Biomedical subjects
Publications and source records attributed to H E Müller.
Consistent major differences were detected by SDS-PAGE, Western blotting and Ouchterlony immunodiffusion in four axenic and microscopically indistinguishable strains of the anaerobic human parasite Blastocystis hominis from different sources. It is concluded that at least two variants with different polypeptide patterns and antigens exist and the biological significance of these findings is discussed.
During the summer season of 1989 about 222 samples of bathing water (Northsea, Baltic Sea and inland waters) were investigated, i.e. 2 times 1998 dilution tubes (1501 positive) were tested. Results with BRILA-MUG and lactose-bouillon were compared and additional 97 samples were tested with one respectively three colonies by confirmative tests. -- The BRILA-MUG one-tube-test (gas, fluorescence and indol) has as expected larger numbers of total coliforms (GC) and faecal coliforms (FC) in comparison with corresponding numbers for E. coli and coliform bacteria according to German law for drinking water (TrinkwV). -- BRILA-MUG and lactose-bouillon with the same identification mode according to "TrinkwV" has corresponding results concerning E. coli and coliform bacteria. -- Following the identification mode according to "TrinkwV" the total coliforms (GC/gas in BRILA-MUG) depending on the origin proved 60-80% as coliform bacteria. Additional API-tests showed that the other bacteria are coliform bacteria too or bacteria which can be considered as indicators for water pollution. -- Faecal coliforms (FC/fluorescence and indol-positive) depending on the origin proved 70-90% as E. coli, if following the identification mode according to "TrinkwV". Using 3 instead of 1 colony per positive dilution tube for identification the positive results increased approximately by 9%. 15% of negative results with the identification mode according to "TrinkwV" proved as E. coli-positive with identification according to API-system, i.e. the corresponding rate of E. coli-positive tubes will therefore be higher than shown above. The BRILA-MUG one-tube-test in connection with the MPN-method can be used successfully to determine the number of total and faecal coliforms according to the EEC-directive for bathing waters. The test needs only a minimum of material and also a minimum of laboratory staff. Differences between this test and other more extensive tests with several biochemical identification steps are negligible. The work load for bathing water tests would not be justified in this case.
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Listeria monocytogenes cells are ingested by protozoa such as Acanthamoeba sp. or Tetrahymena pyriformis. However, they are not killed, but survive within the protozoa and may multiply intracellularly. The protozoa are lysed within about 8 days, releasing viable L. monocytogenes. No co-existence was observed between L. monocytogenes and Tetrahymena. A co-culture of L. monocytogenes and Acanthamoeba sp. showed a decay of locomotive forms and release of listeria from vegetative protozoan cells whereas the bacteria were destroyed in cysts. These phenomena provide an insight into the pathogenesis of listeria infection in man and warm-blooded animals because intracellular processes occurring in protozoa after ingestion of L. monocytogenes may be similar to those observed in mammalian cells.
In order to test the usefulness of BRILA-MUG (= Fluorocult) medium (Merck) for isolation and identification of total coliforms and faecal coliforms in surface water according to the EC guidelines for bathing waters a total of 969 strains of different Enterobacteriaceae and Vibrionaceae species was examined under different culture conditions. These included 486 E. coli (reference strains of O-groups 1-170, enterotoxin and Verotoxin-producing strains), 149 Salmonella (subspecies I-VI), 92 Yersinia, 44 Shigella, 64 Vibrio, 16 Aeromonas, and 118 strains of other Enterobacteriaceae species. After 48 h incubation at 36 degrees C 372 (82.0%) of 486 E. coli showed the typical reactions of gas formation, fluorescence (beta-glucuronidase activity), and indole production. Examination of fluorescence after addition of 1 N NaOH (0.5 ml), or testing of indole production after subculture in tryptophane containing broth improved the amount of typical reactions to 434 (95.4%). Incubation at 44 degrees C for 48 h gave less favourable results as compared with that at 36 degrees C. Out of 483 strains of other species 3.9% Salmonella strains of subspecies II-IV, 6.25% Citrobacter freundii, and 50% Edwardsiella tarda strains yielded reactions typical of E. coli. Shigella and Yersinia strains occasionally produced indole or fluorescence, but never visible gas.
The prevention of possible outbreaks as they occurred hundred years ago is still the object of the control of drinking water by the EC-guideline and the Drinking Water Regulations, respectively. At that time, the principle of faecal indicator bacteria was developed and it proved very useful. Now, however, drinking water is no longer supplied by wells but by plants. In comparison with the situation during the last century the relative risks of infectious diseases by faecal organisms are now drastically lower. An intelligent strategy of prevention of drinking water epidemics has to take into consideration that enteropathogens have become too rare so that it is hardly possible to detect them by the frequently occurring faecal markers E. coli and total coliforms. The alternative is the control of the "good manufacturing practices" (GMP) during the processing of drinking water. The colony counts seem to be better markers than the coliforms. Today, more than 90 per cent of infections by drinking water are due to legionellas and ubiquitous mycobacteria occurring in warm water. Therefore, warm water has to be included in the normal control. Furthermore, legionellae and ubiquitous mycobacteria should be tested regularly. Against that, the investigation of total coliforms and/or faecal streptococci seems superfluous. Finally, the future safety of drinking water demands the investigation of at least two additional chemical contaminants, i.e. asbestos and radon, whereas the cyanide test does not appear necessary.
