Validation of a Loop-Mediated Isothermal Amplification-Based Kit for the Detection of in Environmental Samples According to ISO/TS 12869:2012.

Giorgia Caruso, Maria Anna Coniglio, Pasqualina Lagan��, Teresa Fasciana, Giuseppe Arcoleo, Ignazio Arrigo, Paola Di Carlo, Mario Palermo, Anna Giammanco
Author Information
  1. Giorgia Caruso: U.O.C. of Microbiology and Virology, ARNAS "Civico Di Cristina and Benfratelli", 90127 Palermo, Italy.
  2. Maria Anna Coniglio: Legionella Reference Laboratory, Department of Medical, Surgical Sciences and Advanced Technologies "G.F. Ingrassia", University of Catania, Via Santa Sofia 87, 95123 Catania, Italy.
  3. Pasqualina Lagan��: Legionella Reference Laboratory, University of Messina, 98125 Messina, Italy. ORCID
  4. Teresa Fasciana: Legionella Reference Laboratory, University of Palermo, 90127 Palermo, Italy.
  5. Giuseppe Arcoleo: Enbiotech s.r.l., Via Aquileia 34, 90144 Palermo, Italy.
  6. Ignazio Arrigo: Legionella Reference Laboratory, University of Palermo, 90127 Palermo, Italy. ORCID
  7. Paola Di Carlo: Department of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties, University of Palermo, 90127 Palermo, Italy. ORCID
  8. Mario Palermo: Sicilian Health Department, Public Health and Environmental Risks Service, 90127 Palermo, Italy.
  9. Anna Giammanco: Legionella Reference Laboratory, University of Palermo, 90127 Palermo, Italy.

Abstract

is a freshwater opportunistic pathogen and the leading cause of severe pneumonia known as Legionnaires' disease. It can be found in all water systems and survives in biofilms, free-living amoebae, and a wide variety of facilities, such as air conditioning and showers in hospitals, hotels and spas. The reference cultural method allows for the isolation and identification in many days, and in addition, it does not detect viable but rather non-culturable bacteria, increasing the risk of infection. In this context, a new LAMP-based (loop-mediated isothermal amplification) kit was developed, allowing for the rapid, sensitive, and labor-saving detection of . The kit, " Glow", was validated according to ISO/TS 12869:2012, testing sensitivity, inclusivity and exclusivity, and kit robustness. Sensitivity showed that the " Glow" kit can detect up to 28 plasmid copies/��L. Robustness tests showed consistent results, with both contamination levels and the matrices used giving reproducible results. Furthermore, real samples were evaluated to compare the performance of the two methods. The LAMP kit " Glow" proved a useful option for the rapid, efficient, and labor-saving screening of different typologies of water samples, offering significant advantages over the traditional method, as it is characterized by a high sensitivity, ease of use for laboratory testing, and a large reduction in analysis time, making it an asset to official controls.

Keywords

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