Meta-analysis of the Microbial Diversity Cultured in Bioreactors Simulating the Gut Microbiome.

Understanding the intricate ecological interactions within the gut microbiome and unravelling its impact on human health is a challenging task. Bioreactors are valuable tools that have contributed to our understanding of gut microbial ecology. However, there is a lack of studies describing and comparing the microbial diversity cultivated in these models. This knowledge is crucial for refining current models to reflect the gastrointestinal microbiome accurately. In this study, we analysed the microbial diversity of 1512 samples from 18 studies available in public repositories that employed cultures performed in batches and various bioreactor models to cultivate faecal microbiota. Community structure comparison between samples using t-distributed stochastic neighbour embedding and the Hellinger distance revealed a high variation between projects. The main driver of these differences was the inter-individual variation between the donor faecal inocula. Moreover, there was no overlap in the structure of the microbial communities between studies using the same bioreactor platform. In addition, α-diversity analysis using Hill numbers showed that highly complex bioreactors did not exhibit higher diversities than simpler designs. However, analyses of five projects in which the samples from the faecal inoculum were also provided revealed an amplicon sequence variants enrichment in bioreactors compared to the inoculum. Finally, a comparative analysis of the taxonomy of the families detected in the projects and the GMRepo database revealed bacterial families exclusively found in the bioreactor models. These findings highlight the potential of bioreactors to enrich low-abundance microorganisms from faecal samples, contributing to uncovering the gut microbial "dark matter".

Effect of Ecuadorian natural zeolite on the performance of anaerobic digestion of swine waste in semicontinuous regime.

This study explores the potential of zeolite as an amendment to mitigate ammonium inhibition in the anaerobic digestion of swine waste. Two 50 L reactors, one with and one without zeolite amendment were operated at an OLR of 3.0 g VS L-1d-1 for 130 days, and fed with swine waste from a full-scale pig farm. Under these conditions, zeolite doses of 4 g L-1 allowed total ammonia nitrogen (TAN) concentrations to be kept below 1000 mgNH3-N L-1. The zeolite-amended reactor not only showed an average increase of 8% in methane production under stable conditions but also exhibited 34% reduction in H2S concentrations in the biogas, compared to the reactor without zeolite. The community of archaea originating from the inoculum was conserved in the reactor with zeolite amendment, particularly the acetoclastic methanogens of the genus Methanosaeta. On the other hand, in the reactor without zeolite addition, the microbial community went from being dominated by the acetoclastic methanogen Methanosaeta to having a high relative abundance of hydrogenotrophic methanogens. The zeolite addition also favoured the reactor stability, prevented foaming, and produced an enriched natural zeolite with N, P and K. However, additional studies on the potential of enriched zeolite as a fertilizer are required, which could make the use of zeolite in Anaerobic Digestion of swine waste not only energetically favourable but also economically feasible.

Effect of Operational Parameters on the Cultivation of the Gut Microbiome in Continuous Bioreactors Inoculated with Feces: A Systematic Review.

Since the early 1980s, multiple researchers have contributed to the development of in vitro models of the human gastrointestinal system for the mechanistic interrogation of the gut microbiome ecology. Using a bioreactor for simulating all the features and conditions of the gastrointestinal system is a massive challenge. Some conditions, such as temperature and pH, are readily controlled, but a more challenging feature to simulate is that both may vary in different regions of the gastrointestinal tract. Promising solutions have been developed for simulating other functionalities, such as dialysis capabilities, peristaltic movements, and biofilm growth. This research field is under constant development, and further efforts are needed to drive these models closer to in vivo conditions, thereby increasing their usefulness for studying the gut microbiome impact on human health. Therefore, understanding the influence of key operational parameters is fundamental for the refinement of the current bioreactors and for guiding the development of more complex models. In this review, we performed a systematic search for operational parameters in 229 papers that used continuous bioreactors seeded with human feces. Despite the reporting of operational parameters for the various bioreactor models being variable, as a result of a lack of standardization, the impact of specific operational parameters on gut microbial ecology is discussed, highlighting the advantages and limitations of the current bioreactor systems.

