Water quality assessment of a Central European River - does the Directive 2000/60/EC cover all the needs for a comprehensive classification?

Over the last decades, European rivers have mostly shared the same fate. Until the mid 1980s many of them were in rather bad shape. Nobody would have thought about using the waters for recreation. However, continuous efforts over the last three decades have achieved success and today's water quality has changed to the better. This obvious change in water quality, which was proved by investigations done on the basis of the European Directive 2000/60/EC framework for Community action in the field of water policy, made people consider using the rivers for recreation or irrigation. In the present study a Central European River was investigated from hygienic perspectives (faecal indicators), and its toxicological status was examined - both of which are parameters which are not included in the Directive 2000/60/EC. The pivotal question was whether these parameters provide essential and additional information about the river's status and its quality. In general, the monthly values for all sampling points from the toxicological tests correlated with the determined physical and chemical statuses according to the national water management plan. Once, however, the toxicological values did increase significantly. This detection of a single toxicological burst within a year of observation underlines the applicability of the tests as screening methods for the detection of toxic substances. Moreover, our microbiological investigations show high numbers of faecal indicators and Salmonellae. These results are not consistent with the apparently good quality of the river. Therefore faecal indicators, which are directly related to the discharge of waste water, should be an integral part of the procedure of water classification.

Comparison of extended-spectrum-ß-lactamase (ESBL) carrying Escherichia coli from sewage sludge and human urinary tract infection.

For many years, extended-spectrum-beta-lactamase (ESBL) producing bacteria were a problem mainly located in medical facilities. Within the last decade however, ESBL-producing bacteria have started spreading into the community and the environment. In this study, ESBL-producing Escherichia coli from sewage sludge were collected, analysed and compared to ESBL-E. coli from human urinary tract infections (UTIs). The dominant ESBL-gene-family in both sample groups was bla(CTX-M), which is the most prevalent ESBL-gene-family in human infection. Still, the distribution of ESBL genes and the frequency of additional antibiotic resistances differed in the two sample sets. Nevertheless, phenotyping did not divide isolates of the two sources into separate groups, suggesting similar strains in both sample sets. We speculate that an exchange is taking place between the ESBL E. coli populations in infected humans and sewage sludge, most likely by the entry of ESBL E. coli from UTIs into the sewage system.

Occurrence and hygienic relevance of fungi in drinking water.

Fungi, above all filamentous fungi, can occur almost everywhere, even in water. They can grow in such a quantity in water that they can affect the health of the population or have negative effects on food production. There are several reports of fungal growth in water from different countries, but to our knowledge none from Austria so far. The aim of this study was to gain an overview of the spectrum of filamentous fungi and yeasts in drinking water systems. Thirty-eight water samples from drinking water and groundwater were analysed. Fungi were isolated by using membrane filtration and plating method with subsequent cultivation on agar plates. The different taxa of fungi were identified using routine techniques as well as molecular methods. Fungi were isolated in all water samples examined. The mean value for drinking water was 9.1 CFU per 100 ml and for groundwater 5400 CFU per 100 ml. Altogether 32 different taxa of fungi were found. The taxa which occurred most frequently were Cladosporium spp., Basidiomycetes and Penicillium spp. (74.6%, 56.4% and 48.7%, respectively). This study shows that drinking water can be a reservoir for fungi, among them opportunists, which can cause infections in immunosuppressed patients.

An integrated assessment of options for rural wastewater management in Austria.

This paper reports a recently finished, interdisciplinary project on rural wastewater management in Austria. The objective of the project was to study alternative wastewater management options based on separation of the wastewater into its constituent parts, and to compare them with conventional ones. Thereby, a feasibility study of both conventional and alternative options for wastewater management in six model regions was carried out. Life cycle costs and social acceptance were analysed by using a case study-based assessment approach. However, hygienic and environmental risks were evaluated on a more general level. In order to complement the findings, a survey on urine separation system users in the Solar City of Linz was conducted. Based on these assessments and empirical findings, the paper concludes that options using a full separation of all wastewater fractions should be considered with care. Options based on a separation of only grey water and black water or in the liquid/solid phase can offer ecological and financial advantages over conventional options. Further, options combining wastewater management and regional biogas plants were identified as an interesting solution. However, legal constraints restrict this option currently.

