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Drinking water can be exposed to different biological contaminants from the source, through the pipelines, until reaching the final consumer or industry. Some of these are pathogenic bacteria and viruses which may cause important gastrointestinal or systemic diseases. The microbiological quality of drinking water relies mainly in monitoring three indicator bacteria of faecal origin, Escherichia coli, Enterococcus faecalis and Clostridium perfringens, which serve as early sentinels of potential health hazards for the population. Here we describe the analysis of three chimeric fluorescent protein bullets as biosensor candidates for fast detection of E. coli in drinking water. Two of the chimeric proteins (based on GFP-hadrurin and GFP-pb5 chimera proteins) failed with respect to specificity and/or sensitivity, but the GFP-colS4 chimera protein was able to carry out specific detection of E. coli in drinking water samples in a procedure encompassing about 8 min for final result and this biosensor protein was able to detect in a linear way between 20 and 103 CFU of this bacterium. Below 20 CFU, the system cannot differentiate presence or absence of the target bacterium. The fluorescence in this biosensor system is provided by the GFP subunit of the chimeric protein, which, in the case of the better performing sensor bullet, GFP-colS4 chimera, is covalently bound to a flexible peptide bridge and to a bacteriocin binding specifically to E. coli cells. Once bound to the target bacteria, the excitation step with 395 nm LED light causes emission of fluorescence from the GFP domain, which is amplified in a photomultiplier tube, and finally this signal is converted into an output voltage which can be associated with a CFU value and these data distributed along mobile phone networks, for example. This method, and the portable fluorimeter which has been developed for it, may contribute to reduce the analysis time for detecting E. coli presence in drinking water.

Running water habitats are among the most altered aquatic systems by human activities driving an increase in the organic components and the associated bacterial load as well. To contribute in improving the monitoring activities in running waters, here we tested the validity of the new Micro Biological Survey (MBS) method to specifically assess the bacterial load in running waters focusing on Total Viable Counts (at 22°C and 37°C) and Escherichia coli (at 44°C) in order to propose a new prognostic tool for watercourses. MBS method is an alternative colorimetric method for counting bacterial load in water and food samples that is easy to use and leads to a reliable and simple interpretation of results, being also faster and less expensive than traditional methods. Then, we compared MBS with the traditionally used reference method for the bacterial load, and with the most used biotic index for Italian watercourses based on the benthic invertebrates: the Extended Biotic Index (EBI). The last comparison was performed to validate the use of MBS in biomonitoring activities since the benthic invertebrate multi-species assemblage (and then EBI) alter own structure mainly depending on the organic component variation. During the first part of the study, the assessment of both linearity (regressions among bacterial concentrations) and accuracy (significant correlation between a measured value and a value used as reference) confirmed the validity of the MBS method. Second, the linear regressions between the three investigated microbial parameters vs. both physical-chemical descriptors and EBI, revealed the usefulness of MBS as a valid tool for routine microbiological analyses involved in rapid and easy field monitoring activities. This represents the first attempt to evaluate the river microbial status by exploiting the innovative MBS on running waters to propose it as new valuable monitoring tool in the biomonitoring field.

Legionella spp. is considered an emerging microorganism involved in aquatic environments contamination and cause of Legionnaires' disease. The aims of the study are to evaluate the level of contamination of Legionella spp. in the water system of the largest Hospital of Apulia region during a 4-year surveillance and to establish, by molecular method, the presence of a predominant genotype of L. pn. sg 1. The results showed that Legionella spp. was present in 36% of water samples with Legionella pneumophila serogroup 1 (L. pn. sg 1) the most prevalent species and serogroup and the wards most contaminated are the high risk units. In addition, despite four main clones of L. pn. sg 1 were identified, a predominant genotype existed. In conclusion the study demonstrates the necessity for periodic evaluation on hospitals water system to assess the potential contamination of Legionella spp., performing decontamination in the presence of bacterial contamination, even low, in particular in high risk wards. Moreover, the switching of the disinfection methods may be suggested in order to prevent resistance phenomenon by some L. pn. sg 1 clones.