Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients
<i>Staphylococcus aureus</i> is a commensal and frequent colonizer of the upper respiratory tract. When mechanical ventilation disrupts natural defenses, <i>S. aureus</i> is frequently isolated from the lower airways, but distinguishing between colonization and infection is d...
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MDPI AG
2021-02-01
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| Series: | Toxins |
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| Online Access: | https://www.mdpi.com/2072-6651/13/2/122 |
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| author | Alicia Lacoma Maisem Laabei Jose Francisco Sánchez-Herrero Bernadette Young Gerard Godoy-Tena Meissiner Gomes-Fernandes Lauro Sumoy Oriol Plans Fernando Arméstar Cristina Prat |
| author_facet | Alicia Lacoma Maisem Laabei Jose Francisco Sánchez-Herrero Bernadette Young Gerard Godoy-Tena Meissiner Gomes-Fernandes Lauro Sumoy Oriol Plans Fernando Arméstar Cristina Prat |
| author_sort | Alicia Lacoma |
| collection | DOAJ |
| description | <i>Staphylococcus aureus</i> is a commensal and frequent colonizer of the upper respiratory tract. When mechanical ventilation disrupts natural defenses, <i>S. aureus</i> is frequently isolated from the lower airways, but distinguishing between colonization and infection is difficult. The objectives of this study were (1) to investigate the bacterial genome sequence in consecutive isolates in order to identify changes related to the pathological adaptation to the lower respiratory tract and (2) to explore the relationship between specific phenotypic and genotypic features with the patient’s study group, persistence of the clinical isolate and clinical outcome. A set of 94 clinical isolates were selected and corresponded to 34 patients that were classified as having pneumonia (10), tracheobronchitis (11) and bronchial colonization (13). Clinical strains were phenotypically characterized by conventional identification and susceptibility testing methods. Isolates underwent whole genome sequencing using Illumina HiSeq4000. Genotypic characterization was performed with an in-house pipeline (BacterialTyper). Genomic variation arising within-host was determined by comparing mapped sequences and de novo assemblies. Virulence factors important in staphylococcal colonization and infection were characterized using previously established functional assays. (1) Toxin production was assessed using a THP-1 cytotoxicity assay, which reports on the gross cytotoxicity of individual isolates. In addition, we investigated the expression of the major virulence factor, alpha-toxin (Hla) by Western blot. (2) Adhesion to the important extracellular matrix molecule, fibronectin, was determined using a standardized microtitre plate assay. Finally, invasion experiments using THP-1 and A539 cell lines and selected clinical strains were also performed. Repeated isolation of <i>S. aureus</i> from endotracheal aspirate usually reflects persistence of the same strain. Within-host variation is detectable in this setting, but it shows no evidence of pathological adaptation related to virulence, resistance or niche adaptations. Cytotoxicity was variable among isolates with 14 strains showing no cytotoxicity, with these latter presenting an unaltered Fn binding capacity. No changes on cytotoxicity were reported when comparing study groups. Fn binding capacity was reported for almost all strains, with the exception of two strains that presented the lowest values. Strains isolated from patients with pneumonia presented a lower capacity of adhesion in comparison to those isolated during tracheobronchitis (<i>p</i> = 0.002). Hla was detected in 71 strains (75.5%), with most of the producer strains in pneumonia and bronchial colonization group (<i>p</i> = 0.06). In our cohort, Hla expression (presence or absence) in sequential isolates was usually preserved (70%) although in seven cases the expression varied over time. No relationship was found between low cytotoxicity and intracellular persistence in invasion experiments. In our study population, persistent <i>S. aureus</i> isolation from airways in ventilated patients does not reflect pathological adaptation. There is an important diversity of sequence types. Cytotoxicity is variable among strains, but no association with study groups was found, whereas isolates from patients with pneumonia had lower adhesion capability. Favorable clinical outcome correlated with increased bacterial adhesion in vitro. Most of the strains isolated from the lower airways were Hla producers and no correlation with an adverse outcome was reported. The identification of microbial factors that contribute to virulence is relevant to optimize patient management during lower respiratory tract infections. |
| first_indexed | 2024-03-09T05:22:40Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 2072-6651 |
| language | English |
| last_indexed | 2024-03-09T05:22:40Z |
| publishDate | 2021-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Toxins |