Association mapping of Stagonospora nodorum blotch resistance in modern European winter wheat varieties.

Association mapping in populations relevant for wheat breeding has a large potential for validating and fine-mapping QTLs identified in F2- or DH (double haploid)-derived populations. In this study, associations between markers in the region of QSng.sfr-3BS, a major QTL for resistance to Stagonospora nodorum glume blotch (SNG), and SNG resistance were investigated by linkage and association analyses. After increasing marker density in 240 F(5:7) recombinant inbred lines (RILs), QSng.sfr-3BS explained 43% of the genetic variance and peaked 0.6 cM proximal from the marker SUN2-3B. Association between SNG resistance and markers mapped in the region of QSng.sfr-3BS was investigated in a population of 44 modern European winter wheat varieties. Two genetically distinct subpopulations were identified within these lines. In agreement with linkage analyses, association mapping by a least squares general linear model (GLM) at marker loci in the region of QSng.sfr-3BS revealed the highest association with SNG resistance for SUN2-3B (p < 0.05). Association mapping can provide an effective mean of relating genotypes to complex quantitative phenotypes in hexaploid wheat. Linkage disequilibrium (r (2)) in chromosome 3B extended less than 0.5 cM in 44 varieties, while it extended about 30 cM in 240 RILs, based on 91 SSR and STS marker-pair comparisons. This indicated that the association mapping population had a marker resolution potential at least 390-fold higher compared to the RIL population.

Antibiotic resistance of E. coli in sewage and sludge.

The aim of the study is the evaluation of resistance patterns of E. coli in wastewater treatment plants without an evaluation of basic antibiotic resistance mechanisms. Investigations have been done in sewage, sludge and receiving waters from three different sewage treatment plants in southern Austria. A total of 767 E. coli isolates were tested regarding their resistance to 24 different antibiotics. The highest resistance rates were found in E. coli strains of a sewage treatment plant which treats not only municipal sewage but also sewage from a hospital. Among the antimicrobial agents tested, the highest resistance rates in the penicillin group were found for Ampicillin (AM) (up to 18%) and Piperacillin (PIP) (up to 12%); in the cephalosporin group for Cefalothin (CF) (up to 35%) and Cefuroxime-Axetil (CXMAX) (up to 11%); in the group of quinolones for Nalidixic acid (NA) (up to 15%); and for Trimethoprime/Sulfamethoxazole (SXT) (up to 13%) and for Tetracycline (TE) (57%). Median values for E. coli in the inflow (crude sewage) of the plants were between 2.0 x 10(4) and 6.1 x 10(4)CFU/ml (Coli ID-agar, BioMerieux 42017) but showed a 200-fold reduction in all three plants in the effluent. Nevertheless, more than 10(2)CFU E. coli/ml reached the receiving water and thus sewage treatment processes contribute to the dissemination of resistant bacteria in the environment.

The significance of sunlight for the elimination of indicator bacteria in small-scale bathing ponds in central Europe.

In order to determine the impact of sunlight on the reduction rate of indicator bacteria, bathing waters of different turbidity were artificially contaminated with indicator bacteria and exposed to sunlight. The reduction rates determined for the indicator bacteria showed a significant correlation with radiation intensity and, thus, also the time of day as well as the turbidity of the water. Under conditions of strong radiation intensity and low water turbidity (< 5 FTU), indicator bacteria were eliminated within 1-2 h of contamination with an initial concentration of 10(5) CFU/mL. Moreover, differences between different indicator bacteria were found. Enterococcus faecalis proved to be significantly more resistant to sunlight than the other indicator bacteria tested (Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus). The results of the present study, namely the complete elimination of indicator bacteria within a few hours, showed that sunlight, given an appropriate intensity and good water transparency, was the most important factor in the reduction of hygienically-relevant microorganisms in surface waters and, thus, also in small-scale bathing ponds.

Impact of biocontrol strain Pseudomonas fluorescens CHA0 on rhizosphere bacteria isolated from barley (Hordeum vulgare L.) with special reference to Cytophaga-like bacteria.

