Priscilla Declerck

Evaluation of process parameters of ultrasonic treatment of bacterial suspensions in a pilot scale water disinfection system

In this study, several process parameters that may contribute to the efficiency of ultrasound disinfection are examined on a pilot scale water disinfection system that mimics realistic circumstances as encountered in an industrial environment. The main parameters of sonication are: (i) power; (ii) duration of treatment; (iii) volume of the treated sample. The specific energy (E(s)) is an indicator of the intensity of the ultrasound treatment because it incorporates the transferred power, the duration of sonication and the treated volume. In this study, the importance of this parameter for the disinfection efficiency was assessed through changes in volume of treated water, water flow rate and electrical power of the ultrasonic reactor. In addition, the influences of the initial bacterial concentration on the disinfection efficiency were examined. The disinfection efficiency of the ultrasonic technique was scored on a homogenous and on a mixed bacterial culture suspended in water with two different types of ultrasonic reactors (Telsonic and Bandelin). This study demonstrates that specific energy, treatment time of water with ultrasound and number of passages through the ultrasonic reactor are crucial influential parameters of ultrasonic disinfection of contaminated water in a pilot scale water disinfection system. The promising results obtained in this study on a pilot scale water disinfection system indicate the possible application of ultrasound technology to reduce bacterial contamination in recirculating process water to an acceptable low level. However, the energy demand of the ultrasound equipment is rather high and therefore it may be advantageous to apply ultrasound in combination with another treatment.

Impact of Non- Legionella Bacteria on the Uptake and Intracellular Replication of Legionella pneumophila in Acanthamoeba castellanii and Naegleria lovaniensis

In aquatic environments, Legionella pneumophila survives, in association with other bacteria, within biofilms by multiplying in free-living amoebae. The precise mechanisms underlying several aspects of the uptake and intracellular replication of L. pneumophila in amoebae, especially in the presence of other bacteria, remain unknown. In the present study, we examined the competitive effect of selected non-Legionella bacteria (Escherichia coli, Aeromonas hydrophila, Flavobacterium breve, and Pseudomonas aeruginosa) on the uptake of L. pneumophila serogroup 1 by the amoebae Acanthamoeba castellanii and Naegleria lovaniensis. We also investigated their possible influence on the intracellular replication of L. pneumophila in both amoeba species. Our results showed that the non-Legionella bacteria did not compete with L. pneumophila for uptake, suggesting that the amoeba hosts took in L. pneumophila through a specific and presumably highly efficient uptake mechanism. Living and heat-inactivated P. aeruginosa best supported the replication of L. pneumophila in N. lovaniensis and A. castellanii, respectively, whereas for both amoeba species, E. coli yielded the lowest number of replicated L. pneumophila. Furthermore, microscopic examination showed that 100% of the A. castellanii and only 2% of the N. lovaniensis population were infected with L. pneumophila at the end of the experiment. This study clearly shows the influence of some non-Legionella bacteria on the intracellular replication of L. pneumophila in A. castellanii and N. lovaniensis. It also demonstrates the different abilities of the two tested amoeba species to serve as a proper host for the replication and distribution of the human pathogen in man-made aquatic environments such as cooling towers, shower heads, and air conditioning systems with potential serious consequences for human health.

Novel magnetically induced membrane vibration (MMV) for fouling control in membrane bioreactors

Conventional submerged membrane bioreactors (MBRs) rely on the coarse bubbles aeration to generate shear at the liquid-membrane interface to limit membrane fouling. Unfortunately, it is a very energy consuming method, still often resulting in a rapid decrease of membrane permeability and consequently in higher expenses. In this paper, the feasibility of a novel magnetically induced membrane vibration (MMV) system was studied in a lab-scale MBR treating synthetic wastewater. The effects on membrane fouling of applied electrical power of different operation strategies, of membrane flux and of the presence of multiple membranes on one vibrating engine on membrane fouling were investigated. The filtration performance was evaluated by determining the filtration resistance profiles and critical flux. The results showed clear advantages of the vibrating system over conventional MBR processes by ensuring higher fluxes at lower fouling rates. Intermittent vibration was found a promising strategy for both efficient fouling control and significant energy saving. The optimised MMV system is presumed to lead to significant energy and cost reduction in up-scaled MBR operations.

Survey for the presence of specific free-living amoebae in cooling waters from Belgian power plants

Free-living amoebae (FLA) are distributed ubiquitously in aquatic environments with increasing importance in hygienic, medical and ecological relationships to man. In this study, water samples from Belgian industrial cooling circuits were quantitatively surveyed for the presence of FLA. Isolated, thermotolerant amoebae were identified morphologically as well as using the following molecular methods: enzyme-linked immunosorbent assay and isoenzyme electrophoresis and PCR. Thermophilic amoebae were present at nearly all collection sites, and the different detection methods gave similar results. Naegleria fowleri was the most frequently encountered thermotolerant species, and concentrations of thermotolerant FLA were correlated with higher temperatures.

