Initiating guidance values for novel biological stability parameters in drinking water to control regrowth in the distribution system.

Nine novel biological stability parameters for drinking water have been developed recently. Here, we report data for these nine parameters in treated water from 34 treatment plants in the Netherlands to deduce guidance values for these parameters. Most parameters did not show a strong correlation with another biological stability parameter in the same sample, demonstrating that most parameters hold different information on the biological stability of drinking water. Furthermore, the novel biological stability parameters in treated water varied considerably between plants and five parameters in treated water were significantly lower for drinking water produced from groundwater than surface water. The maximum biomass concentration (MBC7), cumulative biomass potential (CBP14) from the biomass production potential test (BPP-W) and the total organic carbon concentration in treated water from groundwater were predictive parameters for HPC22 and Aeromonas regrowth in the distribution system. Guidance values of 8.6 ng ATP L-1, 110 d·ng ATP L-1 and 4.1 mg C L-1 were deduced for these parameters, under which the HPC22 and Aeromonas numbers remain at regulatory level. The maximum biomass growth (MBG7) from the BPP-W test, the particulate and/or high molecular organic carbon and the iron accumulation rate in treated water from surface water were predictive parameters for HPC22 and Aeromonas regrowth in the distribution system. Deduced guidance values for these biological stability parameters were 4.5 ng ATP L-1, 47 µg C L-1 and 0.34 mg Fe m-2 day-1, respectively. We conclude from our study that a multiple parameter assessment is required to reliable describe the biological stability of drinking water, that the biological stability of drinking water produced from groundwater is described with other parameters than the biological stability of drinking water produced from surface water, and that guidance values for predictive biological stability parameters were inferred under which HPC22 and Aeromonas regrowth is under control.

Incubation of premise plumbing water samples on Buffered Charcoal Yeast Extract agar at elevated temperature and pH selects for Legionella pneumophila.

Worldwide, over 90% of the notified cases of Legionnaires' disease are caused by Legionella pneumophila. However, the standard culture medium for the detection of Legionella in environmental water samples, Buffered Charcoal Yeast Extract (BCYE) agar of pH 6.9 ± 0.4 with or without antimicrobial agents incubated at 36 ± 1 °C, supports the growth of a large diversity of Legionella species. BCYE agar of elevated pH or/and incubation at elevated temperature gave strongly reduced recoveries of most of 26 L. non-pneumophila spp. tested, but not of L. pneumophila. BCYE agar of pH 7.3 ± 0.1, incubated at 40 ± 0.5 °C (BCYE pH 7.3/40 °C) was tested for selective enumeration of L. pneumophila. Of the L. non-pneumophila spp. tested, only L. adelaidensis and L. londiniensis multiplied under these conditions. The colony counts on BCYE pH 7.3/40 °C of a L. pneumophila serogroup 1 strain cultured in tap water did not differ significantly from those on BCYE pH 6.9/36 °C when directly plated and after membrane filtration and showed repeatability's of 13-14%. By using membrane filtration L. pneumophila was detected in 58 (54%) of 107 Legionella-positive water samples from premise plumbing systems under one or both of these culture conditions. The L. pneumophila colony counts (log-transformed) on BCYE pH 7.3/40 °C were strongly related (r2 = 0.87) to those on BCYE pH 6.9/36 °C, but differed significantly (p < 0.05) by a mean of - 0.12 ± 0.30 logs. L. non-pneumophila spp. were detected only on BCYE pH 6.9/36 °C in 49 (46%) of the samples. Hence, BCYE pH 7.3/40 °C can facilitate the enumeration of L. pneumophila and their isolation from premise plumbing systems with culturable L. non-pneumophila spp., some of which, e.g. L. anisa, can be present in high numbers.

Multiplication of Legionella pneumophila Sequence Types 1, 47, and 62 in Buffered Yeast Extract Broth and Biofilms Exposed to Flowing Tap Water at Temperatures of 38°C to 42°C.