Overview of the Nomenclature and Network of Contributors to the Development of Bioreactors for Human Gut Simulation Using Bibliometric Tools: A Fragmented Landscape.

The evolution of complex in vitro models of the human gastrointestinal system to interrogate the biochemical functionality of the gut microbiome has augmented our understanding of its role in human physiology and pathology. With 5718 authors from 52 countries, gut bioreactor research reflects the growing awareness of our need to understand the contribution of the gut microbiome to human health. Although a large body of knowledge has been generated from in vitro models, it is scattered and defined by application-specific terminologies. To better grasp the capacity of bioreactors and further our knowledge of the human gastrointestinal system, we have conducted a cross-field bibliometric search and mapped the evolution of human gastrointestinal in vitro research. We present reference material with the aim of identifying key authors and bioreactor types to enable researchers to make decisions regarding the choice of method for simulating the human gut in the context of microbiome functionality.

Analysis of the Metabolic Response of Planktonic Cells and Biofilms of Klebsiella pneumoniae to Sublethal Disinfection with Sodium Hypochlorite Measured by NMR.

Klebsiella pneumoniae is a pathogenic agent able to form biofilms on water storage tanks and pipe walls. This opportunistic pathogen can generate a thick layer as one of its essential virulence factors, enabling the bacteria to survive disinfection processes and thus develop drug resistance. Understanding the metabolic differences between biofilm and planktonic cells of the K. pneumoniae response to NaClO is key to developing strategies to control its spread. In this study, we performed an NMR metabolic profile analysis to compare the response to a sublethal concentration of sodium hypochlorite of biofilm and planktonic cells of K. pneumoniae cultured inside silicone tubing. Metabolic profiles revealed changes in the metabolism of planktonic cells after a contact time of 10 min with 7 mg L-1 of sodium hypochlorite. A decrease in the production of metabolites such as lactate, acetate, ethanol, and succinate in this cell type was observed, thus indicating a disruption of glucose intake. In contrast, the biofilms displayed a high metabolic heterogeneity, and the treatment did not affect their metabolic signature.

Exploiting the Potential of Bioreactors for Creating Spatial Organization in the Soil Microbiome: A Strategy for Increasing Sustainable Agricultural Practices.

Industrial production of synthetic nitrogen fertilizers and their crop application have caused considerable environmental impacts. Some eco-friendly alternatives try to solve them but raise some restrictions. We tested a novel method to produce a nitrogen bioinoculant by enriching a soil microbial community in bioreactors supplying N2 by air pumping. The biomass enriched with diazotrophic bacteria was diluted and applied to N-depleted and sterilized soil of tomato plants. We estimated microbial composition and diversity by 16S rRNA metabarcoding from soil and bioreactors at different run times and during plant uprooting. Bioreactors promoted the N-fixing microbial community and revealed a hided diversity. One hundred twenty-four (124) operational taxonomic units (OTUs) were assigned to bacteria with a greater Shannon diversity during the reactor's steady state. A total of 753 OTUs were found in the rhizospheres with higher biodiversity when the lowest concentration of bacteria was applied. The apparent bacterial abundance in the batch and continuous bioreactors suggested a more specific functional ecological organization. We demonstrate the usefulness of bioreactors to evidence hidden diversity in the soil when it passes through bioreactors. By obtaining the same growth of inoculated plants and the control with chemical synthesis fertilizers, we evidence the potential of the methodology that we have called directed bioprospecting to grow a complex nitrogen-fixing microbial community. The simplicity of the reactor's operation makes its application promising for developing countries with low technological progress.

Metagenomic Analysis of Biochemical Passive Reactors During Acid Mine Drainage Bioremediation Reveals Key Co-selected Metabolic Functions.

