Microbiological water quality of the River Danube (km 2581 - km 15): Longitudinal variation of pollution as determined by standard parameters Gerhard G. Kavka1, Georg D. Kasimir2, and Andreas H. Farnleitner3 Keywords: Microbiological water quality, Large Rivers, faecal pollution Introduction The examination of microbiological river water quality according to technical standards is obligatory for use-related aspects such as for drinking water production, irrigation or recrea- tion. Microbiological data for total coliforms, faecal coliforms (representing Escherichia coli as predominant species), intestinal enterococci (faecal streptococci) and heterotrophic plate count (colony count) were collected during the Joint Danube Survey (JDS) 2001, organised by the International Commission for the Protection of the Danube River (ICPDR), along the longitudinal stretch of the River Danube from the upper section (km 2581) to the Black Sea and in major tributaries. Data were used for the assessment of microbial pollution along the course of this large river and to establish a microbiological water quality map of the investi- gated river basin. In addition observed microbiological data were compared with data from the Danube survey 1988, organised by IAD. Microbial pollution due to anthropogenic im- pacts, especially from large urban settlements, were analysed within defined sections of the River Danube and furthermore compared to biological and chemical data. Indicator bacteria and analysis methods Faecal indicator bacteria like total coliforms, faecal coliforms (thermotolerant coliforms), E. coli and intestinal enterococci (faecal streptococci) are excreted by humans and warm- blooded animals, pass sewage treatment plants to a great amount and survive for a certain time in the aquatic environment (Kavka & Poetsch 2002). E. coli and faecal coliforms are the best indicators for the assessment of recent faecal pollution, mainly caused by raw and treated sewage and diffuse impacts e. g. from farm land and pasture. E. coli and faecal coliforms in- dicate also the potential presence of pathogenic bacteria, viruses and parasites (Kavka & Po- etsch 2002). Detailed knowledge of faecal pollution in aquatic environments is crucial for watershed management activities in order to maintain safe waters for recreational and eco- nomic purposes (Farnleitner et al. 2001). The concentrations of heterotrophic bacteria (hetero- trophic plate count 22°C, colony count 22°C) correspond commonly with contamination by organic matter (Kohl 1975). Analysis of variations of indicator bacteria in the longitudinal stretch of the Danube River and major tributaries were carried out by uniform standard meth- ods in a laboratory on-board to receive comparable results according to ISO 9308-1 1990, EN ISO 7899-2 2000, EN ISO 6222 1999, DEV K5 – 1971, DIN 38411-5 – 1983 (for more de- tails see Kavka & Poetsch 2002). 1 Federal Ministry of Agriculture, Forestry, Environment and Water Management, Federal Agency for Water Management, A-3252 Petzenkirchen, Austria 2 HYDRA-Institute Österreich, Himmelstrasse 42, A-1190 Vienna 3 Institute of Chemical Engineering, Department for Applied Biochemistry and Gene Technology, University of Technology, Getreidemarkt 166-9, A-1060 Vienna 415 Variations of microbiological parameters along the longitudinal course of the Danube River and in major tributaries The Joint Danube Survey (JDS) 2001 included 98 sampling sites from Germany (Neu-Ulm, stream-km 2581) to the Black Sea (Sulina arm, stream-km 15). The variation of the microbi- ological results of all sampling sites along the flow stretch of the River Danube is presented exemplarily by the parameter faecal coliforms (Fig. 1). Tributaries are symbolised as small bars. The target value for the concentrations is marked on the chart. [CFU/ 100ml] Faecal Coliforms 100 000 000 m m r o s L A e - m i g 10 000 000 . r k r S A s a - A - n v e n e a v o 1 000 000 r s e s s o k o u c M M a R 100 000 a t R k t i e l r u i r e S P 10 000 V 1 000 100 10 1 2600 2400 2200 2000 1800 1600 1400 1200 1000 800 600 400 200 0 [Danube km] Fig. 1: Variation of faecal coliforms along the longitudinal course of the Danube River; small bars = tributaries; dotted line = target value; CFU=colony forming units Total coliforms varied from 60 to 75 000, faecal coliforms from 20 to 41 000, intestinal en- terococci from 5 to 2 200 CFU/ 100ml. The target values amount to 10 000/100ml (= imperative value acc. to EU-Bathing water quality directive 76/160 EEC), 1000/100ml and 400/100ml (= target values acc. to the EU-Bathing water quality directive 06/7/EEC) respec- tively (see table 2). In about 42% of all Danube sampling sites including arms and in 53% of the tributaries no compliance with the requirements concerning faecal indicator bacteria were found. In the Danube arms “ Danube arm, Rackeve-Soroksar Danube ar” and the tributaries “Ipoly, Russenski Lom, Arges, Siret and Prut" a great lot of faecal indicator bacte- ria were detected. Heterotrophic plate counts varied from 240 to 54 000 per 1ml in the Danube River. The target value is 10 000 colonies per 1ml water in consideration to Kohl (1975). In about 13% of sam- pling sites of the Danube River more colonies than acceptable were isolated. In the tributaries Drava, Russenski Lom and Arges the highest colony counts (maximum 1 400 000 /ml) occurred. 