Alejandro Gonzalez-Martinez

Comparative analysis of the bacterial diversity in a lab-scale moving bed biofilm reactor (MBBR) applied to treat urban wastewater under different operational conditions

Different types of carriers were tested as support material in a lab-scale moving bed biofilm reactor (MBBR) used to treat urban wastewater under three different conditions of hydraulic retention time (HRT) and carrier filling ratios (FR). The bacterial diversity developed on the biofilms responsible of the treatment was studied using a cultivation-independent approach based on the polymerase chain reaction-temperature gradient gel electrophoresis technique (PCR-TGGE). Cluster analysis of TGGE fingerprints showed significant differences of community structure dependent upon the different operational conditions applied. Redundancy analysis (RDA) was used to determine the relationship between the operational conditions (type of carrier, HRT, FR) and bacterial biofilm diversity, demonstrating a significant effect of FR=50%. Phylogenetic analysis of PCR-reamplified and sequenced TGGE bands revealed that the prevalent Bacteria populations in the biofilm were related to Betaproteobacteria (46%), Firmicutes (34%),Alphaproteobacteria (14%) and Gammaproteobacteria (9%).

Bacterial community structure and enzyme activities in a membrane bioreactor (MBR) using pure oxygen as an aeration source

A pilot-scale membrane bioreactor was used to treat urban wastewater using pure oxygen instead of air as a source of aeration, to study its influence on bacterial diversity and levels of enzyme activities (acid and alkaline phosphatases, glucosidase, protease, and esterase) in the sludge. The experimental work was developed in two stages influenced by seasonal temperature. Operational parameters (temperature, pH, BOD5, COD, total and volatile suspended solids) were daily monitored, and enzyme activities measured twice a week. Redundancy analysis (RDA) was used to reveal relationships between the level of enzyme activities and the variation of operational parameters, demonstrating a significant effect of temperature and volatile suspended solids. Bacterial diversity was analyzed by temperature-gradient gel electrophoresis of PCR-amplified partial 16S rRNA genes. Significant differences in community structure were observed between both stages. Sequence analysis revealed that the prevalent Bacteria populations were evolutively close to Alphaproteobacteria (44%), Betaproteobacteria (25%) and Firmicutes (17%).

Effect of ciprofloxacin antibiotic on the partial-nitritation process and bacterial community structure of a submerged biofilter

A partial-nitritation bench-scale submerged biofilter was used for the treatment of synthetic wastewater containing a high concentration of ammonium in order to study the influence of the antibiotic ciprofloxacin on the partial-nitritation process and biodiversity of the bacterial community structure. The influence of ciprofloxacin was evaluated in four partial-nitritation bioreactors working in parallel, which received sterile synthetic wastewater amended with 350 ng/L of ciprofloxacin (Experiment 1), synthetic wastewater without ciprofloxacin (Experiment 2), synthetic wastewater amended with 100 ng/L of ciprofloxacin (Experiment 3) and synthetic wastewater amended with 350 ng/L of ciprofloxacin (Experiment 4). The concentration of 100 ng/L of antibiotics demonstrated that the partial-nitritation process, microbial biomass and bacterial structure generated by tag-pyrosequencing adapted progressively to the conditions in the bioreactor. However, high concentrations of ciprofloxacin (350 ng/L) induced a decay of the partial-nitritation process, while the total microbial biomass was increased. Within the same experiment, the bacterial community experienced sequential shifts with a clear reduction of the ammonium oxidation bacteria (AOB) and an evident increase of Commamonas sp., which have been previously reported to be ciprofloxacin-resistant. Our study suggests the need for careful monitoring of the concentration of antibiotics such as ciprofloxacin in partial-nitritation bioreactors, in order to choose and maintain the most appropriate conditions for the proper operation of the system.