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In the monitoring of bathing waters according to EC Guideline 76/160 EWG, total coliform and faecal coliform bacteria have to be regularly determined by a quantitative method. This requirement can be fulfilled with sufficient reliability by the MPN method. Therefore, a procedure had to be developed which on the one hand is practical and not laborious, and on the other hand warrants a sufficiently specific, quantitative bacteriological diagnostic. The commercially available BRILA-MUG-broth appeared suitable for this purpose as it allows a direct determination of total as well as faecal coliform bacteria in the same assay. By this method, total coliform bacteria are recognized by gas production from lactose whereas faecal coliform bacteria additionally produce indole and liberate fluorochrome from MUG. Theoretical considerations and practical experiences showed that the rate of false-positive as well as false-negative results was within a range of 10% if compared with reactions described in the literature. Some factors which may influence the results are discussed.
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A survey was undertaken on the occurrence of Protease in the human fecal flora and its coincidence with other well-documented enteropathogens such as Campylobacter, Salmonella, Shigella, Staphylococcus aureus, Yersinia, protozoa and rotavirus. A total of 2000 fecal specimens was investigated, 1000 from patients suffering from diarrhea and 1000 from healthy persons which served as controls. Proteus mirabilis was isolated more frequently from diarrhea cases than from healthy people. The difference was statistically significant (P less than 0.001). There was no correlation between its occurrence and isolation of Campylobacter, Salmonella, Shigella, Staphylococcus aureus, and protozoa. However, a questionable coincidence was found with rotavirus (P less than 0.2) and a more certain correlation with Yersinia enterocolitica (P less than 0.025). Proteus mirabilis in patients suffering from infection by other known enteropathogens was largely absent, suggesting that the organisms were independent causative agents of intestinal disorders. Notwithstanding this, they may additionally play a role as opportunists in enteric diseases due to some other pathogens.
Listeria monocytogenes, L. innocua, and L. seeligeri are ingested by Tetrahymena pyriformis. They are not killed but survive and lyse bacteriovorous protozoans after about 8 days. The question important in food processing whether intracellular L. monocytogenes are protected against pasteurization was investigated using a model of Tetrahymena containing previously ingested Listeria. The study showed that Tetrahymena containing Listeria are more susceptible against application of heat, but intracellular Listeria, phagocytized within Tetrahymena, are not more protected than free and extracellular bacteria. Therefore, a properly performed pasteurization, i.e. of milk, kills intracellular Listeria as fast as extracellular living organisms. Finally, the pathogenic L. monocytogenes showed a higher susceptibility and lower tolerance against application of heat than apathogenic L. innocua and L. seeligeri.
Regarding the quality of natural recreational water for bathing purposes, the limit values of E. coli and coliforms, faecal coliforms and totalcoliforms, respectively, laid down by the Council of the European Community, differ considerably from those prescribed by some Land Governments of the Federal Republic of Germany and by the DIN Standard Regulation No. 19643 entitled 'Processing and Disinfection of Water in Swimming Pools and Basins in Public Baths'. It must be concluded by virtue of hygienically plausible considerations, as well as for juridical reasons, that only the limit values of the EC-Code of Practice are authoritative, whereas standards of the DIN 19643, especially in respect of E. coli and coliforms, differing from the limit values of the EC-Code of Practice are not legally binding. Moreover, coliforms are not relevant for controlling swimming pools and basins in public baths. Staphylococcus aureus should therefore be investigated instead whereas other parameters such as E. coli, Pseudomonas aeruginosa, and total colony counts are good markers. However, the standards of these organisms as established by the DIN 19643 are more in line with technical practibility and biased aesthetics instead of the hygienically important prinicple of commensurability.
Collaborative studies of comparative investigations of standardized drinking water were performed within Lower Saxony since two years. The results show that the precision of the measurements even in the reference laboratories is substantially less than the measuring tolerance given by the Federal Drinking Water Regulation. Therefore, the standard deviation of the different parameters, variation coefficients, respectively, are to be used for the internal statistical quality control as well as for the judgment of water. Protests against drinking water, bath water, respectively, due to parameters exceeded should be made just if the guide or imperative values are exceeded by the threefold standard deviation experimentally determined in reference laboratories.
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