Active prokaryotic population dynamics exhibit high correlation to reactor performance during methane production from acid hydrolysates of Agave tequilana var. azul bagasse.

AIMS: The purpose of this study was to apply cDNA approach for the characterization of active prokaryotic community to understand microbial scenarios and performance of an AnSBR digester fed with acid hydrolysates of Agave tequilana var. azul bagasse (ATAB). METHODS AND RESULTS: The digester was implemented for methane production under organic loading rate (OLR) disturbances to correlate physicochemical variables with changes in abundance, diversity and population dynamics of active Bacteria and Archaea by principal components analysis (PCA). Results indicated that methane yield increased as well as active syntrophic relationships for interspecies hydrogen/formate (Anaerolinaceae-Methanobacterium beijingense) and acetate (Anaerolinaceae-Methanosaeta concilii) transfers at 8 g-COD l-1  day-1 . However, methane yield was negatively affected at 16 g-COD l-1  day-1 due to the competition for acetate by active Desulfovibrio marrakechensis and volatile fatty acids inhibition. CONCLUSIONS: Microbial scenarios obtained by PCA correlations indicated that methane production from acid hydrolysates of ATAB was feasible at 8 g-COD l-1  day-1 . The digester operation at higher OLR only favoured methanogenesis by the hydrogenotrophic pathway. SIGNIFICANCE AND IMPACT OF THE STUDY: Only cDNA analysis showed Archaea population dynamics, exhibiting high correlation with physicochemical variables towards the understanding of the methanogenic digester performance during OLR disturbances.

Impact of storage conditions on fungal community composition of green coffee beans Coffea arabica L. stored in jute sacks during 1 year.

AIMS: The purpose of this study was to evaluate the effect of warehouse storage conditions on the composition of the fungal community of green coffee beans (GCB) that were stored in jute sacks for 1 year. METHODS AND RESULTS: Molecular characterization of the fungal community composition and population dynamics obtained by Q-PCR, CE-SSCP (Simpson's diversity index D) and Illumina MiSeq sequencing indicated that Saccharomycetales dominated during the first 6 months of storage period with species as Meyerozyma guilliermondii and Pichia kluyveri. However, after 6 months of storage, the filamentous genus Wallemia became dominant. Principal components analysis correlated this fungal dynamic with storage conditions and other variables as chromaticity (colour), water activity, moisture content, reducing sugars concentration, fungal infection and ochratoxin A production. CONCLUSIONS: This study demonstrated that GCB stored in jute sacks after 6 months of storage lead to fungal population dynamics, decreased chromaticity in GCB by bleaching and, then, affected overall quality. SIGNIFICANCE AND IMPACT OF THE STUDY: Storage plays an important role in the quality evolution during the handling of the GCB. In this context, the composition of the microbial community could be considered a marker to assess the trade value of the coffee beans.

Biodegradation of polycyclic aromatic hydrocarbons by an acidophilic Stenotrophomonas maltophilia strain AJH1 isolated from a mineral mining site in Saudi Arabia.

The present study aims at analyzing the degradation of polycyclic aromatic hydrocarbons (PAHs) at acidic conditions (pH = 2) by acidophilic Stenotrophomonas maltophilia strain AJH1 (KU664513). The strain AJH1 was obtained from an enrichment culture obtained from soil samples of mining area in the presence of PAH as sole sources of carbon and energy. Strain AJH1was able to degrade low (anthracene, phenanthrene, naphthalene, fluorene) and high (pyrene, benzo(e)pyrene and benzo(k)fluoranthene) molecular weight PAHs in acidophilic mineral salt medium at pH 2, with removal rates of up to 95% (LMW PAH) and 80% (HMW PAH), respectively. In addition, strain AJH1 treated petroleum wastewater with 89 ± 1.1% COD removal under acidic condition (pH 2) in a continuously stirred reactor. Acidophilic S. maltophilia strain AJH1, hence holds the promise as an effective degrader for biological treatment of PAHs contaminated wastewater at acidic pH.

Dynamics of antibiotic resistance genes and presence of putative pathogens during ambient temperature anaerobic digestion.