Acid mine drainage (AMD) is the major pollutant generated by the mining industry, and it is characterized by low pH and high concentration of metals and sulfate. The use of biochemical passive reactors (BPRs) is a promising strategy for its bioremediation. To date, there are various studies describing the taxonomical composition of BPR microbial communities, generally consisting of an assemblage of sulfate-reducing organisms inside Deltaproteobacteria, and a diverse set of anaerobic (ligno)cellulolytic bacteria; however, insights about its functional metagenomic content are still scarce. In previous studies, a laboratory-scale AMD bioremediation using biochemical passive reactors was designed and performed, tracking operation parameters, chemical composition, and changes, together with taxonomic composition of the microbiomes harbored in these systems. In order to reveal the main functional content of these communities, we used shotgun metagenomics analyses to explore genes of higher relative frequencies and their inferred functions during the AMD bioremediation from three BPRs representing the main microbiome compositions detected in the system. Remarkably, genes encoding for two-component regulatory systems and ABC transporters related to metal and inorganic ions, cellulose degradation enzymes, dicarboxylic acid production, and sulfite reduction complex were all detected at increased frequency. Our results evidenced that higher taxonomic diversity of the microbiome was arising together with a functional redundancy of the specific metabolic roles, indicating its co-selection and suggesting that its enrichment on BPRs may be implicated in the cumulative efficiency of these systems.

High Resistance of a Sludge Enriched with Nitrogen-Fixing Bacteria to Ammonium Salts and Its Potential as a Biofertilizer.

The increasing use of chemical fertilizers causes the loss of natural biological nitrogen fixation in soils, water eutrophication and emits more than 300 Mton CO2 per year. It also limits the success of external bacterial inoculation in the soil. Nitrogen fixing bacteria can be inhibited by the presence of ammonia as its presence can inhibit biological nitrogen fixation. Two aerobic sludges from wastewater treatment plants (WWTP) were exposed to high ammonium salts concentrations (>450 mg L-1 and >2 dS m-1). Microbial analysis after treatment through 16S pyrosequencing showed the presence of Fluviicola sp. (17.70%), a genus of the Clostridiaceae family (11.17%), and Azospirillum sp. (10.42%), which were present at the beginning with lower abundance. Denaturing gradient gel electrophoresis (DGGE) analysis based on nifH genes did not show changes in the nitrogen-fixing population. Nitrogen-Fixing Bacteria (NFB) were identified and associated with other microorganisms involved in the nitrogen cycle, presumably for survival at extreme conditions. The potential use of aerobic sludges enriched with NFB is proposed as an alternative to chemical fertilizer as this bacteria could supplement nitrogen to the plant showing competitive results with chemical fertilization.

Valorization of pulp and paper industry wastewater using sludge enriched with nitrogen-fixing bacteria.

Nitrogen-fixing bacteria (NFB) can reduce nitrogen at ambient pressure and temperature. In this study, we treated effluent from a paper mill in sequencing batch reactors (SBRs) and monitored the abundance and activity of NFB with a view to producing a sludge that could work as a biofertilizer. Four reactors were inoculated with activated sludge enriched with NFB and fed with a high C/N waste (100:0.5) from a paper mill. Though the reactors were able to reduce the organic load of the wastewater by up to 89%, they did not have any nitrogen-fixing activity and showed a decrease in the putative number of NFB (quantified with qPCR). The most abundant species in the reactors treating high C/N paper mill wastewater was identified by Illumina MiSeq 16S rRNA gene amplicon sequencing as Methyloversatilis sp. (relative abundance of 4.4%). Nitrogen fixation was observed when the C/N ratio was increased by adding sucrose. We suspect that real-world biological nitrogen fixation (BNF) will only occur where there is a C/N ratio ≤100:0.07. Consequently, operators should actively avoid adding or allowing nitrogen in the waste streams if they wish to valorize their sludge and reduce running costs. PRACTITIONER POINTS: Efficient biological wastewater treatment of low nitrogen paper mill effluent was achieved without nutrient supplementation. The sludge was still capable of fixing nitrogen although this process was not observed in the wastewater treatment system. This high C/N wastewater treatment technology could be used with effluents from cassava flour, olive oil, wine and dairy industries.

