Talis Juhna

Biological Instability in a Chlorinated Drinking Water Distribution Network

The purpose of a drinking water distribution system is to deliver drinking water to the consumer, preferably with the same quality as when it left the treatment plant. In this context, the maintenance of good microbiological quality is often referred to as biological stability, and the addition of sufficient chlorine residuals is regarded as one way to achieve this. The full-scale drinking water distribution system of Riga (Latvia) was investigated with respect to biological stability in chlorinated drinking water. Flow cytometric (FCM) intact cell concentrations, intracellular adenosine tri-phosphate (ATP), heterotrophic plate counts and residual chlorine measurements were performed to evaluate the drinking water quality and stability at 49 sampling points throughout the distribution network. Cell viability methods were compared and the importance of extracellular ATP measurements was examined as well. FCM intact cell concentrations varied from 5×103 cells mL−1 to 4.66×105 cells mL−1 in the network. While this parameter did not exceed 2.1×104 cells mL−1 in the effluent from any water treatment plant, 50% of all the network samples contained more than 1.06×105 cells mL−1. This indisputably demonstrates biological instability in this particular drinking water distribution system, which was ascribed to a loss of disinfectant residuals and concomitant bacterial growth. The study highlights the potential of using cultivation-independent methods for the assessment of chlorinated water samples. In addition, it underlines the complexity of full-scale drinking water distribution systems, and the resulting challenges to establish the causes of biological instability.

Effect of Phosphorus on Survival of Escherichia coli in Drinking Water Biofilms

ABSTRACT The effect of phosphorus addition on survival of Escherichia coli in an experimental drinking water distribution system was investigated. Higher phosphorus concentrations prolonged the survival of culturable E. coli in water and biofilms. Although phosphorus addition did not affect viable but not culturable (VBNC) E. coli in biofilms, these structures could act as a reservoir of VBNC forms of E. coli in drinking water distribution systems.

Drinking water and biofilm disinfection by Fenton-like reaction

A Fenton-like disinfection process was conducted with Fentons reagent (H2O2) at pH 3 or 5 on autochthonous drinking water biofilms grown on corroded or non-corroded pipe material. The biofilm disinfection by Fenton-like oxidation was limited by the low content of iron and copper in the biomass grown on non-corroded plumbing. It was slightly improved by spiking the distribution system with some additional iron source (soluble iron II or ferrihydrite particles appeared as interesting candidates). However successful in situ disinfection of biofilms was only achieved in fully corroded cast iron pipes using H2O2 and adjusting the pH to 5. These new results provide additional support for the use of Fentons processes for cleaning drinking water distribution systems contaminated with biological agents or organics.

Disinfection effect of electrochemically generated chlorine on surface associated Escherichia coli in a drinking water system

AbstractFor many years, electrochemical treatment has been proposed as a potential alternative to conventional drinking water chlorination due to its simplicity, ease of use and ability to generate active disinfectant from ions naturally found in the drinking water The aim of this study was to evaluate the survival of Escherichia coli on the surfaces of water distribution system after exposure to in situ electrochemically generated chlorine. To analyse the effect of chlorine and its reaction intermediates, completely mixed reactor with or without ingenuous biofilm was supplied with natural drinking water containing low amount of chloride ions (<10 mg/L) and treated with non-stoichiometric titanium oxide electrodes (TiO2−x) at low current density (4.1–8 mA/cm2) which generate predominantly chlorine species. Various cell viability markers (cultivability, ability to divide as such and respiratory activity) were assessed in this study. The results showed that electrochemical disinfection was very effective to...

A pipeline for developing and testing staining protocols for flow cytometry, demonstrated with SYBR Green I and propidium iodide viability staining