AIMS: This study was focused on evaluating the persistency of antimicrobial resistance (AR) genes and putative pathogenic bacteria in an anaerobic digesters operating at mesophilic ambient temperature, in two different year seasons: summer and winter. METHODS AND RESULTS: Abundance and dynamic of AR genes encoding resistance to macrolides (ermB), aminoglycosides (aphA2) and beta-lactams (blaTEM -1 ) and persistency of potentially pathogenic bacteria in pilot-scale anaerobic digesters were investigated. AR genes were determined in the influent and effluent in both conditions. Overall, after 60 days, reduction was observed for all evaluated genes. However, during the summer, anaerobic digestion was more related to the gene reduction as compared to winter. Persistency of potentially pathogenic bacteria was also evaluated by metagenomic analyses compared to an in-house created database. Clostridium, Acinetobacter and Stenotrophomonas were the most identified. CONCLUSIONS: Overall, considering the mesophilic ambient temperature during anaerobic digestion (summer and winter), a decrease in pathogenic bacteria detection through metagenomic analysis and AR genes is reported. Although the mesophilic anaerobic digestion has been efficient, the results may suggest medically important bacteria and AR genes persistency during the process. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report to show AR gene dynamics and persistency of potentially pathogenic bacteria through metagenomic approach in cattle manure ambient temperature anaerobic digestion.

Anaerobic treatment of tequila vinasses under seasonal operating conditions: start-up, normal operation and restart-up after a long stop and starvation period.

This study examines the performance of an anaerobic fixed-film bioreactor under seasonal operating conditions prevailing in medium and small size Tequila factories: start-up, normal operation and particularly, during the restart-up after a long stop and starvation period. The proposed start-up procedure attained a stable biofilm in a rather short period (28 days) despite unbalanced COD/N/P ratio and the use of non-acclimated inoculum. The bioreactor was restarted-up after being shut down for 6 months during which the inoculum starved. Even when biofilm detachment and bioreactor clogging were detected at the very beginning of restart-up, results show that the bioreactor performed better as higher COD removal and methane yield were attained. CE-SSCP and Q-PCR analyses, conducted on the biofilm prokaryotic communities for each operating condition, confirmed that the high COD removal results after the bioreactor clogging and the severe starvation period were mainly due to the stable archaeal and resilient bacterial populations.

Stability of airborne microbes in the Louvre Museum over time.

The microbial content of air has as yet been little described, despite its public health implications, and there remains a lack of environmental microbial data on airborne microflora in enclosed spaces. In this context, the aim of this study was to characterize the diversity and dynamics of airborne microorganisms in the Louvre Museum using high-throughput molecular tools and to underline the microbial signature of indoor air in this human-occupied environment. This microbial community was monitored for 6 month during occupied time. The quantitative results revealed variations in the concentrations of less than one logarithm, with average values of 10(3) and 10(4) Escherichia coli/Aspergillus fumigatus genome equivalent per m(3) for bacteria and fungi, respectively. Our observations highlight the stability of the indoor airborne bacterial diversity over time, while the corresponding eukaryote community was less stable. Bacterial diversity characterized by pyrosequencing 454 showed high diversity dominated by the Proteobacteria which represented 51.1%, 46.9%, and 38.4% of sequences, for each of the three air samples sequenced. A common bacterial diversity was underlined, corresponding to 58.4% of the sequences. The core species were belonging mostly to the Proteobacteria and Actinobacteria, and to the genus Paracoccus spp., Acinetobacter sp., Pseudomonas sp., Enhydrobacter sp., Sphingomonas sp., Staphylococcus sp., and Streptococcus sp.

Heterogeneity and spatial distribution of bacterial background contamination in pulp and process water of a paper mill.

Identifying the source and the distribution of bacterial contaminant communities in water circuits of industrial applications is critical even when the process may not show signs of acute biofouling. The endemic contamination of facilities can cause adverse effects on process runability but may be masked by the observed daily variability. The distribution of background communities of bacterial contaminants may therefore be critical in the development of new site-specific antifouling strategies. In a paper mill as one example for a full-scale production process, bacterial contaminants in process water and pulp suspensions were mapped using molecular fingerprints at representative locations throughout the plant. These ecological data were analyzed in the process-engineering context of pulp and water flow in the facilities. Dispersal limits within the plant environment led to the presence of distinct groups of contaminant communities in the primary units of the plant, despite high flows of water and paper pulp between units. In the paper machine circuit, community profiles were more homogeneous than in the other primary units. The variability between sampled communities in each primary unit was used to identify a possible point source of microbial contamination, in this case a storage silo for reused pulp. Part of the contamination problem in the paper mill is likely related to indirect effects of microbial activity under the local conditions in the silo rather than to the direct presence of accumulated microbial biomass.

