Macrolide resistance and molecular typing of Mycoplasma pneumoniae infections during a 4 year period in Spain

Mycoplasma pneumoniae (MP) causes community-acquired pneumonia affecting mainly children, and tends to produce cyclic outbreaks. The widespread use of macrolides is increasing resistance rates to these antibiotics. Molecular tools can help in diagnosis, typing and resistance detection, leading to better patient management. To assess the MP genotypes and resistance pattern circulating in our area while comparing serological and molecular diagnosis of MP. Molecular and serological diagnosis of MP was performed in 821 samples collected in Badalona (Barcelona, Spain) from 2013 to 2017. Multiple locus variable number tandem repeat analysis (MLVA) and macrolide resistance detection by pyrosequencing were performed in those cases positive by PCR. Presence of respiratory viruses and relevant clinical data were also recorded. MP was detected in 16.8% of cases by PCR, with an overall agreement with serology of 76%. Eleven different MLVA types were identified, with 4-5-7-2 (50.1%) and 3-5-6-2 (29.2%) being the most abundant, with the latter showing a seasonal increase during the study. A total of 8% of the strains harboured a point substitution associated with macrolide resistance, corresponding mainly to an A2063G 23S rRNA mutation and directly related to previous macrolide therapy. Analysis of respiratory viruses showed viral coinfections in most cases. Serological and molecular tools combined could improve MP diagnosis and the analysis of its infection patterns. Macrolide resistance is associated with previous therapy. Given that MP pneumonia usually resolves spontaneously, it should be reconsidered whether antibiotic treatment is suitable for all cases

Accessory gene regulator (Agr) functionality in Staphylococcus aureus derived from lower respiratory tract infections

Altres ajuts: This work has been funded by the project PI13/01418 which is part of "Plan Nacional de I+D+I" and co-funded by ISCIII-Subdirección General de Evaluacioón and "Fondo Europeo de Desarrollo Regional"(FEDER).D. Domínguez-Villanueva is funded by "Plan Nacional de I+D+I" and co-funded by ISCIII-Subdirección General de Evaluación and "Fondo Europeo de Desarrollo Regional"(FEDER). M. Gomes-Fernandes is funded by CAPES Foundation, Ministry of Education of Brazil (Brasılia, Brazil). Maisem Laabei was supported by a joint ERS/SEPAR fellowship (LTRF2015).This work also received a grant from the Spanish Society of Pneumology and Thoracic Surgery (SEPAR054/2011).Objective. Characterization of Staphylococcus aureus clinical isolates derived from lower respiratory tract infections (LRTIs), and correlation between the functionality of the accessory gene regulator (Agr) and genotypic and phenotypic characteristics, clinical variables and clinical outcome. Methods. S aureus isolates derived from LRTIs and control groups (nasal carriage and bacteraemia) were genotyped using StaphyType DNA microarray. Agr activity was evaluated using the CAMP synergistic haemolysis assay and the Vesicle Lysis Test (VLT). Discordant strains were analysed by quantitative reverse- transcriptase real-time PCR (qRT-PCR). Results. Agr was functional in 79.7% and 84.5% of strains according to the CAMP and VLT assays respectively. Higher concordance with RNAIII expression measured by qRT-PCR was observed with the VLT assay (76.2%) compared with the CAMP assay (23.8%). No statistically significant differences were observed in Agr functionality between the study groups, nor the phenotypical/genotypical bacterial characteristics. No association between increased mortality/respiratory complications and Agr function was observed. Conclusions. Agr activity was high (82.2%) in isolates from LRTIs suggesting the importance of this global regulator in lower respiratory tract colonisation and infection. However, equally high Agr activity was observed in isolates derived from nasal carriage and bacteraemia, contradictory to previous observations. Agr functionality measured by the VLT assay was superior to CAMP assay

An engineered HIV-1 Gag-based VLP displaying high antigen density induces strong antibody-dependent functional immune responses