AIMS: To assess the impact of the biocontrol strain Pseudomonas fluorescens CHA0 on a collection of barley rhizosphere bacteria using an agar plate inhibition assay and a plant microcosm, focusing on a CHA0-sensitive member of the Cytophaga-like bacteria (CLB). METHODS AND RESULTS: The effect of strain CHA0 on a collection of barley rhizosphere bacteria, in particular CLB and fluorescent pseudomonads sampled during a growth season, was assessed by a growth inhibition assay. On average, 85% of the bacteria were sensitive in the May sample, while the effect was reduced to around 68% in the July and August samples. In the May sample, around 95% of the CLB and around 45% of the fluorescent pseudomonads were sensitive to strain CHA0. The proportion of CHA0-sensitive CLB and fluorescent pseudomonad isolates decreased during the plant growth season, i.e. in the July and August samples. A particularly sensitive CLB isolate, CLB23, was selected, exposed to strain CHA0 (wild type) and its genetically modified derivatives in the rhizosphere of barley grown in gnotobiotic soil microcosms. Two dry-stress periods were imposed during the experiment. Derivatives of strain CHA0 included antibiotic or exopolysaccharide (EPS) overproducing strains and a dry-stress-sensitive mutant. Despite their inhibitory activity against CLB23 in vitro, neither wild-type strain CHA0, nor any of its derivatives, had a major effect on culturable and total cell numbers of CLB23 during the 23-day microcosm experiment. Populations of all inoculants declined during the two dry-stress periods, with soil water contents below 5% and plants reaching the wilting point, but they recovered after re-wetting the soil. Survival of the dry-stress-sensitive mutant of CHA0 was most affected by the dry periods; however, this did not result in an increased population density of CLB23. CONCLUSIONS: CLB comprise a large fraction of barley rhizosphere bacteria that are sensitive to the biocontrol pseudomonad CHA0 in vitro. However, in plant microcosm experiments with varying soil humidity conditions, CHA0 or its derivatives had no major impact on the survival of the highly sensitive CLB strain, CLB23, during two dry-stress periods and a re-wetting period; all co-existed well in the rhizosphere of barley plants. SIGNIFICANCE AND IMPACT OF THE STUDY: Results indicate a lack of interaction between the biocontrol pseudomonad CHA0 and a sensitive CLB when the complexity increases from agar plate assays to plant microcosm experiments. This suggests the occurrence of low levels of antibiotic production and/or that the two bacterial genera occupy different niches in the rhizosphere.

Improved method for the fluorimetric detection of beta-D-galactosidase in water.

A very convenient method to quantify coliform bacteria in water can probably be designed via the determination of the activity of the enzyme beta-D-galactosidase, whose natural occurrence is, apart from less frequently occurring aeromonads mainly restricted to this type of microorganisms. 4-methylumbelliferyl-beta-D-galactoside is used as substrate, which is hydrolyzed during the enzymatic reaction; the released 4-methylumbelliferone can be quantified fluorimetrically. In the present study the influence of various physical and chemical parameters on the determination is investigated and the experimental conditions are optimized. Most important entities are the pH value during hydrolysis, the presence of nutrients and co-factors in the sample, and the modification of the substrate. Statistical evaluation of the results obtained by changing single or multiple parameters reflects clearly their positive or negative influence on the enzyme activity. Thus, deliberate addition of surfactants, specific nutrients, salts and co-enzymes results in a significantly increased activity of beta-D-galactosidase towards the substrate, which can be advantageously exploited to increase the sensitivity of the analytical method together with a decrease of the detection limit. The influence of the parameters and the optimized conditions of the improved analytical methods are presented.

The viable-but-nonculturable state induced by abiotic stress in the biocontrol agent Pseudomonas fluorescens CHA0 does not promote strain persistence in soil.