Comparison of bacterial communities of conventional and A-stage activated sludge systems

The bacterial community structure of 10 different wastewater treatment systems and their influents has been investigated through pyrosequencing, yielding a total of 283486 reads. These bioreactors had different technological configurations: conventional activated sludge (CAS) systems and very highly loaded A-stage systems. A-stage processes are proposed as the first step in an energy producing municipal wastewater treatment process. Pyrosequencing analysis indicated that bacterial community structure of all influents was similar. Also the bacterial community of all CAS bioreactors was similar. Bacterial community structure of A-stage bioreactors showed a more case-specific pattern. A core of genera was consistently found for all influents, all CAS bioreactors and all A-stage bioreactors, respectively, showing that different geographical locations in The Netherlands and Spain did not affect the functional bacterial communities in these technologies. The ecological roles of these bacteria were discussed. Influents and A-stage bioreactors shared several core genera, while none of these were shared with CAS bioreactors communities. This difference is thought to reside in the different operational conditions of the two technologies. This study shows that bacterial community structure of CAS and A-stage bioreactors are mostly driven by solids retention time (SRT) and hydraulic retention time (HRT), as suggested by multivariate redundancy analysis.

Comparison of bacterial diversity in full scale anammox bioreactors operated under different conditions.

Bacterial community structure of full‐scale anammox bioreactor is still mainly unknown. It has never been analyzed whether different anammox bioreactor configurations might result in the development of different bacterial community structures among these systems. In this work, the bacterial community structure of six full‐scale autotrophic nitrogen removal bioreactors located in The Netherlands and China operating under three different technologies and with different influent wastewater characteristics was studied by the means of pyrotag sequencing evaluation of the bacterial assemblage yielded a great diversity in all systems. The most represented phyla were the Bacteroidetes and the Proteobacteria, followed by the Planctomycetes. 14 OTUs were shared by all bioreactors, but none of them belonged to the Brocadiales order. Statistical analysis at OTU level showed that differences in the microbial communities were high, and that the main driver of the bacterial assemblage composition was different for the distinct phyla identified in the six bioreactors, depending on bioreactor technology or influent wastewater characteristics.

Microbial community analysis of a full-scale DEMON bioreactor

Full-scale applications of autotrophic nitrogen removal technologies for the treatment of digested sludge liquor have proliferated during the last decade. Among these technologies, the aerobic/anoxic deammonification process (DEMON) is one of the major applied processes. This technology achieves nitrogen removal from wastewater through anammox metabolism inside a single bioreactor due to alternating cycles of aeration. To date, microbial community composition of full-scale DEMON bioreactors have never been reported. In this study, bacterial community structure of a full-scale DEMON bioreactor located at the Apeldoorn wastewater treatment plant was analyzed using pyrosequencing. This technique provided a higher-resolution study of the bacterial assemblage of the system compared to other techniques used in lab-scale DEMON bioreactors. Results showed that the DEMON bioreactor was a complex ecosystem where ammonium oxidizing bacteria, anammox bacteria and many other bacterial phylotypes coexist. The potential ecological role of all phylotypes found was discussed. Thus, metagenomic analysis through pyrosequencing offered new perspectives over the functioning of the DEMON bioreactor by exhaustive identification of microorganisms, which play a key role in the performance of bioreactors. In this way, pyrosequencing has been proven as a helpful tool for the in-depth investigation of the functioning of bioreactors at microbiological scale.

Archaeal Diversity in Biofilm Technologies Applied to Treat Urban and Industrial Wastewater: Recent Advances and Future Prospects

Biological wastewater treatment (WWT) frequently relies on biofilms for the removal of anthropogenic contaminants. The use of inert carrier materials to support biofilm development is often required, although under certain operating conditions microorganisms yield structures called granules, dense aggregates of self-immobilized cells with the characteristics of biofilms maintained in suspension. Molecular techniques have been successfully applied in recent years to identify the prokaryotic communities inhabiting biofilms in WWT plants. Although methanogenic Archaea are widely acknowledged as key players for the degradation of organic matter in anaerobic bioreactors, other biotechnological functions fulfilled by Archaea are less explored, and research on their significance and potential for WWT is largely needed. In addition, the occurrence of biofilms in WWT plants can sometimes be a source of operational problems. This is the case for membrane bioreactors (MBR), an advanced technology that combines conventional biological treatment with membrane filtration, which is strongly limited by biofouling, defined as the undesirable accumulation of microbial biofilms and other materials on membrane surfaces. The prevalence and spatial distribution of archaeal communities in biofilm-based WWT as well as their role in biofouling are reviewed here, in order to illustrate the significance of this prokaryotic cellular lineage in engineered environments devoted to WWT.

