Gertjan Medema

Removal of microorganisms by deep well injection

The removal of bacteriophages MS2 and PRD1, spores of Clostridium bifermentans R5 and . Escherichia coli WR1 by deep well injection into a sandy aquifer, was studied at a pilot field site in the southeast of the Netherlands. Injection water was seeded with the microorganisms for 5 days. Breakthrough was monitored for 93 days at 4 monitoring wells with their screens at a depth of about 310 m below surface. Within the first 8 m of soil passage, concentrations of MS2 and PRD1 were reduced by 6 log , that of R5 spores by 5 log and that of WR1 by 7.5 log . 10 10 10

Pyrosequencing reveals bacterial communities in unchlorinated drinking water distribution system: an integral study of bulk water, suspended solids, loose deposits, and pipe wall biofilm.

The current understanding of drinking water distribution system (DWDS) microbiology is limited to pipe wall biofilm and bulk water; the contributions of particle-associated bacteria (from suspended solids and loose deposits) have long been neglected. Analyzing the composition and correlation of bacterial communities from different phases helped us to locate where most of the bacteria are and understand the interactions among these phases. In the present study, the bacteria from four critical phases of an unchlorinated DWDS, including bulk water, pipe wall biofilm, suspended solids, and loose deposits, were quantified and identified by adenosine triphosphate analysis and pyrosequencing, respectively. The results showed that the bulk water bacteria (including the contribution of suspended solids) contributed less than 2% of the total bacteria. The bacteria associated with loose deposits and pipe wall biofilm that accumulated in the DWDS accounted for over 98% of the total bacteria, and the contributions of bacteria in loose deposits and pipe wall biofilm were comparable. Depending on the amount of loose deposits, its contribution can be 7-fold higher than the pipe wall biofilm. Pyrosequencing revealed relatively stable bacterial communities in bulk water, pipe wall biofilm, and suspended solids throughout the distribution system; however, the communities present in loose deposits were dependent on the amount of loose deposits locally. Bacteria within the phases of suspended solids, loose deposits, and pipe wall biofilm were similar in phylogenetic composition. The bulk water bacteria (dominated by Polaromonas spp.) were clearly different from the bacteria from the other three phases (dominated by Sphingomonas spp.). This study highlighted that the integral DWDS ecology should include contributions from all of the four phases, especially the bacteria harbored by loose deposits. The accumulation of loose deposits and the aging process create variable microenvironments inside loose deposits structures for bacteria to grow. Moreover, loose deposits protect the associated bacteria from disinfectants, and due to their mobility, the associated bacteria reach taps easily.

GAC adsorption filters as barriers for viruses, bacteria and protozoan (oo)cysts in water treatment.

Granular Activated Carbon (GAC) adsorption filtration is commonly used in drinking water treatment to remove NOM and micro-pollutants and on base of the process conditions a certain capacity to eliminate pathogenic micro-organisms was expected. The experiences with the mandatory quantitative microbial risk assessment of Dutch drinking water revealed a lack of knowledge on the elimination capacity of this process for pathogens. The objective of the current study was to determine the capacity of GAC filtration to remove MS2, Escherichia coli and spores of Clostridium bifermentans as process indicators for pathogens and more directly of (oo)cysts of Cryptosporidium parvum and Giardia lamblia. Challenge tests with fresh and loaded GAC were performed in pilot plant GAC filters supplied with pre-treated surface water at a contact time which was half of the contact time of the full-scale GAC filters. MS2 phages were not removed and the removal of E. coli and the anaerobic spores was limited ranging from < or =0.1-1.1 log. The (oo)cysts of C. parvum and G. lamblia, however, were removed significantly (1.3-2.7 log). On base of the results of the experiments and the filtration conditions the removal of the indicator bacteria and (oo)cysts was largely attributed to attachment. The model of the Colloid Filtration Theory was used to describe the removal of the dosed biocolloids in the GAC filters, but the results demonstrated that there is a lack of quantitative knowledge about the influence of collector characteristics on the two major CFT parameters, the single collector and the sticking efficiency.