The effects of oxygen limitation, low redox potential, and high NaCl stress for 7 days in vitro on the rifampin-resistant biocontrol inoculant Pseudomonas fluorescens CHA0-Rif and its subsequent persistence in natural soil for 54 days were investigated. Throughout the experiment, the strain was monitored using total cell counts (immunofluorescence microscopy), Kogure's direct viable counts, and colony counts (on rifampin-containing plates). Under in vitro conditions, viable-but-nonculturable (VBNC) cells of CHA0-Rif were obtained when the strain was exposed to a combination of low redox potential (230 mV) and oxygen limitation. This mimics a situation observed in the field, where VBNC cells of the strain were found in the water-logged soil layer above the plow pan. Here, VBNC cells were also observed in vitro when CHA0-Rif was subjected to high NaCl levels (i.e., NaCl at 1.5 M but not 0.7 M). In all treatments, cell numbers remained close to the inoculum level for the first 12 days after inoculation of soil, regardless of the cell enumeration method used, but decreased afterwards. At the last two samplings in soil, VBNC cells of CHA0-Rif were found in all treatments except the one in which log-phase cells had been used. In the two treatments that generated high numbers of VBNC cells in vitro, VBNC cells did not display enhanced persistence compared with culturable cells once introduced into soil, which suggests that this VBNC state did not represent a physiological strategy to improve survival under adverse conditions.

Metabolism and effect of nitrates.

The effect of nitrate on the health of mice in a long term study is described. Nitrate was given to the mice in the form of calcium nitrate in drinking water in varying concentrations, and the mice were observed over the course of 18 months. Three groups were formed: 1. control group, 2. group I (100 mg nitrate/l drinking water) and group II (1,000 mg nitrate/l drinking water). The parameters studied were liver function, kidney function, total iron, ammonium, total protein and electrophoresis of the various serum proteins, body weight, and N-glycolyl-neuraminic acid as a tumor marker. Nitrate is broken down via nitrate, hydroxylamine to ammonium and finally to urea. The concentration of urea increased with time and higher concentrations of nitrate load. The exposed mice clearly lost weight and died prematurely. This was true only for the mice in group II. With a load of 100 mg/l drinking water, no change in the physiological parameters could be seen within the time period studied.

[Modification of the nitrate content of groundwater near the surface by agricultural soil use in the example of Styria]. / Beeinflussung des Nitratgehaltes oberflächennahen Grundwassers durch landwirtschaftliche Bodennutzung am Beispiel der Steiermark.

In a one year study, nitrate concentrations of superficial ground water were determined in Styria. Of 3931 samples, 21.1% were above 30 mg, 10.1% above 50 mg and 1.5% above 100 mg NO3/1. The contaminated areas lie exclusively in the Styrian tertiary basin, especially in the loose fluvoglacial deposits downstream of Graz. These areas are further characterized by a high population density, intensive cultivation of maize and intensive animal farming (pig breeding). Aside from toxicological aspects, increased nitrate concentrations are an indicator of human influences on the environment, which no longer permit the exclusion of substances potentially deleterious to health in the ground water. The areas are of great interest from the point of view of water supply and distribution because of the great quantities of water to be found there.

Hygienic investigation of coastal waters of the upper Adriatic Sea.

In the course of the bathing season of 1989, investigations of bathing waters were carried out in two-week intervals. From the point of view of public health, the chemical-physical and microbiological results do not suggest objections against bathing at the beach areas investigated (Grado, Lignano). However, these favorable results do not imply intact ecological conditions. National and international standards are designed for humans and allow only very limited conclusions about the living conditions of the marine ecosystem. This also means that ecological investigations are not sufficient to permit conclusions about hygienic conditions. In spite of this seeming contradiction, hygienic and ecological concerns are clearly identical. Hygienic measures must not be limited to local "cosmetic" corrections but must target foremost unfavorable basic conditions.

[In vitro sensitivity of Aeromonas spp. to antibiotics]. / In-vitro-Empfindlichkeit von Aeromonas spp. gegen Antibiotika.

Aeromonas strains, isolated from stool samples and aquatic biotopes were tested for their susceptibility to 20 antibiotics. The beta-lactamase-resistant cephalosporins, imipenem, aminoglycosides, ciprofloxacin, tetracycline, chloramphenicol and sulfamethoxazole + trimethoprim are highly efficient.

Cryptosporidiosis in children in Idukki District in southern India.