Effect of a transient perturbation on marine bacterial communities with contrasting history.

AIMS: To evaluate the importance of the bacterial composition on the resilience of the organic matter assimilation in the sea. METHODS AND RESULTS: Chemostats were inoculated with coastal and offshore bacterial communities. Bacterial density and protein synthesis increased before stabilizing, and this response to confinement was more marked in the offshore chemostats. Before the toluene perturbation the community structure in the coastal chemostats remained complex whereas the offshore chemostats became dominated by Alteromonas sp. After the perturbation, bacterial protein synthesis was inhibited before peaking briefly at a level fivefold to that observed before the perturbation and then stabilizing at a level comparable to that before the perturbation. Alteromonas dominated both the coastal and the offshore communities immediately after the perturbation and the coastal communities did not recover their initial complexity. CONCLUSIONS: Cell lysis induced by the toluene perturbation favoured the growth of Alteromonas which could initiate growth rapidly in response to the nutrient pulse. Despite their different community structure in situ, the resilience of protein synthesis of coastal and offshore bacterial communities was dependent on Alteromonas, which dominated in the chemostats. SIGNIFICANCE AND IMPACT OF THE STUDY: Here we show that although Alteromonas sp. dominated in artificial offshore and coastal communities in chemostats, their response time to the shock was different. This suggests that future perturbation studies on resilience in the marine environment should take account of ecosystem history.

The microbial signature of aerosols produced during the thermophilic phase of composting.

AIMS: The microbial diversity of bioaerosols released during operational activities at composting plants is poorly understood. Identification of bacteria and fungi present in such aerosols is the prerequisite for the definition of microbial indicators that could be used in dispersal and exposure studies. METHODS AND RESULTS: A culture-independent analysis of composting bioaerosols collected at five different industrial open sites during the turning of composting piles in fermentation was performed by building 16S rDNA and 18S rDNA libraries. More than 800 sequences were analysed. Although differences in the phylotypes distribution were observed from one composting site to another, similarities in the structure of microbial diversity were remarkable. The same phyla dominated in the five bioaerosols: Ascomycota among fungi, Firmicutes and Actinobacteria among bacteria. For each phylum, some dominant phylotypes were common to at least four bioaerosols. These common phylotypes belonged to Thermomyces, Aspergillus, Penicillium, Geobacillus, Planifilum, Thermoactinomyces, Saccharopolyspora, Thermobifida and Saccharomonospora. CONCLUSIONS: The microbial signature of aerosols produced during the thermophilic phase of composting was determined. The similarities observed may be explained by the selection of thermophilic and sporulating species. SIGNIFICANCE AND IMPACT OF THE STUDY: Several bacteria and fungi identified in this study may represent potential indicators of composting bioaerosols in air.

Structural and functional responses of sewage microbial communities used for the treatment of a complex mixture of volatile organic compounds (VOCs).

AIMS: The aim of this work was to assess the impact of the applied mass loading on the selection of an efficient microbial community able to degrade a complex mixture of volatile organic compounds (VOCs). METHODS AND RESULTS: Two reactors were used and were supplied with a gaseous effluent containing 11 VOCs with different concentrations. The response of the microflora was monitored as a function of time: biodegradation activity, bacterial density and diversity. The results showed that the applied mass loading seems to have an impact on the functioning and the genetic structure of the bacterial community. CONCLUSIONS: A high mass loading seems to induce a low efficient functioning in terms of elimination efficiency and a simplification of the genetic structure of the total bacterial community with the apparition of a dominant microflora. A low mass loading seems to favour a better functioning and allows to keep a healthier bacterial diversity. SIGNIFICANCE AND IMPACT OF THE STUDY: In the treatment processes of gaseous effluents, it would be judicious to define the functioning parameters of the process to keep the diversity of important functional bacterial groups. These results provide also useful information about changes in microbial communities following natural or anthropogenic alterations in different ecosystems.