The increasing use of flow cytometry (FCM) for analyses of environmental samples has resulted in a large variety of staining protocols with varying results and limited comparability. Viability assessment with FCM is in this context of particular interest because incorrect staining could severely affect the outcome/interpretation of the results. Here we propose a pipeline for the development and optimization of staining protocols for environmental samples, demonstrated with the common viability dye combination of SYBR Green I (SG) and propidium iodide (PI). Optimization steps included the assessment of dye solvents, determination of suitable PI concentration, and determining the optimal staining temperature and staining time. We demonstrated that dimethyl sulfoxide (DMSO) could impair membrane integrity, when used for SGPI stock solution preparation, and TRIS buffer was chosen as an alternative. Moreover we selected 6μM as optimal PI final concentration: less than 3μM resulted in incomplete staining of damaged cells, while concentrations higher that 12μM resulted in false PI-positive staining of intact cells. Low temperatures (25°C) resulted in a slow reaction and did not enable the staining of all bacteria, while high temperatures (44°C) caused damage to cells and false PI-positive results. Hence, 35°C was selected as optimal staining temperature. We further showed that a minimum of 15min were necessary to obtain stable staining results. Moreover, we showed that addition of EDTA resulted in 1-39% more PI-positive results compared to an EDTA-free sample, and argue that insufficient evidence currently exist in favor of adding EDTA to all samples in general. Altogether, the data clearly shows the need to be careful, precise and reproducible when staining cells for flow cytometric analyses, and the need to assess and optimize staining protocols with both viable and non-viable bacteria.

Hydrogen‐producing Escherichia coli strains overexpressing lactose permease: FT‐IR analysis of the lactose‐induced stress

The lactose permease gene (lacY) was overexpressed in the septuple knockout mutant of Escherichia coli, previously engineered for hydrogen production from glucose. It was expected that raising the lactose transporter activity would elevate the intracellular lactose concentration, inactivate the lactose repressor, induce the lactose operon, and as a result stimulate overall lactose consumption and conversion. However, overexpression of the lactose transporter caused a considerable growth delay in the recombinant strain on lactose, resembling to some extent the “lactose killing” phenomenon. Therefore, the recombinant strain was subjected to selection on lactose‐containing media. Selection on plates with 3% lactose yielded a strain with a decreased content of the recombinant plasmid but with an improved ability to grow and produce hydrogen on lactose. Macromolecular analysis of its biomass by means of Fourier transform‐infrared spectroscopy demonstrated that increase of the cellular polysaccharide content might contribute to the adaptation of E. coli to lactose stress.

Identifying the underlying causes of biological instability in a full-scale drinking water supply system

Changes in bacterial concentration and composition in drinking water during distribution are often attributed to biological (in)stability. Here we assessed temporal biological stability in a full-scale distribution network (DN) supplied with different types of source water: treated and chlorinated surface water and chlorinated groundwater produced at three water treatment plants (WTP). Monitoring was performed weekly during 12 months in two locations in the DN. Flow cytometric total and intact cell concentration (ICC) measurements showed considerable seasonal fluctuations, which were different for two locations. ICC varied between 0.1-3.75 × 105 cells mL-1 and 0.69-4.37 × 105 cells mL-1 at two locations respectively, with ICC increases attributed to temperature-dependent bacterial growth during distribution. Chlorinated water from the different WTP was further analysed with a modified growth potential method, identifying primary and secondary growth limiting compounds. It was observed that bacterial growth in the surface water sample after chlorination was primarily inhibited by phosphorus limitation and secondly by organic carbon limitation, while carbon was limiting in the chlorinated groundwater samples. However, the ratio of available nutrients changed during distribution, and together with disinfection residual decay, this resulted in higher bacterial growth potential detected in the DN than at the WTP. In this study, bacterial growth was found to be higher (i) at higher water temperatures, (ii) in samples with lower chlorine residuals and (iii) in samples with less nutrient (carbon, phosphorus, nitrogen, iron) limitation, while this was significantly different between the samples of different origin. Thus drinking water microbiological quality and biological stability could change during different seasons, and the extent of these changes depends on water temperature, the water source and treatment. Furthermore, differences in primary growth limiting nutrients in different water sources could contribute to biological instability in the network, where mixing occurs.

