{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T12:56:17Z","timestamp":1791896177565,"version":"4.3.4"},"reference-count":27,"publisher":"MDPI AG","issue":"21","license":[{"start":{"date-parts":[[2019,10,26]],"date-time":"2019-10-26T00:00:00Z","timestamp":1572048000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Sensors"],"abstract":"<jats:p>Development of sensitive methods for the determination of E. coli bacteria contamination in water distribution systems is of paramount importance to ensure the microbial safety of drinking water. This work presents a new sensing platform enabling the fast detection of bacteria in field samples by using specific antibodies as the biorecognition element and dark field microscopy as the detection technique. The development of the sensing platform was performed using non-pathogenic bacteria, with the E. coli DH5\u03b1 strain as the target, and Bacillus sp. 9727 as the negative control. The identification of the captured bacteria was made by analyzing the dark field microscopy images and screening the detected objects by using object circularity and size parameters. Specificity tests revealed the low unspecific attachment of either E. coli over human serum albumin antibodies (negative control for antibody specificity) and of Bacillus sp. over E. coli antibodies. The system performance was tested using field samples, collected from a wastewater treatment plant, and compared with two quantification techniques (i.e., Colilert-18 test and quantitative polymerase chain reaction (qPCR)). The results showed comparable quantification capability. Nevertheless, the present method has the advantage of being faster, is easily adaptable to in-field analysis, and can potentially be extended to the detection of other bacterial strains.<\/jats:p>","DOI":"10.3390\/s19214652","type":"journal-article","created":{"date-parts":[[2019,10,28]],"date-time":"2019-10-28T04:44:31Z","timestamp":1572237871000},"page":"4652","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":12,"title":["Dark Field Microscopy-Based Biosensors for the Detection of E. coli in Environmental Water Samples"],"prefix":"10.3390","volume":"19","author":[{"given":"Rita","family":"La Spina","sequence":"first","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), Via E. Fermi 2749, 21027 Ispra, Varese, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3162-2983","authenticated-orcid":false,"given":"Diana C.","family":"Ant\u00f3nio","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), Via E. Fermi 2749, 21027 Ispra, Varese, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1931-1477","authenticated-orcid":false,"given":"Cloe","family":"Desmet","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), Via E. Fermi 2749, 21027 Ispra, Varese, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6367-561X","authenticated-orcid":false,"given":"Andrea","family":"Valsesia","sequence":"additional","affiliation":[{"name":"European Commission, Joint Research Centre (JRC), Via E. 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Licence: CC BY-NC-SA 3.0 IGO."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"307","DOI":"10.3390\/pathogens4020307","article-title":"Waterborne pathogens: Detection methods and challenges","volume":"4","author":"Jacques","year":"2015","journal-title":"Pathogens"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1007\/s13206-015-9208-9","article-title":"Recent trends in the detection of pathogenic Escherichia coli O157: H7","volume":"9","author":"Wu","year":"2015","journal-title":"BioChip J."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1056","DOI":"10.1111\/jam.13510","article-title":"On the challenges of detecting whole Staphylococcus aureus cells with biosensors","volume":"123","author":"Templier","year":"2017","journal-title":"J. Appl. Microbiol."},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Byrne, B., Stack, E., Gilmartin, N., and O\u2019Kennedy, R. (2009). Antibody-Based Sensors: Principles, Problems and Potential for Detection of Pathogens and Associated Toxins. Sensors, 9.","DOI":"10.3390\/s90604407"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"464","DOI":"10.1111\/jam.13508","article-title":"Absolute bacterial cell enumeration using flow cytometry","volume":"123","author":"Ou","year":"2017","journal-title":"J. Appl. Microbiol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"9721","DOI":"10.1021\/ac302471c","article-title":"Counting Bacteria Using Functionalised Gold Nanoparticles as the Light-Scattering Reporter","volume":"84","author":"Xu","year":"2012","journal-title":"Anal. Chem."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1007\/s11270-018-3834-x","article-title":"Label-Free Darkfield-Based Technique to Assist in the Detection of Giardia Cysts","volume":"229","author":"Belini","year":"2018","journal-title":"Water Air Soil Pollut."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1016\/j.bios.2017.08.025","article-title":"A low cost mobile phone dark-field microscope for nanoparticle-based quantitative studies","volume":"99","author":"Sun","year":"2018","journal-title":"Biosens. Bioelectron."