Synergetic effect of pH and biochemical components on bacterial diversity during mesophilic anaerobic fermentation of biomass-origin waste.

AIMS: To investigate the synergetic effect of pH and biochemical components on bacterial community structure during mesophilic anaerobic degradation of solid wastes with different origins, and under acidic or neutral conditions. METHODS AND RESULTS: The bacterial community in 16 samples of solid wastes with different biochemical compositions and origins was evaluated during mesophilic anaerobic degradation at acidic and neutral pH. Denaturing gradient gel electrophoresis (DGGE) and single-strand conformation polymorphism (SSCP) were used to compare the communities. Multivariate analysis of the DGGE and SSCP results revealed that most of the dominant microbes were dependent on the content of easily degradable carbohydrates in the samples. Furthermore, the dominant microbes were divided into two types, those that preferred an acid environment and those that preferred a neutral environment. A shift in pH was found to change their preference for medium substrates. Although most of the substrates with similar origin and biochemical composition had similar microbial diversity during fermentation, some microbes were found only in substrates with specific origins. For example, two microbes were only found in substrate that contained lignocellulose and animal protein without starch. These microbes were related to micro-organisms that are found in swine manure, as well as in other intestinal or oral niches. In addition, the distribution of fermentation products was less sensitive to the changes in pH and biochemical components than the microbial community. CONCLUSIONS: Bacterial diversity during anaerobic degradation of organic wastes was affected by both pH and biochemical components; however, pH exerted a greater effect. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study reveal that control of pH may be an effective method to produce a stable bacterial community and relatively similar product distribution during anaerobic digestion of waste, regardless of variation in the waste feedstocks.

Structural divergence of bacterial communities from functionally similar laboratory-scale vermicomposts assessed by PCR-CE-SSCP.

AIMS: To evaluate bacterial community structure and dynamics in triplicate vermicomposts made from the same start-up material, along with certain physico-chemical changes. METHODS AND RESULTS: The physico-chemical parameters (pH, temperature, carbon, nitrogen, soluble substances and cellulose) evolved similarly in the triplicate vermicomposts, indicating a steady function. The 16S bacterial gene abundance remained constant over time. To monitor changes in the bacterial community structure, fingerprinting based on capillary electrophoresis single-strand conformation polymorphism was employed. A rise in bacterial diversity occurred after precomposting and it remained stable during the maturation phase. However, a rapid shift in the structure of the bacterial community in the vermicompost replicates was noted at the beginning that stabilized with the process maturation. Multivariate analyses showed different patterns of bacterial community evolution in each vermicompost that did not correlate with the physico-chemical changes. CONCLUSIONS: The broad-scale functions remained similar in the triplicates, with stable bacterial abundance and diversity despite fluctuation in the community structure. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has demonstrated that microbial fingerprinting with multivariate analysis can provide significant understanding of community structure and also clearly suggests that an ecosystem's efficacy could be the outcome of functional redundancy whereby a number of species carry out the same function.

Bacterial community dynamics and product distribution during pH-adjusted fermentation of vegetable wastes.

AIMS: To estimate the effect of pH on the structures of bacterial community during fermentation of vegetable wastes and to investigate the relationship between bacterial community dynamics and product distribution. METHODS AND RESULTS: The bacterial communities in five batch tests controlled at different pH values [uncontrolled (about pH 4), 5, 6, 7 and 8] were monitored by denaturing gradient gel electrophoresis (DGGE) and single-strand conformation polymorphism (SSCP). The two fingerprinting methods provided consistent results and principal component analysis indicated a close similarity of bacterial community at pH 7 and 8 in addition to those at pH 4-6. This clustering also corresponded to dominant metabolic pathway. Thus, pH 7-8 shifted from alcohol-forming to acid-forming, especially butyric acid, whereas both alcohol-forming and acid-forming dominated at pH 5-6, and at pH 4, fermentation was inhibited. Shannon-weaver index was calculated to analyse the DGGE profiles, which revealed that the bacterial diversities at pH 7 and 8 were the highest while those at pH 5 and 4 (uncontrolled) were the lowest. According to sequencing results of the bands excised from DGGE gels, lactic acid bacteria and Clostridium sp. were predominant at all pH values, but varieties in species were observed as pH changed and time prolonged. CONCLUSIONS: The bacterial community during fermentation was materially influenced by pH and the diverse product distribution was related to the shift of different bacterial population. SIGNIFICANCE AND IMPACT OF THE STUDY: The study reveals that the impact of pH on fermentation product distribution is implemented primarily by changes of bacterial community. It also provides information about the comparison of two fingerprinting methods, DGGE and SSCP.