Resistance and induction of viable but non culturable states (VBNC) during inactivation of E. coli and Klebsiella pneumoniae by addition of H2O2 to natural well water under simulated solar irradiation.

Inactivation of E. coli and Klebsiella pneumoniae by addition of H2O2 10 mg L-1 into natural well water samples containing natural total iron concentrations (around 0.3 mg L-1) under simulated solar light was followed by bacterial culturability (plate count) and viability (DVC-FISH). Results showed that culturability of both bacteria was totally reduced while viability was only completely depleted for E. coli in well water samples depending of total iron concentration. Post-irradiation effects in presence of residual H2O2 showed that viability of both bacteria kept dropping being totally reduced for E. coli cells while K. pneumoniae decreased only 1-log. SEM micrographs showed that E. coli and K. pneumoniae cells underwent morphological changes and size reduction according to VBNC states. Different dark and photo-induced processes where physical-chemical features of groundwater samples play an important role could be responsible of bacteria abatement.

Enrichment of Nitrogen-Fixing Bacteria in a Nitrogen-Deficient Wastewater Treatment System.

Anthropogenic nitrogen fixation is essential to sustain a global population of 7.7 billion. However, there has been a long-standing desire to find cheaper and more environmentally friendly alternatives to the Haber-Bosch process. In this study, we developed a new strategy of nitrogen fixation by enriching free-living N2-fixing bacteria (NFB) in reactors fed with low nitrogen wastewater, analogous to those usually found in certain industrial effluents such as paper mills. Our reactors fixed appreciable quantities of nitrogen with a rate of 11.8 mg N L-1 day-1. This rate is comparable to recent "breakthrough" nitrogen-fixing technologies and far higher than observed in low C/N reactors (fed with organic matter and nitrogen). NFB were quantified using quantitative polymerase chain reaction (qPCR) of the nifH (marker gene used to identify biological nitrogen fixation) and 16S rRNA genes. The nifH gene was enriched by a factor of 10 in the nitrogen-fixing reactors (compared to controls) attaining 13% of the bacterial population (1:4.2 copies of nifH to 16S rRNA). The Illumina MiSeq 16S rRNA gene amplicon sequencing of reactors showed that the microbial community was dominated (19%) by Clostridium pasteurianum. We envisage that nitrogen-enriched biomass could potentially be used as a biofertilizer and that the treated wastewater could be released to the environment with very little post-treatment.

Photo-Fenton process at natural conditions of pH, iron, ions, and humic acids for degradation of diuron and amoxicillin.

Effect of ferric ions at concentrations typically found in natural waters (0.05 to 1.06 mg L-1) and low H2O2 concentrations (between 0.5 and 17.9 mg L-1) on simulated sunlight-induced (300 W m-2) photo-Fenton degradation at initial neutral pH (7.0) of amoxicillin and diuron in Milli-Q water was studied using an rotatable central composite experimental design 22 with a central and two axial points. H2O2 concentration was the parameter playing the key role on the degradation of both pollutants. Despite that initial pH was 7.0 in Milli-Q water, this latter decreased rapidly in the first minutes, reaching values of 3.5 and 5.0 for diuron and amoxicillin respectively after 15 min of simulated sunlight irradiation. In contrast, in presence of bicarbonate/carbonate (HCO3-/CO3=), fluoride (F-), and humic acids (HAs) at concentrations found often in surface and well waters with ferric ion and H2O2 concentrations of 0.3 and 9.7 and 15.2 mg L-1 respectively, both pollutants exhibited a strong degradation keeping the circumneutral pH. Amoxicillin and diuron degradation byproducts found by HPLC/MS were compatible with HO• and/or CO3-• radical attack. Several photo-induced processes such as photo-Fenton (by dissolved ferric-HA complexes), heterogeneous photocatalysis (by colloidal iron), UV-B H2O2 photolysis, irradiated-dissolved organic matter, and their reactions with pollutants would be the main oxidative route responsible of degradations. These findings demonstrated that it could be possible using iron concentrations often found in natural waters to oxidize via photo-Fenton processes among other events, organic pollutants at natural pH conditions.

