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Microorganisms 2017, 5(3), 62; doi:10.3390/microorganisms5030062

Diagnostic Value of Endotracheal Aspirates Sonication on Ventilator-Associated Pneumonia Microbiologic Diagnosis

1
Centro de Investigación Biomedica En Red-Enfermedades Respiratorias (CibeRes, CB06/06/0028), Institut d’Investigacions Biomèdiques Agust Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
2
Department of Medicine, School of Medicine, University of Barcelona, 08036 Barcelona, Spain
3
Pulmonary and Critical Care Unit, Respiratory Institute, Hospital Clinic, 08036 Barcelona, Spain
4
Dipartimento Scienze Chirurgiche e Diagnostiche Integrate (DISC), Università degli Studi di Genova, 16132 Genova, Italy
5
Respiratory Unit, Ospedale Garbagnate Salvini, Azienda Socio Sanitaria Territoriale Rhodense, 20024 Garbagnate Milanese, Italy
6
Department of Anesthesiology and Intensive Care Medicine, Humanitas Clinical and Research Center, Via Manzoni 56, 20089 Rozzano, Milano, Italy
7
Department of Anesthesiology and Intensive Care Medicine, Università Cattolica del Sacro Cuore, Fondazione Policlinico Universitario Agostino Gemelli, 00135 Rome, Italy
8
Barcelona Institute for Global Health (ISGlobal) Hospital Clínic, 08036 Barcelona, Spain
9
Department of Basic Clinical Practice, School of Medicine, University of Barcelona, 08036 Barcelona, Spain
10
Department of Pathophysiology and Transplantation, Università degli Studi di Milano, 20122 Milan, Italy
11
Department of Internal Medicine, Respiratory Unit and Cystic Fibrosis Adult Center, Fondazione IRCCS Cá Granda, Ospedale Maggiore Policlinico, 20122 Milan, Italy
These authors contributed equally to this work.
*
Authors to whom correspondence should be addressed.
Received: 10 August 2017 / Revised: 15 September 2017 / Accepted: 17 September 2017 / Published: 20 September 2017
(This article belongs to the Special Issue Microbial Biofilms and Human Infections)
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Abstract

Microorganisms are able to form biofilms within respiratory secretions. Methods to disaggregate such biofilms before utilizing standard, rapid, or high throughput diagnostic technologies may aid in pathogen detection during ventilator associated pneumonia (VAP) diagnosis. Our aim was to determine if sonication of endotracheal aspirates (ETA) would increase the sensitivity of qualitative, semi-quantitative, and quantitative bacterial cultures in an animal model of pneumonia caused by Pseudomonas aeruginosa or by methicillin resistant Staphylococcus aureus (MRSA). Material and methods: P. aeruginosa or MRSA was instilled into the lungs or the oropharynx of pigs in order to induce severe VAP. Time point assessments for qualitative and quantitative bacterial cultures of ETA and bronchoalveolar lavage (BAL) samples were performed at 24, 48, and 72 h after bacterial instillation. In addition, at 72 h (autopsy), lung tissue was harvested to perform quantitative bacterial cultures. Each ETA sample was microbiologically processed with and without applying sonication for 5 min at 40 KHz before bacterial cultures. Sensitivity and specificity were determined using BAL as a gold-standard. Correlation with BAL and lung bacterial burden was also determined before and after sonication. Assessment of biofilm clusters and planktonic bacteria was performed through both optical microscopy utilizing Gram staining and Confocal Laser Scanning Microscopy utilizing the LIVE/DEAD®BacLight kit. Results: 33 pigs were included, 27 and 6 from P. aeruginosa and MRSA pneumonia models, respectively. Overall, we obtained 85 ETA, 69 (81.2%) from P. aeruginosa and 16 (18.8%) from MRSA challenged pigs. Qualitative cultures did not significantly change after sonication, whereas quantitative ETA cultures did significantly increase bacterial counting. Indeed, sonication consistently increased bacterial burden in ETAs at 24, 48, and 72 h after bacterial challenge. Sonication also improved sensitivity of ETA quantitative cultures and maintained specificity at levels previously reported and accepted for VAP diagnosis. Conclusion: The use of sonication in ETA respiratory samples needs to be clinically validated since sonication could potentially improve pathogen detection before standard, rapid, or high throughput diagnostic methods used in routine microbial diagnostics. View Full-Text
Keywords: biofilm; endotracheal aspirate; sonication; ventilator associated pneumonia; animal model; Pseudomonas aeruginosa; Staphylococcus aureus biofilm; endotracheal aspirate; sonication; ventilator associated pneumonia; animal model; Pseudomonas aeruginosa; Staphylococcus aureus
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Fernández-Barat, L.; Motos, A.; Ranzani, O.T.; Bassi, G.L.; Aguilera Xiol, E.; Senussi, T.; Travierso, C.; Chiurazzi, C.; Idone, F.; Muñoz, L.; Vila, J.; Ferrer, M.; Pelosi, P.; Blasi, F.; Antonelli, M.; Torres, A. Diagnostic Value of Endotracheal Aspirates Sonication on Ventilator-Associated Pneumonia Microbiologic Diagnosis. Microorganisms 2017, 5, 62.

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