Legionella pneumophila proliferates in freshwater environments at temperatures ranging from 25 to 45°C. To investigate the preference of different sequence types (ST) for a specific temperature range, growth of L. pneumophila serogroup 1 (SG1) ST1 (environmental strains), ST47, and ST62 (disease-associated strains) was measured in buffered yeast extract broth (BYEB) and biofilms grown on plasticized polyvinyl chloride in flowing heated drinking water originating from a groundwater supply. The optimum growth temperatures in BYEB were approximately 37°C (ST1), 39°C (ST47), and 41°C (ST62), with maximum growth temperatures of 42°C (ST1) and 43°C (ST47 and ST62). In the biofilm at 38°C, the ST47 and ST62 strains multiplied equally well compared to growth of the environmental ST1 strain and an indigenous L. pneumophila non-SG1 strain, all attaining a concentration of approximately 107 CFU/cm-2 Raising the temperature to 41°C did not impact these levels within 4 weeks, but the colony counts of all strains tested declined (at a specific decline rate of 0.14 to 0.41 day-1) when the temperature was raised to 42°C. At this temperature, the concentration of Vermamoeba vermiformis in the biofilm, determined with quantitative PCR (qPCR), was about 2 log units lower than the concentration at 38°C. In columns operated at a constant temperature, ranging from 38 to 41°C, none of the tested strains multiplied in the biofilm at 41°C, in which also V. vermiformis was not detected. These observations suggest that strains of ST47 and ST62 did not multiply in the biofilm at a temperature of ≥41°C because of the absence of a thermotolerant host. IMPORTANCE: Growth of Legionella pneumophila in tap water installations is a serious public health concern. The organism includes more than 2,100 varieties (sequence types). More than 50% of the reported cases of Legionnaires' disease are caused by a few sequence types which are very rarely detected in the environment. Strains of selected virulent sequence types proliferated in biofilms on surfaces exposed to warm (38°C) tap water to the same level as environmental varieties and multiplied well as pure culture in a nutrient-rich medium at temperatures of 42 and 43°C. However, these organisms did not grow in the biofilms at temperatures of ≥41°C. Typical host amoebae also did not multiply at these temperatures. Apparently, proliferation of thermotolerant host amoebae is needed to enable multiplication of the virulent L. pneumophila strains in the environment at elevated temperatures. The detection of these amoebae in water installations therefore is a scientific challenge with practical implications.

Transport of MS2 phage, Escherichia coli, Clostridium perfringens, Cryptosporidium parvum, and Giardia intestinalis in a gravel and a sandy soil.

To define protection zones around groundwater abstraction wells and safe setback distances for artificial recharge systems in watertreatment, quantitative information is needed about the removal of microorganisms during soil passage. Column experiments were conducted using natural soil and water from an infiltration site with fine sandy soil and a river bank infiltration site with gravel soil. The removal of phages, bacteria, bacterial spores, and protozoan (oo)-cysts was determined at two velocities and compared with field data from the same sites. The microbial elimination rate (MER) in both soils was generally >2 log, but MER in the gravel soil was higher than that in the fine sandy soil. This was attributed to enhanced attachment, related to higher metal-hydroxides content. From the high sticking efficiencies (>1) and the low influence of flow rate on MER it was deduced that straining played a significant role in the removal of Escherichia coli and Cryptosporidium parvum oocysts in the gravel soil. Lower removal of oocysts than the 4-5 times smaller E. coli and spores in the fine sand indicates that the contribution of straining is variable and needs further attention in transport models. Thus, simple extrapolation of grain size and particle size to the extent of microbial transport underground is inappropriate. Finally, the low MER of indigenous E. coli and Clostridium perfringens observed in the soil columns as well as under field conditions and the second breakthrough peak found for Cryptosporidium and spores in the fine sandy soil upon a change in the feedwater pH indicate a significant role of detachment and retardation to microbial transport and the difficulty of extrapolation of quantitative column test results to field conditions.