A composite taxonomical and functional framework of microbiomes under acid mine drainage bioremediation systems.

Mining-industry is one of the most important activities in the economic development of many countries and produces highly significant alterations on the environment, mainly due to the release of a strong acidic metal-rich wastewater called acid mine drainage (AMD). Consequently, the establishment of multiple wastewater treatment strategies remains as a fundamental challenge in AMD research. Bioremediation, as a constantly-evolving multidisciplinary endeavor had been complemented during the last decades by novel tools of increasingly higher resolution such as those based on omics approaches, which are providing detailed insights into the ecology, evolution and mechanisms of microbial communities acting in bioremediation processes. This review specifically addresses, reanalyzes and reexamines in a composite comparative manner, the available sequence information and associated metadata available in public databases about AMD impacted microbial communities; summarizing our understanding of its composition and functions, and proposing potential genetic enhancements for improved bioremediation strategies. 16 S rRNA gene-targeted sequencing data from 9 studies previously published including AMD systems reported and studied around the world, were collected and reanalyzed to compare and identify the core and most abundant genera in four distinct AMD ecosystems: surface biofilm, water, impacted soils/sediments and bioreactor microbiomes. We determined that the microbial communities of bioreactors were the most diverse in bacterial types detected. The metabolic pathways predicted strongly suggest the key role of syntrophic communities with denitrification, methanogenesis, manganese, sulfate and iron reduction. The perspectives to explore the dynamics of engineering systems by high-throughput sequencing and biochemical techniques are discussed and foreseen application of synthetic biology and omics exploration on improved AMD biotransformation are proposed.

Addition of Hydrogen Peroxide to Groundwater with Natural Iron Induces Water Disinfection by Photo-Fenton at Circumneutral pH and other Photochemical Events.

Samples of natural groundwater (with low turbidity, neutral pH and 0.3 mg L-1 iron concentration) inoculated with Escherichia coli K-12 were exposed to simulated solar light both in the presence and in the absence 10 mg L-1 of H2 O2. Results demonstrated that the viability of E. coli (by DVC-FISH) was grounded to zero after 360 min of irradiation. This abatement could be caused by the oxidative stress induced by ·OH radicals or another photo-induced reactive oxygen species. Two 23 factorial experimental designs enabled the evaluation of the effects of chemical factors on the inactivation of E. coli. The first experimental design considered the pH, iron and H2 O2 , while the second evaluated the ions fluoride, carbonate and phosphate found in groundwater. pH was found to play a key role in the inactivation of E. coli. The best reduction in viability was obtained at the lower pH (6.75), while a nonsignificant effect was observed when iron or H2 O2 concentrations were raised. At higher concentrations, anions, such as carbonate and phosphate, negatively affected the E. coli abatement. However, a higher concentration of fluoride accelerated it. In all experiments, the pH was observed to rise to values higher than 8.0 units after 360 min of treatment.

2,4-D abatement from groundwater samples by photo-Fenton processes at circumneutral pH using naturally iron present. Effect of inorganic ions.