Bacterial community structure of a lab-scale anammox membrane bioreactor

Autotrophic nitrogen removal technologies have proliferated through the last decade. Among these, a promising one is the membrane bioreactor (MBR) Anammox, which can achieve very high solids retention time and therefore sets a proper environment for the cultivation of anammox bacteria. In this sense, the MBR Anammox is an efficient technology for the treatment of effluents with low organic carbon and high ammonium concentrations once it has been treated under partial nitrification systems. A lab‐scale MBR Anammox bioreactor has been built at the Technological University of Delft, The Netherlands and has been proven for efficient nitrogen removal and efficient cultivation of anammox bacteria. In this study, next‐generation sequencing techniques have been used for the investigation of the bacterial communities of this MBR Anammox for the first time ever. A strong domination of Candidatus Brocadia bacterium and also the presence of a myriad of other microorganisms that have adapted to this environment were detected, suggesting that the MBR Anammox bioreactor might have a more complex microbial ecosystem that it has been thought. Among these, nitrate‐reducing heterotrophs and primary producers, among others, were identified. Definition of the ecological roles of the OTUs identified through metagenomic analysis was discussed.

Treatment of Effluents Polluted by Nitrogen with New Biological Technologies Based on Autotrophic Nitrification-Denitrification Processes

In recent years, various technologies have been developed for the removal of nitrogen from wastewater that is rich in nitrogen but poor in organic carbon, such as the effluents from anaerobic digesters and from certain industries. These technologies have resulted in several patents. The core of these technologies is some of the processes and patents described in this paper: Aerobic denitrification, Sharon, Anammox, OLAND, CANON, NOx process, DEMON. More specifically, one of the first innovative options described for removing nitrogen include partial nitrification under aerobic conditions (partial Sharon process) followed by autotrophic anaerobic oxidation (Anammox process). The partial Sharon-Anammox process can be performed under alternating oxic and anoxic conditions in the same bioreactor or in two steps in two separate bioreactors. This overview focuses on the technical and biological aspects of these new types of treatment system, and compares them to other technologies. Given the fact that nitrification is a sensitive process, special attention is paid to conditions such as temperature, dissolved oxygen, hydraulic retention time, free ammonia, nitrous acid concentration, and pH. A discussion of the pros and cons of such treatment systems is also included since autotrophic nitrogen removal has advantages as well as drawbacks. The paper concludes with a discussion of future research that could improve these systems by enhancing performance and reducing costs.

Detection of comammox bacteria in full-scale wastewater treatment bioreactors using tag-454-pyrosequencing

The nitrogen cycle has been expanded with the recent discovery of Nitrospira strains that can conduct complete ammonium oxidation (commamox). Their importance in the nitrogen cycle within engineered ecosystems has not yet been analyzed. In this research, the community structure of the Bacteria domain of six full-scale activated sludge systems and three autotrophic nitrogen removal systems in the Netherlands and China has been investigated through tag-454-pyrosequencing. The phylogenetic analyses conducted in the present study showed that just a few of the Nitrospira sequences found in the bioreactors were comammox. Multivariate redundancy analysis of nitrifying genera showed an outcompetition of Nitrosomonas and non-comammox Nitrospira. Operational data from the bioreactors suggested that comammox could be favored at low temperature, low nitrogen substrate, and high dissolved oxygen. The non-ubiquity and low relative abundance of comammox in full-scale bioreactors suggested that this phylotype is not very relevant in the nitrogen cycle in wastewater treatment plants.