Involvement of microbial populations during the composting of olive mill wastewater sludge.

Olive mill waste water sludge obtained by the electro-Fenton oxidation of olive mill waste water was composted in a bench scale reactor. The evolution of microbial species within the composter was investigated using a respirometric test and by means of both cultivation-dependent and independent approaches (Polymerase Chain Reaction-Single Strand Conformation Polymorphism, PCR SSCP). During the period of high respiration rate (7-24 days), cultivation method showed that thermophilic bacteria as well as actinomycetes dominated over eumycetes. During the composting process, the PCR-SSCP method showed a higher diversity of the bacterial community than the eukaryotic one. After 60 days of composting, the compost exhibited a microbial stability and a clear absence of phytotoxicity.

Impact of increasing NaCl concentrations on the performance and community composition of two anaerobic reactors.

The anaerobic treatment of saline effluents using halophilic and halotolerant microbial consortia is of major interest. Inhibition of anaerobic digestion is known to occur at high salt content. However, it seems that the suitable adaptation of an anaerobic sludge makes possible the treatment of saline wastewater. In this study, a non-saline anaerobic sludge was inoculated in two anaerobic batch reactors operating with a different substrate (distillery vinasse and ethanol) and subjected to increasing NaCl concentrations. The performance of the digesters appeared to be highly dependent on the nature of the substrate, and a similar level of inhibition (i.e. around 90% of the specific loading rate and specific methanogenic activity) was stated at 10 g l(-1) of NaCl with distillery vinasse and 60 g l(-1) of NaCl with ethanol. The characterization of the microflora and its adaptation to increasing NaCl conditions were also investigated using molecular tools based on the analysis of genomic 16S rDNA. The microbial communities revealed a high diversity that could be maintained in both reactors despite the increase in NaCl concentrations.

Survival of Listeria monocytogenes and Enterococcus faecium in sludge evaluated by real-time PCR and culture methods.

AIMS: This study evaluates the behaviour in spiked sludge of a pathogenic bacteria, Listeria monocytogenes, by cultural and molecular techniques, and compares its survival with the one of a faecal indicator, Enterococcus faecium. METHODS AND RESULTS: Listeria monocytogenes strain Scott A and E. faecium(T) were followed for 17 days after inoculation in sludge. Kinetics of survival depended on the bacteria and on the technique used [most probable number method, direct plate count or real-time quantitative PCR (qPCR)]. The concentration of L. monocytogenes decreased rapidly regardless of the technique, but the decrease was much more dramatic with culture techniques than with qPCR. On the contrary, the concentrations of culturable E. faecium(T) were stable. CONCLUSIONS: The results suggest that the cells of L. monocytogenes strain Scott A might have entered a viable, but nonculturable (VBNC) status, whereas cells of the indicator bacteria, E. faecium(T), maintained themselves better and stayed culturable. SIGNIFICANCE AND IMPACT OF THE STUDY: The difference of survival kinetics in the sludge of a faecal indicator (E. faecium) and a pathogenic bacterium (L. monocytogenes) may be linked to the fact that they either enter or do not enter into a VBNC status.

Microbial diversity in hypersaline wastewater: the example of tanneries.

In contrast to conventional wastewater treatment plants and saline environments, little is known regarding the microbial diversity of hypersaline wastewater. In this study, the microbial communities of a hypersaline tannery effluent, and those of three treatment systems operating with the tannery effluent, were investigated using 16S rDNA phylogenetic markers. The comparative analysis of 377 bacterial sequences revealed the high diversity of this type of hypersaline environment, clustering within 193 phylotypes (> or = 97% similarity) and covering 14 of the 52 divisions of the bacterial domain, i.e. Proteobacteria, Bacteroidetes, Firmicutes, Actinobacteria, Chlorobi, Planctomycetes, Spirochaetes, Synergistes, Chloroflexi, Thermotogae, Verrucomicrobia, OP3, OP11 and TM7. Most of the phylotypes were related to halophilic and pollutant-degrading bacteria. Using statistical analysis, the diversity of this type of environment was compared to that of other environmental samples selected on the basis of their salinity, oxygen content and organic load.