BIOBUTANOL PRODUCTION FROM AGRICULTURAL WASTE: A SIMPLE APPROACH FOR PRE-TREATMENT AND HYDROLYSIS

One of the main concerns regarding extensive production of biobutanol has been associated with the high costs of the substrate (preparation of fermentable sugars) and the relatively low tolerance of Clostridia to butanol. In this study a simple, mild approach was tested to obtain fermentable sugars from agricultural waste. Giant hogweed and hay was pre-treated with simple boiling and enzymatically hydrolysed. The results demonstrated that after adaptation of the genus Clostridium bacteria to the new substrate, the growth kinetics and sugar consumption of these bacteria were similar to the ones obtained in traditional culture media. Samazinoties pasaules naftas rezervēm, arvien lielāka uzmanība tiek pievērsta alternatīvās enerģijas ieguves veidiem. Viena no alternatīvām degvielām var būt biobutanols, ko fermentācijas laikā dabiski veido klostrīdiju ģints baktērijas. Līdz šim par vienu no kavējošiem aspektiem biobutanola plašākai lietošanai tika uzskatītas salīdzinoši dārgās ražošanas un substrāta izmaksas. Pētījumā parādīts, kā iespējams iegūt biobutanolu, izmantojot pārtikai nepiemēroto biomasu (siens, latvāņi). Procesa vienkāršošanai un izmaksu samazināšanai tradicionālās ķīmiskās un augstspiediena priekšapstrādes un hidrolīzes tehnoloģijas tika aizvietotas ar vienkāršu substrāta vārīšanu un enzimātisko hidrolīzi. Konstatēts, ka pēc nelielas klostrīdiju adaptēšanas jaunajos vides apstākļos klasiskajā augšanas barotnē un vidē ar pievienotu hidrolizātu būtiski neatšķīrās šūnu augšana un fermentējamo cukuru patēriņš. Tādējādi secināts, ka optimizācijas procesā iespējams samazināt biobutanola iegūšanas izmaksas un vienkāršot tehnoloģiskos procesu.

Drinking Water Disinfection with Electrolysis

Nowadays electrochemical disinfection has gained an increasing attention as an alternative to conventional drinking water disinfection, since it is regarded as environmentally friendly, amendable to automation, inexpensive, easily operated and is known to inactivate a wide variety of microorganisms from bacteria to viruses and algae. We found that along with increasing the number of electrodes in our equipment from 2 to 24, the resistance of chlorine-generating electrolytic cell and specific work of electric current decreased. During the electrolysis the amount of generated Cl2 increased along with the increase of chloride ion concentration in the solution and the intensity of electric current. The technological process parameters (flow rate, current intensity) have been established to obtain a predetermined amount of generated chlorine during the electrolysis process. A comparison of flow and circulating (3 times) regimes for electrolysis of tap water with chloride ion concentration below 10 mg/L showed that circulation is necessary to generate active chlorine (above 1 mg/L). At the same time, when no circulation was performed, even a 0.9 A treatment was not enough to generate detectable levels of free chlorine. Electrochemical disinfection of tap water with non-stoichiometric titanium oxide electrodes was effective enough to inactivate both metabolically active and cultivable bacteria E. coli to undetectable levels within 15 minutes at 0.5 A current intensity. Mūsdienās ūdens elektroķīmiskajai dezinfekcijai, kā alternatīvai tradicionālajām dezinfekcijas metodēm, tiek pievērsta liela uzmanība, jo tā ir videi draudzīga, viegli automatizējama, salīdzinoši lēta, viegli vadāma un ir zināma tās dezinficējošā iedarbība uz plašu mikroorganismu klāstu - no baktērijām līdz vīrusiem un aļģēm. Eksperimentāli konstatēts, ka, palielinot elektrodu skaitu elektrolīzes iekārtā no 2 līdz 24, tās pretestība un strāvas īpatnējais darbs samazinājās. Izdalītā Cl2 daudzums elektrolīzes laikā palielinājās, palielinoties hlorīda jonu koncentrācijai šķīdumā un elektriskās strāvas stiprumam. Variējot elektrolīzes procesa tehnoloģiskos parametrus (ūdens plūsmas ātrumu, strāvas stiprumu), iespējams sasniegt noteiktu izdalītā hlora daudzumu. Salīdzinot ūdens, kurš satur hlorīda jonus mazāk par 10 mg/L, apstrādi ar elektrolīzi caurplūdes režīmā ar cirkulācijas režīmu (3 reizes), konstatēts, ka ūdens apstrādi vēlams veikt cirkulācijas režīmā, lai būtu iespējams saražot vairāk aktīvā hlora (koncentrācijā, lielākā par 1 mg/L). Noteikts, ka process, veicot ūdens apstrādi ar elektrolīzi caurplūdes režīmā pat, ja strāvas stiprums 0,9 A, nenodrošināja aktīvā hlora veidošanos detektējamos daudzumos. Elektroķīmiskā dezinfekcija, izmantojot nestehiometriskā titāna oksīda elektrodus, bija pietiekami efektīva, lai pilnībā inaktivētu metaboliski aktīvās un kultivējamās E. coli baktērijas 15 minūšu laikā, ja strāvas stiprums 0,5 A.