Rapid and reliable identification of waterborne Legionella species by MALDI-TOF mass spectrometry

Detection and enumeration of Legionella bacteria in drinking water is regulated in Germany by ISO 11731-2. The mandatory method for species identification employs parallel subculturing of suspicious colonies on selective media requiring the handling of a large number of cultivation plates. After changes to the drinking water quality regulation in Germany in 2012 the demand for Legionella contamination testing increased drastically. A more reliable, faster and less laborious method for species identification is therefore desirable. Matrix-assisted laser desorption ionization followed by time of flight detection mass spectrometry (MALDI-TOF MS) promises an accelerated identification of bacteria with high reliability and reduced expenditure. Our study shows that MS based species identification results are in full concordance with cultural and biochemical detection and differentiation and that valuable additional information can be gained, even though the ISO regulation demands an extended incubation period for primary bacterial cultures that is actually in contrast to the prerequisites of the MALDI Biotyper system. In addition, the established identification algorithm is very economical and improves time-to-result. Based on our findings, the amendment of MALID-TOF MS identification to ISO11731-2 as an alternative identification method should be taken into consideration. (C) 2016 Elsevier B.V. All rights reserved

Legionella contamination in warm water systems: A species-level survey

Therefore, in many countries it is mandatory to monitor warm water systems for their presence. The method of examination in Germany is regulated by guideline ISO 11731 and DIN EN ISO 11731-2, and the results are reported as concentration of Legionella spp. Only limited information is available on the presence of individual species of Legionellae in the examined systems, since most investigations and research focus solely on Legionella pneuntophila as the most important human pathogen. In this study 76,220 samples obtained from 13,397 warm water systems originating from 24 different zip code districts covering an area of more than 71,0001km(2) in southern Germany were examined. This resulted in the identification of 47,924 Legionella isolates to the species level using a MALDI-TOF mass spectrometry-based method. Legionella species distribution was analyzed with respect to warm water system type, geographic region (defined as zip code district) and temperature during sample taking. Overall, 20.7% of the samples were found positive for Legionella species and 14 different species of Legionella were recovered. These were not equally present throughout the geographic area investigated, but instead an individual regional diversity of Legionella species was observed for the examined zip code districts. Although Legionella pnewnophila represented 84% of all contaminations found, depending on the geographical region its proportion varied substantially between 57.5% and 91.2%. The occurrence of other species was also of importance since they accounted for up to 42% of contaminations regionally, with Legionella londiniensis being most prominent representing up to 38.8% of recovered colonies. In addition, the influence of temperature on the individual species was disparate, but the temperature range between 50 degrees C and 59 degrees C was identified as the optimal condition for facilitating emergence of the majority of recovered Legionella species. The identification of Legionella to the species level by MALDI-TOF allowed for a more concise depiction of the regional distribution and the ecology of this genus, and may be of additional value when counter measures need to be initiated

Colistin- and carbapenem-resistant Klebsiella oxytoca harboring blaVIM-2 and an insertion in the mgrB gene isolated from blood culture

A carbapenemase-producing colistin-resistant Klebsiella oxytoca isolate was recovered from a blood culture of a female patient without previous report of risk factors to obtain multidrug-resistant Gram-negative bacilli. A combination of biochemical and molecular methods was used to identify the resistance mechanism of this isolate. Carbapenemase production was mediated by Verona integron-encoded metallo-S-lactamase (VIM)-2. Colistin resistance was not due to plasmid- borne mcr-1 gene, but we found an integration of IS5-like sequence in the mgrB gene of K. oxytoca. This gene is known to be an important regulator of the PhoPQtwo-component system, and the disruption of this gene is most likely the cause of lipid A modification resulting in colistin resistance of our isolate. To the best of our knowledge this constitutes the first report of a carbapenemase-producing K. oxytoca with colistin resistance, a case that demonstrates the limited treatment options for infections with multidrug-resistant organisms. (C) 2017 Elsevier GmbH. All rights reserved