| spelling | doaj.art-e4a35b37abf84e958999d27feb5e32622023-12-03T12:39:26ZengMDPI AGToxins2072-66512021-02-0113212210.3390/toxins13020122Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated PatientsAlicia Lacoma0Maisem Laabei1Jose Francisco Sánchez-Herrero2Bernadette Young3Gerard Godoy-Tena4Meissiner Gomes-Fernandes5Lauro Sumoy6Oriol Plans7Fernando Arméstar8Cristina Prat9Microbiology Department, Hospital Universitari Germans Trias i Pujol, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, SpainDepartment of Biology and Biochemistry, University of Bath, Bath BA2 7AY, UKHigh Content Genomics and Bioinformatics Unit, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, 08916 Badalona, SpainNuffield Department of Medicine, University of Oxford, John Radcliffe Hospital, Oxford OX3 9DU, UKMicrobiology Department, Hospital Universitari Germans Trias i Pujol, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, SpainMicrobiology Department, Hospital Universitari Germans Trias i Pujol, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, SpainHigh Content Genomics and Bioinformatics Unit, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, 08916 Badalona, SpainIntensive Care Unit, Hospital Universitari Germans Trias i Pujol, Institut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, SpainIntensive Care Unit, Hospital Universitari Germans Trias i Pujol, Institut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, SpainMicrobiology Department, Hospital Universitari Germans Trias i Pujol, Institut d’Investigació en Ciències de la Salut Germans Trias i Pujol, Universitat Autònoma de Barcelona, 08916 Badalona, Spain<i>Staphylococcus aureus</i> is a commensal and frequent colonizer of the upper respiratory tract. When mechanical ventilation disrupts natural defenses, <i>S. aureus</i> is frequently isolated from the lower airways, but distinguishing between colonization and infection is difficult. The objectives of this study were (1) to investigate the bacterial genome sequence in consecutive isolates in order to identify changes related to the pathological adaptation to the lower respiratory tract and (2) to explore the relationship between specific phenotypic and genotypic features with the patient’s study group, persistence of the clinical isolate and clinical outcome. A set of 94 clinical isolates were selected and corresponded to 34 patients that were classified as having pneumonia (10), tracheobronchitis (11) and bronchial colonization (13). Clinical strains were phenotypically characterized by conventional identification and susceptibility testing methods. Isolates underwent whole genome sequencing using Illumina HiSeq4000. Genotypic characterization was performed with an in-house pipeline (BacterialTyper). Genomic variation arising within-host was determined by comparing mapped sequences and de novo assemblies. Virulence factors important in staphylococcal colonization and infection were characterized using previously established functional assays. (1) Toxin production was assessed using a THP-1 cytotoxicity assay, which reports on the gross cytotoxicity of individual isolates. In addition, we investigated the expression of the major virulence factor, alpha-toxin (Hla) by Western blot. (2) Adhesion to the important extracellular matrix molecule, fibronectin, was determined using a standardized microtitre plate assay. Finally, invasion experiments using THP-1 and A539 cell lines and selected clinical strains were also performed. Repeated isolation of <i>S. aureus</i> from endotracheal aspirate usually reflects persistence of the same strain. Within-host variation is detectable in this setting, but it shows no evidence of pathological adaptation related to virulence, resistance or niche adaptations. Cytotoxicity was variable among isolates with 14 strains showing no cytotoxicity, with these latter presenting an unaltered Fn binding capacity. No changes on cytotoxicity were reported when comparing study groups. Fn binding capacity was reported for almost all strains, with the exception of two strains that presented the lowest values. Strains isolated from patients with pneumonia presented a lower capacity of adhesion in comparison to those isolated during tracheobronchitis (<i>p</i> = 0.002). Hla was detected in 71 strains (75.5%), with most of the producer strains in pneumonia and bronchial colonization group (<i>p</i> = 0.06). In our cohort, Hla expression (presence or absence) in sequential isolates was usually preserved (70%) although in seven cases the expression varied over time. No relationship was found between low cytotoxicity and intracellular persistence in invasion experiments. In our study population, persistent <i>S. aureus</i> isolation from airways in ventilated patients does not reflect pathological adaptation. There is an important diversity of sequence types. Cytotoxicity is variable among strains, but no association with study groups was found, whereas isolates from patients with pneumonia had lower adhesion capability. Favorable clinical outcome correlated with increased bacterial adhesion in vitro. Most of the strains isolated from the lower airways were Hla producers and no correlation with an adverse outcome was reported. The identification of microbial factors that contribute to virulence is relevant to optimize patient management during lower respiratory tract infections.https://www.mdpi.com/2072-6651/13/2/122<i>Staphylococcus aureus</i>mechanical ventilationpneumoniatoxicityadhesionpersistence |
| spellingShingle | Alicia Lacoma Maisem Laabei Jose Francisco Sánchez-Herrero Bernadette Young Gerard Godoy-Tena Meissiner Gomes-Fernandes Lauro Sumoy Oriol Plans Fernando Arméstar Cristina Prat Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients Toxins <i>Staphylococcus aureus</i> mechanical ventilation pneumonia toxicity adhesion persistence |
| title | Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients |
| title_full | Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients |
| title_fullStr | Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients |
| title_full_unstemmed | Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients |
| title_short | Genotypic and Phenotypic Characterization of <i>Staphylococcus aureus</i> Isolates from the Respiratory Tract in Mechanically-Ventilated Patients |
| title_sort | genotypic and phenotypic characterization of i staphylococcus aureus i isolates from the respiratory tract in mechanically ventilated patients |
| topic | <i>Staphylococcus aureus</i> mechanical ventilation pneumonia toxicity adhesion persistence |
| url | https://www.mdpi.com/2072-6651/13/2/122 |
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