This study evaluated, at laboratory scale, if the using iron naturally present (0.3 mg L-1) and adding 10 mg L-1 of hydrogen peroxide was effective to remove 24.3 mgL-1 of 2,4-dichlorophenoxyacetic acid (2,4-D) from groundwater samples by simulated solar irradiation (global intensity = 300 W m-2). Under these conditions, the degradation of 2,4-D reached 75.2 % and the apparition of its main oxidation byproduct 2,4-dichlorophenol (DCP) was observed. On the other hand, pH exhibited an increasing from 7.0 to 8.3 during the experiment. Experiments using Milli-Q water at pH 7.0, iron, and H2O2 concentrations of 0.3 and 10 mg L-1, respectively, were carried out in order to study the effect of ions such as carbonate species, phosphate, and fluoride in typical concentrations often found in groundwater. Ion concentrations were combined by using a factorial experimental design 23. Results showed that carbonates and fluoride did not produce a detrimental effect on the 2,4-D degradation, while phosphate inhibited the process. In this case, the pH increased also from 7.0 to 7.95 and 8.99. Effect of parameters such as pH, iron concentration, and hydrogen peroxide concentration on the 2,4-D degradation by the photo-Fenton process in groundwater was evaluated by using a factorial experimental design 23. Results showed that the pH was the main parameter affecting the process. This study shows for the first time that using the photo-Fenton process at circumneutral pH and iron naturally present seems to be a promising process to remove pesticides from groundwater.

Crecimiento rápido autotrófico de microorganismos anaerobios oxidadores de amonio en presencia de nitrito, usando inóculos de Colombia / Faster autotrophic growth of anaerobic ammonium-oxidizing microorganisms in presence of nitrite, using inocula from Colombia

Anammox is a nitrite dependent process, catalyzed by bacteria of the order Brocadiales. Anammox bacteria oxidize ammonia under anoxic conditions, with nitrite as electron acceptor producing dinitrogen gas. Here, we demonstrated the presence of anammox bacteria by enriched them in a SBR reactor, with anaerobic samples taken from de bottom of a pond used in primary wastewater treatment. The enrichment reached nitrogen (N) removal rates of nearly 1.92kg N/m3/day. (The stoichiometry of the reaction matched previous anammox studies). The enriched bacterial communities were analyzed by Fluorescence In situ Hybridization (FISH), and showed a nearly 90% enrichment at the end of the experiment (day 90). As far as we know this is the first time that the anammox bacteria were enriched using Colombian inocula. The enrichment was achieved in relatively short time with high yields and have an excellent potential for application in wastewater treatment opening the opportunity to treat nitrogen-rich effluents by partial nitritation and anammox, thereby decreasing operational costs with respect to aeration (nitrification) and addition of organic electron donor (heterotrophic denitrification). This more sustainable treatment is a good alternative to control nutrient pollution in water bodies in tropical countries.

La oxidación anaerobia del amonio (anammox), es un proceso nitrito dependiente, catalizado por bacterias del filo planctomicetes. Estas bacterias oxidan el amonio en ausencia de oxígeno, con nitrito como aceptor de electrones produciendo nitrógeno molecular. En Colombia, demostramos la presencia de estas bacterias mediante el enriquecimiento de cultivos en reactores por lotes, con inóculos nativos, provenientes de muestras anaeróbias tomadas del fondo de una laguna para el tratamiento primario de aguas residuales. El enriquecimiento logrado alcanzó remociones de nitrógeno (N), en el orden de 1.92kg - N /m3/día (la estequiometria de la reacción estuvo acorde con estudios previos de anammox). La comunidad bacteriana enriquecida, se analizó mediante hibridación en sitio con fluorescencia (FISH), y mostró que el enriquecimiento contenía aprox. 90 % de bacterias anammox al final del experimento (Día 90). Esta es la primera vez que en Colombia se logra el enriquecimiento de estas bacterias con inóculos locales, hasta nuestro conocimiento. El enriquecimiento fue alcanzado en relativamente corto tiempo con altos rendimientos y tiene un excelente potencial de aplicación en el tratamiento de aguas residuales, abriendo oportunidades para el tratamiento de efluentes ricos en nitrógeno mediante nitritación parcial y anammox, disminuyendo los costos en los procesos de aireación (nitrificación) y en la de adición de donadores orgánicos (denitrificación heterótrofa). El uso de estos tratamientos más sostenibles es una buena alternativa para el control de contaminación por nutrientes en los cuerpos de agua, en países tropicales.