The microbial signature of drinking waters: myth or reality?

This paper presents a new software developed for analyzing single strand conformation polymorphism (SSCP) electrophoresis patterns delivered by the genetic analyzer ABI310 (Applied Biosystems). SSCP is a molecular typing technique based on the PCR amplification of microbial 16S rDNA and used for the monitoring of complex microbial ecosystems dynamics. The software--a home-made MATLAB toolbox called MODIMECO--developed for the analysis of SSCP patterns is presented. MODIMECO includes a number of basic signal processing abilities as well as largely used statistical tools such as the well known principal component analysis. The use of the SSCP for assessing the hypothesis of the existence of a microbial signature of drinking waters illustrates the typical advantages of using such software tools. Results are discussed and conclusions drawn.

Link between spatial structure of microbial communities and degradation of a complex mixture of volatile organic compounds in peat biofilters.

AIMS: To investigate the relationships between the operation of the volatile organic compound (VOC) removal biofilter and the structure of microbial communities, and to study the impact on degradation activities and the structuring of microbial communities of biofilter malfunctions related to the qualitative composition of the polluted air. METHODS AND RESULTS: A microbiological study and a measurement of biodegradation activities were simultaneously carried out on two identical peat-packed columns, seeded with two different inocula, treating polluted air containing 11 VOCs. For both reactors, the spatial structure of the microbial communities was investigated by means of single-strand conformation polymorphism (SSCP) analysis. For both reactors, stratification of degradation activities in function of depth was observed. Oxygenated compounds were removed at the top of the column and aromatics at the bottom. Comparison of SSCP patterns clearly showed a shift in community structure in function of depth inside both biofilters. This distribution of biodegradation activities correlates with the spatialization of microbial density and diversity. Although the operating conditions of both reactors were identical and the biodegradation activities similar, the composition of microflora differed for biofilters A and B. Subdivision of biofilter B into two independent parts supplied with polluted air containing the complex VOC mixture showed that the microflora having colonized the bottom of biofilter B retained their potential for degrading oxygenated compounds. CONCLUSIONS: This work highlights the spatialization of biodegradation functions in a biofilter treating a complex mixture of VOCs. This distribution of biodegradation activities correlates with the spatialization of microbial density and diversity. SIGNIFICANCE AND IMPACT OF THE STUDY: This vertical structure of microbial communities must be taken into consideration when dealing with the malfunctioning of bioreactors. These results are also useful information about changes in microbial communities following natural or anthropogenic alterations in different ecosystems (soils and sediments) where structuring of microbial communities according to depth has been observed.

Effect of solid hold-up on nitrite accumulation in a biofilm reactor--molecular characterization of nitrifying communities.

Biological ammonium oxidation was carried out in two inverse turbulent bed reactors fed with synthetic mineral wastewater containing a high ammonium concentration (100 mg N-NH4+/L). Both reactors were started-up and operated in the same conditions except for the solid carrier concentration: the solid hold-up ratios applied, defined as the ratios of static to expanded bed height, were 0.1 and 0.3 in reactors R10 and R30 respectively. These two solid hold-up ratios generate different particle-to-particle collision frequencies and, therefore, detachment forces. The influence of solid hold-up on biofilm growth and nitrifying performance was studied from a macroscopic (i.e. nitrate and/or nitrite production) and microbiological point of view. After 60 days of operation, both reactors contained the same amount of biomass. However, R10 produced only nitrate while nitrite accumulated in R30. A comparison of microbial populations in the reactors showed that R10 contained both ammonium and nitrite oxidizing populations such as Nitrosomonas and Nitrospira, whereas in R30, ammonium oxidizing populations were much greater than those of nitrite oxidizers. The major ammonium-oxidizing organism was not the same in both reactors.