Concentrated stool samples from 560 children, aged from two weeks to 10 years, from Idukki District, southern India, were examined to detect Cryptosporidium oocysts, using a phenol-auramin stain and fluorescent microscopy. Smears of stool samples were also stained by a modified Ziehl-Neelsen method and were examined by light-microscopy. One sample was collected from each of 266 children with diarrhoea and 294 age-matched children without diarrhoea. Cryptosporidium oocysts were detected in 4.5% of all stools, in 6% of diarrhoeal stools and in 3% of stools from healthy controls. Results of the study show that Cryptosporidium infection is common in children with diarrhoea in southern India, and there is also a high frequency of asymptomatic cyst-passers in this area.

The germ-spectrum problem and antibiotic-resistance in Bida and Minna hospitals--Nigeria.

In the present paper the occurrence of different species of Pseudomonas, of Acinetobacter calcoaceticum var. anitratum and Aeromonas hydrophila in hospitals is described. The possible human pathogenic significance is discussed.

Leptospirosis in the Melut district--upper Nile province (south Sudan)--an overview.

The authors examined 195 domestic animals for leptospirosis serologically in Sudan (170 cattle, 7 sheep and 18 goats) and 771 wild animals (36 species) with 13 serovars from 13 serogroups. 54% of domestic animals namely, 108 cattle and 1 sheep were positive in titer levels of 1:400 and higher. Cattle sera with the serovars of the serogroup Icterohaemorrhagiae were 2.4% positive with Cynopteri 2.4%, Autumnalis 3.5%, Australis 1.2%, Pomona 2.9%, Grippotyphosa 2.9%, Hebdonadis 17.1%, Bataviae 9.4% and Tarassovi 50.6%. Only one Numida meleagris of the 54 wild birds (14 species) was positive in low titer level of 1:100 with L. canicola. Seventy (9.8%) of the 717 wild mammals (22 species) examined were positive namely with the serovars of the serogroups Icterohaemorrhagiae 3.4%, Javanica 3.0%, Pomona 2.0%, Grippotyphosa 0.7%, Hebdomadis 0.4%, Ballum, Australis, Pyrogenes, Bataviae each 0.3% and Cynopteri 0.1%, whereby sera from 7 animals reacted simultaneously with the serovars of two serogroups. With regard to the species, positive reactions were found in Erinaceus albiventris, Eidolon helbum, Cercopithecus aethiops, Genetta spec., Canis adustus et aurus, Felis silvestris, Leptailurus serval, Arvicanthis niloticus, Mastomys spec., Mus spec., Gazella spec., Tragelaphus spec. The positivity in rodents was very low - 1.2%.

[Significance of the occurrence of Aeromonas of various biotypes in the Third World]. / Zur Bedeutung von Auftreten von Aeromonaden in verschiedenen Biotopen der 3. Welt.

In investigations carried out in the refugee camps of the Polisario (South Algeria), in the neighbourhood of the inhabitants of the waste disposal sites of Cairo and in the Ayoun el Atrous and Nema (Mauritania) regions also in the Bida and Minna hospitals. Aeromonas hydrophila and Aeromonas spec. were repeatedly found in the investigated material. Therefore, it was of particular interest for us to investigate this type of organism.

[Water studies of inhabitants of refuse disposal sites in Cairo. 1]. / Wasseruntersuchungen bei den Bewohnern der Mülldeponien in Kairo. 1. Mitteilung.

In two waste disposal sites in Cairo, drinking water samples from springs, drinking vessels and kitchens were investigated. Total germ count lay seldom below one thousand and ten million pro ml. Apart from various faecal germs, Aeromonas hydrophila was also found. The high occurrence of diarrhoeal diseases among the inhabitants of the waste disposal sites can be traced with certainty to the extremely bad drinking water quality.

Microbiological and chemical analysis of drinking water in southern India (with special consideration of Aeromonas species).

Analyses of drinking water taken from central and individual water supplies in Southern India (Idukki/Kerala) showed bacterial counts of 10(3)-10(6). E. coli were found in 11% and 28% of the samples. Aeromonads were identified more frequently than E. coli at 44% in central and 38% in individual water supplies. Due to this high rate of contamination, Aeromonas spp. must be considered as a possible cause of diarrhoea and other infections in this area.