The detrimental influence of bacteria (E. coli, Shigella and Salmonella) on the degradation of organic compounds (and vice versa) in TiO2 photocatalysis and near-neutral photo-Fenton processes under simulated solar light.

TiO2 photocatalytic and near-neutral photo-Fenton processes were tested under simulated solar light to degrade two models of natural organic matter - resorcinol (R) (which should interact strongly with TiO2 surfaces) and hydroquinone (H) - separately or in the presence of bacteria. Under similar oxidative conditions, inactivation of Escherichia coli, Shigella sonnei and Salmonella typhimurium was carried out in the absence and in the presence of 10 mg L(-1) of R and H. The 100% abatement of R and H by using a TiO2 photocatalytic process in the absence of bacteria was observed in 90 min for R and in 120 min for H, while in the presence of microorganisms abatement was only of 55% and 35% for R and H, respectively. Photo-Fenton reagent at pH 5.0 completely removed R and H in 40 min, whereas in the presence of microorganisms their degradation was of 60% to 80%. On the other hand, 2 h of TiO2 photocatalytic process inactivated S. typhimurium and E. coli cells in three and six orders of magnitude, respectively, while S. sonnei was completely inactivated in 10 min. In the presence of R or H, the bacterial inactivation via TiO2 photocatalysis was significantly decreased. With photo-Fenton reagent at pH 5 all the microorganisms tested were completely inactivated in 40 min of simulated solar light irradiation in the absence of organics. When R and H were present, bacterial photo-Fenton inactivation was less affected. The obtained results suggest that in both TiO2 and iron photo-assisted processes, there is competition between organic substances and bacteria simultaneously present for generated reactive oxygen species (ROS). This competition is most important in heterogeneous systems, mainly when there are strong organic-TiO2 surface interactions, as in the resorcinol case, suggesting that bacteria-TiO2 interactions could play a key role in photocatalytic cell inactivation processes.

Combined anaerobic ammonium and methane oxidation for nitrogen and methane removal.

Anammox (anaerobic ammonium oxidation) is an environment-friendly and cost-efficient nitrogen-removal process currently applied to high-ammonium-loaded wastewaters such as anaerobic digester effluents. In these wastewaters, dissolved methane is also present and should be removed to prevent greenhouse gas emissions into the environment. Potentially, another recently discovered microbial pathway, n-damo (nitrite-dependent anaerobic methane oxidation) could be used for this purpose. In the present paper, we explore the feasibility of simultaneously removing methane and ammonium anaerobically, starting with granules from a full-scale anammox bioreactor. We describe the development of a co-culture of anammox and n-damo bacteria using a medium containing methane, ammonium and nitrite. The results are discussed in the context of other recent studies on the application of anaerobic methane- and ammonia-oxidizing bacteria for wastewater treatment.

Simultaneous nitrite-dependent anaerobic methane and ammonium oxidation processes.

Nitrite-dependent anaerobic oxidation of methane (n-damo) and ammonium (anammox) are two recently discovered processes in the nitrogen cycle that are catalyzed by n-damo bacteria, including "Candidatus Methylomirabilis oxyfera," and anammox bacteria, respectively. The feasibility of coculturing anammox and n-damo bacteria is important for implementation in wastewater treatment systems that contain substantial amounts of both methane and ammonium. Here we tested this possible coexistence experimentally. To obtain such a coculture, ammonium was fed to a stable enrichment culture of n-damo bacteria that still contained some residual anammox bacteria. The ammonium supplied to the reactor was consumed rapidly and could be gradually increased from 1 to 20 mM/day. The enriched coculture was monitored by fluorescence in situ hybridization and 16S rRNA and pmoA gene clone libraries and activity measurements. After 161 days, a coculture with about equal amounts of n-damo and anammox bacteria was established that converted nitrite at a rate of 0.1 kg-N/m(3)/day (17.2 mmol day(-1)). This indicated that the application of such a coculture for nitrogen removal may be feasible in the near future.

