Studies on Detection Methods for Legionella Species from Environmental Water

Abstract

We investigated selective cultivation media and previous treatments of samples suitable for detection of Legionella species from environmental water and for elimination of co-existing microbes which gave rise to an interference with the evaluation of Legionella sp. growth. Twenty thousand U of polymyxin B (PL-B)/ml and 100 micrograms of oxytetracycline (OTC)/ml seem to be useful as additives to MWY selective agar medium. Both antibiotics markedly inhibited the growth of co-existing microbes with almost no influence on the growth of Legionella sp. In the studies on the resistance of 8 strains of Legionella sp., 24 strains of co-existing microbes and 2 standard strains of Staphylococcus aureus and Escherichia coli to acid treatment (0.2 M HCl-KCl, pH 2.2, 25 degrees C, 4 minutes) and heating (50 degrees C, 20 minutes), acid treatment or heating alone showed no inhibition on the growth of almost all strains examined. However, combination with acid treatment after heating resulted in an apparent extinction of almost all microbes except for Legionella sp., Seven strains from co-existing microbes showed an apparent growth inhibition against 8 strains of Legionella sp. with different serotypes and were all identified as Pseudomonas aeurginosa, which were all eliminated by means of the combination with acid treatment after heating. From these results, it was concluded that the combined pre-treatment of water samples with acid after heating and the addition of PL-B and OTC into the selective cultivation medium is an useful method for detection of Legionella sp. from environmental water.

Legionella: from environmental habitats to disease pathology, detection and control

“…Often, pre-enrichment with various inhibitors is needed to eliminate populations of other bacteria (Roberts et al, 1987;Kusnetsov et al, 1994a). Kasuga et al (1999), for example, recently reported that combined pretreatment with acid after heating and the addition of polymyxin B and oxytetracycline into the selective cultivation medium is a useful method for the detection of Legionella sp. from environmental water samples.…”

Interpreting the Results of the Conventional Plate Culture and Gene Detection Methods for <i>Legionella</i> Detection in Environmental Water Samples

“…Combining these pretreatments is also possible. In this case, heat treatment is performed first, and then samples are cooled to room temperature and treated with acid buffer (Kasuga et al, 1999;Inoue et al, 2014) .…”

Legionellaprevalence and risk of legionellosis in Japanese households

“…Swab samples were suspended in 5 ml of 50-fold-diluted PBS. The suspension was then subjected to heat treatment at 50°C for 20 min according to the methods of Kasuga et al [13] followed by acid buffer treatment in HCl-KCl buffer (pH 2·2) for 4 min according to the methods of Bopp et al [14]. The samples were subsequently inoculated onto GVPC agar (Oxoid, Hampshire, UK) plates by the spread plate method, and the plates were incubated at 36°C for at least 7 days.…”