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"14742","DOI":"10.1038\/s41598-017-15256-1","article-title":"Microscopic Differentiation of Plasmonic Nanoparticles for the Ratiometric Read-out of Target DNA","volume":"7","author":"Wu","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Wang, G., Wang, C., Yang, R., Liu, W., and Sun, S. (2017). A sensitive and stable surface plasmon resonance sensor based on monolayer protected silver film. Sensors, 17.","DOI":"10.3390\/s17122777"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1016\/0014-5793(77)80354-2","article-title":"The number of rRNA genes in Escherichia coli","volume":"79","author":"Kiss","year":"1977","journal-title":"FEBS Lett."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1016\/j.bios.2012.04.034","article-title":"Comparison of sensing strategies in SPR biosensor for rapid and sensitive enumeration of bacteria","volume":"37","author":"Torun","year":"2012","journal-title":"Biosens. Bioelectron."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"873","DOI":"10.1080\/01621459.1958.10501484","article-title":"The Estimation of the Parameters of a Linear Regression System Obeying Two Separate Regimes","volume":"53","author":"Quandt","year":"1958","journal-title":"J. Am. Stat. Assoc."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"730","DOI":"10.1080\/01621459.1978.10480085","article-title":"Estimating mixtures of normal distributions and switching regressions","volume":"73","author":"Quandt","year":"1978","journal-title":"J. Am. Stat. Assoc."},{"key":"ref_16","unstructured":"McLachlan, G., and Peel, D. (2005). Finite Mixture Models, Waley."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Mengersen, K.L., Robert, C., and Titterington, M. (2011). Mixtures: Estimation and Applications, Wiley.","DOI":"10.1002\/9781119995678"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/j.2517-6161.1977.tb01600.x","article-title":"Maximum likehood from Incomplete data via the EM algorithm","volume":"39","author":"Dempster","year":"1977","journal-title":"J. R. Stat. Soc. Serie B (Methodological)"},{"key":"ref_19","unstructured":"R. Core Team (2018). A Language and Environment for Statistical Computing, R Development Core Team."},{"key":"ref_20","unstructured":"Wickham, H., Fran\u00e7ois, R., Lionel, H., and M\u00fcller, K. (2019, October 26). Dplyr: A Grammar of Data Manipulation; R package version 0.7.8. Available online: https:\/\/dplyr.tidyverse.org\/."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"29","DOI":"10.18637\/jss.v032.i06","article-title":"Mixtools: An R Package for Analysing Mixture Models","volume":"32","author":"Benaglia","year":"2009","journal-title":"J. Stat. Softw."},{"key":"ref_22","unstructured":"Browne, R.P., Elsherbiny, A., and McNicholas, P.D. (2019, October 26). Mixture: Mixture Models for Clustering and Classification. R package version 1.5. Available online: https:\/\/rdrr.io\/cran\/mixture\/man\/mixture.html."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"221","DOI":"10.2166\/wh.2005.042","article-title":"Comparison of two methods for evaluating the quality of stored drinking water in Abidjan, Cote d\u2032Ivoire, and review of other comparisons in the literature","volume":"3","author":"Macy","year":"2005","journal-title":"J. Water Heath"},{"key":"ref_24","unstructured":"(2019, October 26). Commission Directive (EU) 2015\/1787 of 6 October 2015 amending Annexes II and III to Council Directive 98\/83\/EC on the Quality of Water Intended for Human Consumption. Available online: https:\/\/eur-lex.europa.eu\/legal-content\/EN\/TXT\/?uri=celex:32015L1787."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"7437","DOI":"10.1128\/AEM.00651-10","article-title":"Comparison of rapid quantitative PCR-Based and conventional culture-based methods for enumeration of enterococcus spp. and escherichia coli in recreational waters","volume":"76","author":"Noble","year":"2010","journal-title":"Appl. Environ. Microbiol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"203","DOI":"10.1089\/10665270050081478","article-title":"A greedy algorithm for aligning DNA sequences","volume":"7","author":"Zhang","year":"2000","journal-title":"J. Comput. Biol."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1757","DOI":"10.1093\/bioinformatics\/btn322","article-title":"Database indexing for production MegaBLAST searches","volume":"24","author":"Morgulis","year":"2008","journal-title":"Bioinformatics"}],"container-title":["Sensors"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8220\/19\/21\/4652\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T13:29:33Z","timestamp":1760189373000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8220\/19\/21\/4652"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2019,10,26]]},"references-count":27,"journal-issue":{"issue":"21","published-online":{"date-parts":[[2019,11]]}},"alternative-id":["s19214652"],"URL":"https:\/\/doi.org\/10.3390\/s19214652","relation":{},"ISSN":["1424-8220"],"issn-type":[{"value":"1424-8220","type":"electronic"}],"subject":[],"published":{"date-parts":[[2019,10,26]]}}}