The bacterial flora of non-carbonated, natural mineral water from the springs to reservoir and glass and plastic bottles

Abstract

Quantitative and qualitative determinations of the bacterial flora of non-carbonated natural mineral water at the most important steps during bottling at a large water source yielded the following results: (i) Colony counts (on 1 :10 diluted plate count agar, incubated at 20°C for 14 days) for water of the five springs and the mixed water were less than 1 to 4 cfu ml −1. The Gram-negative bacterial flora ( n = 50 isolates) showed a very different but constant spring specific species distributions with predominance of either eutrophic fluorescent pseudomonads, oligotrophic non-fluorescent pseudomonads or oligotrophic yellow bacteria. (ii) In the reservoir and immediately after bottling the counts were in the range of 10 cfu m1 −1. But nearly 30% of the species of the spring water were no longer detectable and there was a significant increase of Gram-positive bacteria. (iii) After 1 week of storage at 2°C colony counts of more than 10 5 cfu ml −1 were found in plastic bottles, but only about 10 4 cfu ml −1 in glass bottles Besides, a very distinct change of the composition of the microflora occurred. In glass bottles slow-growing oligotrophic non-fluorescent pseudomonads, yellow bacteria and Acinetobacter predominated. In plastic bottles fast-growing eutrophic and mesotrophic fluorescent pseudomonads, Flexibacter and Acinebacter were dominating. In mineral water, bottled into thoroughly cleaned glass bottles, colony counts of more than 10 5 cfu ml −1 were found within 4 days. In bottles, cleaned mechanically as usual, the increase was significantly slower with a maximum of only 5 × 10 3 cfu ml −1 after 8 days. The results of inoculation experiments in sterile filtered mineral and distilled water led to the suggestion that the difference between the two types of bottles is caused firstly by an inhibition of growth due to residues of cleaning detergents in the glass bottles. Growth promotion by dissolved organic substances in the plastic bottles only played a minor role. After repairing of the pump at a depth of 300 m in a warm mineral water spring, the colony counts at 20, 37 and 42°C on 1: 10 diluted and normal plate count agar increased beyond the limits required by the EC directive for mineral water stored a month. Then colony counts decreased slowly and reached the initial level after 1 year, except for the colony counts 1 : 10 diluted agar at 20°C which stabilized at a relatively high number and a significant alteration of the microflora. Before the repair of the pump, the flora consisted exclusively of non-fluorescent pseudomonads, whereas non-motile and motile yellow bacteria were found afterwards.