Bacterial community analysis of activated sludge: an evaluation of four commonly used DNA extraction methods

The effectiveness of three commercially available direct DNA isolation kits (Mobio, Fast, Qiagen) and one published direct DNA extraction protocol (Bead) for extracting bacterial DNA from different types of activated sludge was investigated and mutually compared. The DNA quantity and purity were determined using real-time PCR targeting the bacterial 16S rDNA gene. Microbial community fingerprints were assessed by automated ribosomal intergenic spacer analysis. The resulting community profiles were analyzed with canonical correspondence analysis. Our results clearly demonstrate that direct DNA extraction methods can significantly influence the DNA quantity, purity, and observed community patterns of microbiota in activated sludge. Fast and Mobio generated high amounts of good quality DNA compared to Bead and Qiagen. Mobio also resulted in the detection of the highest number of species while Fast scored the best in discriminating between the community patterns of different activated sludge types. With respect to the characterization of community profiles, our analyses demonstrated a strong sludge type dependent variability among methods. Taking into account our results, we recommend Fast as the most suitable DNA extraction method for activated sludge samples used for bacterial community studies.

Receptor‐mediated uptake of Legionella pneumophila by Acanthamoeba castellanii and Naegleria lovaniensis

Aims:  Investigation of the attachment and uptake of Legionella pneumophila by Acanthamoeba castellanii and Naegleria lovaniensis, as these are two critical steps in the subsequent bacterial survival in both amoeba hosts.

Biofouling ecology as a means to better understand membrane biofouling

Despite more than a decade of worldwide research on membrane fouling in membrane bioreactors, many questions remain to be answered. Biofouling, which is referred to as the unwanted deposition and growth of biofilms, remains the main problem. Due to its complexity, most of the existing anti-biofouling strategies are not completely successful. To unravel this complexity and finally to developed well-adapted control strategies, a microbial-based description of the biofouling development is needed. Therefore, in this review, the biofouling formation will be described as a typical biofilm formation in five steps including the formation of a conditioning film, the bacterial attachment, the production of extracellular polymeric substances, the biofilm maturation, and the bacterial detachment. Moreover, important processes such as hydrodynamics and bacterial communication or quorum sensing will be taken into account. It is finally discussed whether biofouling formation is an active or inactive biofilm process together with suggestion for further research.

Spherical nucleic acid enhanced FO-SPR DNA melting for detection of mutations in Legionella pneumophila.

A home-built fiber optic surface plasmon resonance platform (FO-SPR) was applied to directly screen PCR amplified DNA for mutations. The FO-SPR sensor was used for real-time monitoring of DNA duplex melting during high resolution temperature cycling. The signal of the DNA melting was enhanced by means of gold nanoparticle labels. This FO-SPR genetic assay allowed for detection of single-point mutations (SNP) in less than 20 min. The concept was demonstrated for the analysis of 9 different serogroups of the bacterium Legionella pneumophila, a common human pathogen responsible for atypical pneumonia. FO-SPR allowed us to detect genetic mutations inhibiting PCR, which could lead to amplification bias when molecular diagnostics are applied for L. pneumophila detection. All serogroups were found to display unique melting temperatures, indicating that mutations have accumulated in the target sequence. In a next step, clinical samples of L. pneumophila were analyzed using the FO-SPR sensor. This technology was proven to be reliable for the detection of mutations for those samples that previously displayed ambiguous qPCR quantification results. When these results were benchmarked, FO-SPR results were found to be consistent with Sanger sequencing but not with fluorescence based DNA melting. The presented results convincingly advocate the advantages of FO-SPR as a high resolution and fast genetic screening tool that can compete with the current standard techniques for SNP detection.

Analysis of the microbial community structure in a membrane bioreactor during initial stages of filtration.

Membrane biofouling was investigated during the early stages of filtration in a laboratory-scale membrane bioreactor operated on molasses wastewater. The bacterial diversity and composition of the membrane biofilm and activated sludge were analyzed using terminal restriction fragment length polymorphism coupled with 16S rRNA clone library construction and sequencing. The amount of extracellular polymeric substances produced by bacteria was investigated using spectroscopic methods. The results reveal that the bacterial community of activated sludge differs significantly from that of the membrane biofilm, especially at the initial phase. Phylogenetic analysis based on 16S rRNA gene sequences identified 25 pioneer OTUs responsible for membrane surface colonization. Also, the relationship between the identified bacterial strains and the system specifications was explored.

Quantitative Detection and Differentiation of Free-Living Amoeba Species Using SYBR Green–Based Real-Time PCR Melting Curve Analysis

Real-time polymerase chain reaction melting curve analysis (MCA) allows differentiation of several free-living amoebae species. Distinctive characteristics were found for Naegleria fowleri, N. lovaniensis, N. australiensis, N. gruberi, Hartmanella vermiformis, and Willaertia magna. Species specificity of the amplicons was confirmed using agarose gel electrophoresis and sequence-based approaches. Amplification efficiency ranged from 91% to 98%, indicating the quantitative potential of the assay. This MCA approach can be used for quantitative detection of free-living amoebae after cultivation but also as a culture-independent detection method.