Standardization of a quantification method for Salmonella spp. and Shigella spp. in specific liquid media / Estandarización de un método de recuento para Salmonella spp. y Shigella spp. en medios de cultivo líquidos especializados

Introduction: Chlorination is the most widely used disinfection process for drinking water production. The formation of chlorination carcinogenic by-products and chlorine intoxication by direct manipulation in small communities has motivated the study of alternative disinfection processes. In this sense, processes of advanced oxidation (PAOs) have yielded promising results. Escherichia coli (E. coli) is customarily used as faecal bacterial indicator to determine the efficiency of disinfection processes. However, it has been shown that E. coli is less resistant to disinfection than other enteric bacteria such as Shigella  spp. and Salmonella  spp. Additionally, the viable non-culturable (VNC) state yields bacteria which are not detectable on many culture media. Objective: The main objective is to standardize a method for counting Salmonella  spp. and Shigella  spp. in specific liquid media to reliably quantify the bacteriological potential risk related to disinfection processes based on PAO. Methods: The study followed a randomized bi-factorial experimental design and the Duncan multiple comparison test. This design allowed the selection of specific liquid media to fittingly standardize the counting of Salmonella  spp. and Shigella  spp. Results: We found that the best broth for counting Salmonella typhimurium strain at different concentrations in pure and mixed cultures was the Rappaport broth RP, the EE broth also allowed growing the two bacterial species tested in this research. Nonetheless, the latter results suggest the use of additional tests for this particular broth. Discussion: There was a variation in the counting results when pure cultures were used compared to those obtained from mixtures of microorganisms. It was also noted that Salmonella typhimurium and Shigella sonnei, were recovered from minimal concentrations in both RP and EE broths, respectively. To some extent, this suggests an additional confirmative method when using the EE® broth...

Introducción: La cloración es el método más usado para desinfectar aguas de consumo. La formación de subproductos cancerígenos y las intoxicaciones por manipulación directa en pequeñas comunidades, han motivado el estudio de procesos alternativos. Los procesos de oxidación avanzada (PAOS), han arrojado resultados prometedores, utilizando el indicador bacteriano Escherichia coli (E. coli), con el método recuento en placa. Sin embargo, también se ha demostrado que E. coli es menos resistente a la desinfección que otras bacterias entéricas como Shigella y Salmonella y que estos procesos generan bacterias viables que no se cultivan durante el proceso, y no se descubren en medios sólidos. Objetivo: Estandarizar un método de recuento de Salmonella sp. y Shigella sp., en medios de cultivo líquidos especializados, que permita valorar de forma confiable el riesgo bacteriológico en procesos de desinfección PAOS. Métodos: En el presente trabajo se ensayaron y seleccionaron medios líquidos especializados, con los que se estandarizó el recuento de Salmonella sp. y Shigella sp., mediante un diseño experimental aleatorizado bifactorial y la prueba de comparaciones múltiples de Duncan. Resultados: Se encontró que el mejor caldo para recuperar a S. typhimurium a diferentes concentraciones, en cultivos puros y mezclas, fue el caldo Rappaport de Merck (RP). El caldo de enriquecimiento para entero bacterias de Oxoid (EE), permitió un buen crecimiento de las dos especies objeto de esta investigación. Lo cual sugiere el empleo de pruebas adicionales cuando se use caldo EE para NMP. Discusión: Se observó una variación en el recuento cuando se usaron cultivos puros, comparado con la obtenida a partir de mezclas de microorganismos. Sin embargo, S. typhimurium. y Shigella sonnei logran ser recuperadas de concentraciones mínimas en los caldos RP, respectivamente...