. The DNA base composition was 66.9 mol% G+C. Phylogenetic analysis revealed that the nearest relatives were Stenotrophomonas maltophilia LMG 11114, Stenotrophomonas nitritireducens DSM 12575T and Pseudomonas pictorum ATCC 23328T (similarity of 98.1-98.8%). Xanthomonas species, S. maltophilia LMG 958T and Stenotrophomonas africana CIP 104854T showed high 16S rRNA sequence similarities (96.4-97.3%). The high similarity found in cellular fatty acid profiles and identical partial 16S rRNA sequences (500 bp) for strains AMX 17 and AMX 19T indicate that they belong to the same species. DNA-DNA hybridizations revealed respectively 26.7, 31, 65.8 and 43.6% homology between isolate AMX 19T and S. africana CIP 104854T, S. maltophilia CIP 60.77T, S. nitritireducens DSM 12575T and P. pictorum ATCC 23328T. These results allow the proposal of strain AMX 19T (= DSM 13117T = ATCC 700916T = CIP 106456T) as representative of a novel species of the genus Stenotrophomonas, with the name Stenotrophomonas acidaminiphila sp. nov.

Resistencia a la exposición al oxígeno de lodos anaerobios suspendidos / Oxygen exposure resistance in suspended anaerobic sludge

La tolerancia al oxígeno de una biomasa anaerobia suspendida en presencia o ausencia de un sustrato primario (sacarosa) se evaluó en términos de la recuperación de la actividad metanogénica aceticlástica especial (AME) de la biomasa anaerobia y un índice de inhibición 50 por ciento (II50) asociado a la AME. Incubada en presencia de sacarosa. La biomasa anaerobia suspendida mostró resistencia a la exposición al oxígeno; la recuperación de la AME fue >45 por ciento para [O²] iniciales <20 por ciento en el espacio gaseoso, y de 10 a 12 por ciento para [O²] iniciales >20 por ciento en el espacio gaseoso. Cuando fue incubada son fuente, la biomasa suspendida fue mucho más inhibida después de la exposición al oxígeno para [O²] iniciales >20 por ciento. El efecto inhibitorio fue descrito por un II50 elevado /28,6) en contraste con un bajo II50 (5,9) cuando se incubó en presencia de sacarosa. La tolerancia de la biomasa suspendida en este trabajo parece ser del mismo orden de la biomasa anaerobia inmovilizada (gránulos anaerobios) en condiciones de incubación en presencia de sustrato: los II50 fueron 5,9 para todos anaerobios suspendidos (sacarosa) y 5,3 y 2,4 para todos granulares incubados con acetato etanol, respectivamente. La respiración aerobia heterótrofa de los lodos anaerobio floculentos incubados con sacarosa fue cerca de 4 veces mayor que la respiración basal, y la inhibición de la AME descrita por el II50 parece seguir una relación inversa con la respiración aerobia heterótrofa. La relación inversa entre II50 y respiración aerobia heterótrofa se ajustó para datos de todos granulares en la literatura y todos floculentos de este trabajo, y sigue un modelo semi-empírico general, con un coeficiente de correlación de 0,82. Esta relación parece reforzar que uno de los mecanismos principales de protección de los consorcios anaerobios contra la inhibición por oxígeno es la respiración aerobia heterótrofa