Microbial Hazards in Irrigation Water: Standards, Norms, and Testing to Manage Use of Water in Fresh Produce Primary Production

Accessibility to abundant sources of high-quality water is integral to the production of safe and wholesome fresh produce. However, access to safe water is becoming increasingly difficult in many parts of the world, and this can lead to the production of fresh produce contaminated with pathogenic microorganisms, resulting in increased risk of human disease. Water, an important raw material in the fresh produce chain, is used in considerable amounts in many operations, including irrigation and application of pesticides and fertilizers, but also as a transport medium and for cooling and washing in postharvest practices. In several reported outbreaks related to uncooked fruit and vegetable products, water has been identified as a likely source of the outbreak. The present study, initiated by the ILSI Europe Emerging Microbiological Issues Task Force in collaboration with 8 other ILSI branches and support of WHO/FAO, was undertaken to review the status of, and provide suggestions for, consideration by different stakeholders on water and sanitation and its impact on food safety and public health. A limited number of guidelines and regulations on water quality for agricultural production are available, and many of them are still heavily based on microbial standards and (debated) parameters such as fecal coliforms. Data gaps have been identified with regard to baseline studies of microbial pathogens in water sources in many regions, the need for agreement on methods and microbial parameters to be used in assessing water quality, the fate of pathogens in water, and their transfer and persistence on irrigated/processed produce.

Method for rapid detection of viable Escherichia coli in water using real-time NASBA

A rapid real-time NASBA method was developed for detection of Escherichia coli in water samples. In this method, a fragment of the clpB-mRNA is amplified and a specific molecular beacon probe is used to detect the amplified mRNA fragment during the NASBA reaction. The method was shown to be specific and sensitive (1 viable E. coli in 100ml) and can be performed within 3-4h. Different inactivation processes (starvation, heat, UV-irradiation and chlorine) were employed to study the relationship between culturability and the ability to detect E. coli using NASBA. Detection of clpB-mRNA correlated with culturability after starvation or chlorine treatment. After UV-irradiation or heat-inactivation, detection of the increase in production of clpB-mRNA in viable E. coli cells after heat-shock induction correlated with culturability. Application of the NASBA method on tap water, treated sewage and surface water samples showed that culture and NASBA yielded comparable results in these different matrices. This study demonstrates that the NASBA method has high potential as a rapid test for microbiological water quality monitoring.

Unsuitability of quantitative Bacteroidales 16S rRNA gene assays for discerning fecal contamination of drinking water.

ABSTRACT Bacteroidales species were detected in (tap) water samples from treatment plants with three different PCR assays. 16S rRNA gene sequence analysis indicated that the sequences had an environmental rather than fecal origin. We conclude that assays for Bacteroidales 16S rRNA genes are not specific enough to discern fecal contamination of drinking water in the Netherlands.

Zero Risk Does Not Exist: Lessons Learned from Microbial Risk Assessment Related to Use of Water and Safety of Fresh Produce

Risk assessments related to use of water and safety of fresh produce originate from both water and food microbiology studies. Although the set-up and methodology of risk assessment in these 2 disciplines may differ, analysis of the current literature reveals some common outcomes. Most of these studies from the water perspective focus on enteric virus risks, largely because of their anticipated high concentrations in untreated wastewater and their resistance to common wastewater treatments. Risk assessment studies from the food perspective, instead, focus mainly on bacterial pathogens such as Salmonella and pathogenic Escherichia coli. Few site-specific data points were available for most of these microbial risk assessments, meaning that many assumptions were necessary, which are repeated in many studies. Specific parameters lacking hard data included rates of pathogen transfer from irrigation water to crops, pathogen penetration, and survival in or on food crops. Data on these factors have been investigated over the last decade and this should improve the reliability of future microbial risk estimates. However, the sheer number of different foodstuffs and pathogens, combined with water sources and irrigation practices, means that developing risk models that can span the breadth of fresh produce safety will be a considerable challenge. The new approach using microbial risk assessment is objective and evidence-based and leads to more flexibility and enables more tailored risk management practices and guidelines. Drawbacks are, however, capacity and knowledge to perform the microbial risk assessment and the need for data and preferably data of the specific region. Practical Application This manuscripts intends to give an extensive overview of approaches and challenges of past and future quantitative microbial risk assessment studies in the fresh produce chain related to the use of water in order to aid further research efforts in this area.

