Comparison of a biochemical test system with mass spectrometry for identification of bacteria pertinent to a freshwater stream environment

The amphipod Gammarus fossarum, stone crayfish Austropotamobius torrentium, an aquatic prosobranch gastropod mollusk Bithynia tentaculata, water from a pristine stream of the Nature Park Medvednica (Croatia), as well as scrapings from the sedimentary rocks, were sampled for microbiological evaluation. Rapid identification of environmental bacteria via commercial phenotypic tests allows for a wide choice of tests selection, and API 20E, an identification system for Enterobacteriaceae and other non-fastidious Gram-negative rods developed for clinical specimens, seems to be increasingly used for the identification of water-borne pathogens. However, due to several shortcomings of this system, such as the wrong identifications and the need of comparison with the diagnostic schemes based on reactions in conventional phenotypic tests, more advanced methods for identification are sought after. Therefore, a mass spectrometry MS-based method that focuses on the ribosomal proteins as a proteotypic system to utilize MALDI-TOF was used to achieve a highly accurate identification based on molecules that are translated from DNA but not subject to the expression variability seen in phenotypic methods. An excellent accordance of 57% was observed between the two methods in identification to the genus (Pseudomonas, Aeromonas, Shewanella spp.), while no correspondence was noted in identification to the species. Although this study furnished more accurate information on the bacterial populations in a surveyed freshwater stream (prevalence of Aeromonas and Pseudomonas spp.), and better clarified the existing relationships between the bacterial flora in tested samples, even a high-precision system had difficulties distinguishing between the species of the genus. Therefore, although implementation of MALDI-TOF MS can be a first step for identification which shortens the turnaround time, other methods for rapid analysis are warranted in order to obtain comparison of data and achieve an unambiguous identification.