42% of the tributaries did not comply with the requirements. The concentration ranges of indicator bacteria analysed during JDS 2001 and IAD Survey 1988, arranged by the International Association for Danube Research, are presented in table 1. Both investigations were carried out along the Danube River by uniform methods. JDS took place in August/September 2001, the IAD survey in March 1988. The ranges of results of the two surveys are comparable, although the date sets has been recovered from different seasons. 416 Table 1: JDS 2001 and IAD-Survey 1988; concentration ranges of bacteriological parameters in the longitudinal course of the Danube River; JDS = Joint Danube Survey –ICPDR; IAD-Survey = Danube survey of the International Association for Danube Research Danube Survey JDS 2001 IAD-Survey 1988 No of measured sites 79 25 Concentration ranges 2 4 2 4 Total coliforms 100ml 6.0x10 – 7.5x10 * 4.0x10 – 5.3x10 ** 2 4 1 4 Faecal coliforms 100ml 2.0x10 – 4.1x10 * 3.0x10 – 2.3x10 ** 1 3 Intestinal enterococci 100ml 5.0 - 2.2x103* 1.0x10 - 2.7x10 *** 2 4 3 5 Plate count 22°C 1ml 2.4x10 – 5.4x10 * 1.1x10 – 1.2x10 ** *Kavka & Poetsch (2002); **Albinger (1988); ***Trzilova & Miklosovicova (1988) Microbiological Water Quality Map Microbiological water quality of River Danube and its tributaries has been assessed by the concentrations of standard microbiological parameters, classified by 5 quality classes (table 2) and represented in a microbiological map (fig. 2). Table 2: Class limit values for microbial pollutions of rivers assessed by bacteriological stan- dard parameters according to Kohl (1975, modified), Kavka & Poetsch (2002, modi- fied) and in consideration of EU-Bathing water quality directive 2006/7/EEC (target values for E. coli and enterococci concerning a good microbiological bathing water quality = target values in this table) and directive 76/160 EEC (imperative value for total coliforms concerning bathing water quality = target value in this table); TV=Target Value; FS= faecal streptococci a) faecal pollution, b) pollution by easily degradable organic matter a) Classification Class of faecal pollution I II (TV) III IV V Parameter Faecal little moderately critical strongly excessively pollution Escherichia coli in 100ml > 100 > 1 000 > 10 000 (Faecal Coli- < 100 > 100 000 water - 1 000 - 10 000 - 100 000 forms*) Intestinal Ente- in 100ml > 40 > 400 > 4 000 < 40 > 40 000 rococci (FS) water - 400 - 4 000 - 40 000 Total in 100ml > 500 > 10 000 > 100 000 < 500 >1000 000 Coliforms water -10 000 - 100 000 - 1000 000 * Faecal coliforms are mainly represented by Escherichia coli as predominant species 417 b) Classification CLASS of organic pollution I II III IV V Parameter Organic little moderately critical strongly excessively pollution Heterotrophic in 1 ml > 500 > 10 000 > 100 000 Plate Count < 500 > 750 000 water - 10 000 - 100 000 -750 000 22°C Figure 2 presents a map of faecal pollution of the Danube River at 98 measured sites during JDS 2001. 5 Classes of faecal pollution were assessed by the concentrations of faecal coli- forms and additionally by the concentrations of intestinal enterococci (faecal streptococci) (acc. to table 2). Because one sample per site was taken, direct assessment of the bathing wa- ter quality at the sampling locations cannot be given. The EU- bathing water quality directive requires replications and their respective percentiles. The microbiological samples were taken from the middle of the River Danube. Additional investigations from both river banks would allow a better detecting of the impact of tributaries, raw sewage or waste water treatment plants. In the German and Austrian section of the Danube River (km 2581 – 1880) faecal indicator bacteria indicated a little to moderate faecal contamination and organic pollution in the Da- nube and the tributary Inn (classes I and II) (figure 2). Only at one station in Hainburg, Da- nube km 1881, concentrations of faecal indicator bacteria were relative high (critical faecal pollution, class III). Influences from local contamination by the settlement Hainburg, from the tributary Schwechat River, receiving treated waste water, and probably from the sewage treatment plant of Vienna, are supposed to be responsible. The tributaries Schwechat and Mo- rava River were critical polluted by indicator bacteria. Results were corresponding with data received during the Danube survey “River Rhine to the Hungarian stretch of River Da- nube ” Sampling sites along the Danube course and in side-arms in Slovakia, Hungary, Croatia, Ser- bia, and Romania upstream Iron Gate I (stream km 1880 – 956) were moderately to strongly faecal polluted (classes II, III, IV) (figure 2).