Comparison of Particle-Associated Bacteria from a Drinking Water Treatment Plant and Distribution Reservoirs with Different Water Sources

This study assessed the characteristics of and changes in the suspended particles and the associated bacteria in an unchlorinated drinking water distribution system and its reservoirs with different water sources. The results show that particle-associated bacteria (PAB) were present at a level of 0.8–4.5 × 103 cells ml−1 with a biological activity of 0.01–0.04 ng l−1 ATP. Different PAB communities in the waters produced from different sources were revealed by a 16S rRNA-based pyrosequencing analysis. The quantified biomass underestimation due to the multiple cells attached per particle was ≥ 85%. The distribution of the biologically stable water increased the number of cells per particle (from 48 to 90) but had minor effects on the PAB community. Significant changes were observed at the mixing reservoir. Our results show the characteristics of and changes in suspended PAB during distribution, and highlight the significance of suspended PAB in the distribution system, because suspended PAB can lead to a considerable underestimation of biomass, and because they exist as biofilm, which has a greater mobility than pipe-wall biofilm and therefore presents a greater risk, given the higher probability that it will reach the customers’ taps and be ingested.

Global occurrence and emission of rotaviruses to surface waters.

Group A rotaviruses (RV) are the major cause of acute gastroenteritis in infants and young children globally. Waterborne transmission of RV and the presence of RV in water sources are of major public health importance. In this paper, we present the Global Waterborne Pathogen model for RV (GloWPa-Rota model) to estimate the global distribution of RV emissions to surface water. To our knowledge, this is the first model to do so. We review the literature to estimate three RV specific variables for the model: incidence, excretion rate and removal during wastewater treatment. We estimate total global RV emissions to be 2 × 1018 viral particles/grid/year, of which 87% is produced by the urban population. Hotspot regions with high RV emissions are urban areas in densely populated parts of the world, such as Bangladesh and Nigeria, while low emissions are found in rural areas in North Russia and the Australian desert. Even for industrialized regions with high population density and without tertiary treatment, such as the UK, substantial emissions are estimated. Modeling exercises like the one presented in this paper provide unique opportunities to further study these emissions to surface water, their sources and scenarios for improved management.

Quantification of Waterborne Pathogens and Associated Health Risks in Urban Water

Citizens are exposed to microbial hazards in urban waters. To quantify health risks associated with this exposure, pathogen concentrations in an urban river, lake, rainwater sedimentation pond, a pond in a park, and a wadi, were assessed. E. coli concentrations were variable in all locations, with mean values ranging between 1.2 × 10(2) (lake) and 1.7 × 10(4) (sedimentation pond) cfu (colony forming units)/100 mL. High concentrations of Campylobacter were found, being the lowest in the lake (4.2 × 10(1) gc (genomic copies)/L) and the highest in the wadi (1.7 × 10(4) gc/L). Cryptosporidium was not found in any sample. Low levels of adenovirus 40/41 were found in some samples in the river (1.8 × 10(1) gc/L) and lake (7.2 × 10° gc/L), indicating human fecal contamination. Legionella pneumophila was found in the sedimentation pond, with higher concentrations after rain events (1.3 × 10(2) gc/L). Cyanochlorophyll-a was found in the lake (7.0 × 10(-1) μg/L), the sedimentation pond (1.1 × 10° μg/L), and the pond in the park (2.9 × 10(1) μg/L), where low levels of microcystin were found (2.1 × 10° μg/L). Campylobacter data were used to estimate gastrointestinal risks from recreational exposure. This revealed risks above the annual disease incidence of campylobacteriosis in The Netherlands, being highest in the wadi and river. Measures are proposed to reduce the health risks.