Antigen display on the surface of Virus-Like Particles (VLPs) improves immunogenicity compared to soluble proteins. We hypothesised that immune responses can be further improved by increasing the antigen density on the surface of VLPs. In this work, we report an HIV-1 Gag-based VLP platform engineered to maximise the presence of antigen on the VLP surface. An HIV-1 gp41-derived protein (Min), including the C-terminal part of gp41 and the transmembrane domain, was fused to HIV-1 Gag. This resulted in high-density MinGag-VLPs. These VLPs demonstrated to be highly immunogenic in animal models using either a homologous (VLP) or heterologous (DNA/VLP) vaccination regimen, with the latter yielding 10-fold higher anti-Gag and anti-Min antibody titres. Despite these strong humoral responses, immunisation with MinGag-VLPs did not induce neutralising antibodies. Nevertheless, antibodies were predominantly of an IgG2b/IgG2c profile and could efficiently bind CD16-2. Furthermore, we demonstrated that MinGag-VLP vaccination could mediate a functional effect and halt the progression of a Min-expressing tumour cell line in an in vivo mouse model

Performance characteristics of five antigen-detecting rapid diagnostic test (Ag-RDT) for SARS-CoV-2 asymptomatic infection: a head-to-head benchmark comparison.

BACKGROUND: Mass testing for early identification and isolation of infectious COVID-19 individuals is efficacious for reducing disease spread. Antigen-detecting rapid diagnostic tests (Ag-RDT) may be suitable for testing strategies; however, benchmark comparisons are scarce. METHODS: We used 286 nasopharyngeal specimens from unexposed asymptomatic individuals collected between December 2020 and January 2021 to assess five Ag-RDTs marketed by Abbott, Siemens, Roche Diagnostics, Lepu Medical, and Surescreen. RESULTS: For the overall sample, the performance parameters of Ag-RDTs were as follows: Abbott assay, sensitivity 38.6% (95%CI 29.1-48.8) and specificity 99.5% (97-100%); Siemens, sensitivity 51.5% (41.3-61.6) and specificity 98.4% (95.3-99.6); Roche, sensitivity 43.6% (33.7-53.8) and specificity 96.2% (92.4-98.5); Lepu, sensitivity 45.5% (35.6-55.8) and specificity 89.2% (83.8-93.3%); Surescreen, sensitivity 28.8% (20.2-38.6) and specificity 97.8% (94.5-99.4%). For specimens with cycle threshold (Ct) 99% and <50%, respectively. CONCLUSIONS: When screening unexposed asymptomatic individuals, two Ag-RDTs achieved sensitivity ≥80% for specimens with Ct<30 and specificity ≥96%. The estimated negative predictive value suggests the suitability of Ag-RDTs for mass screenings of SARS-CoV-2 infection in the general population

Noves aproximacions als estudis d’epidemiologia molecular i al diagnòstic de les infeccions causades per Staphylococcus aureus. Aportacions al coneixement dels factors de virulència i patogenicitat

En les últimes dècades Staphylococcus aureus s’ha consolidat com un dels microorganismes amb més capacitat de causar infeccions, donant lloc des de malalties lleus i localitzades, de curs benigne, fins a infeccions greus, disseminades, amb elevada mortalitat. En aquesta tesi, s’han abordat tant aspectes clínics i epidemiològics dels pacients amb infeccions documentades per S.aureus, en especial bacterièmia i infeccions de pell i parts toves, com aspectes relacionats amb el patogen. En aquest últim apartat, s’ha estudiat l’epidemiologia molecular de les soques, factors de virulència i alguns tests fenotípics que ens ajudaran a mesurar la seva expressió. Es van incloure un total de 293 bacterièmies per S.aureus. La fagotipia va permetre tipar el 37.5% de les soques SARM i el 43.3% dels SASM. La detecció del gen mecA va resultar positiva en 28 soques, en concordança amb les dades obtingudes de la sensibilitat antibiòtica per CMI. En l’anàlisi del tipus SCCmec de les soques SARM, 27 d’elles corresponien al SCCmecIVc i només una presentava el tipus SCCmec IVa i ACME negatiu. Mitjançant spa typing es va demostrar el predomini d’un grup clonal, CC002 (31%) englobant dos spa tipus predominants: t002, t067 en soques SASM i el predomini de CC067 (37.14%) incloent els spa tipus t0067 i t002 en SARM. Comparant els patrons d’electroforesi en camp polsant es van agrupar les soques SARM en les clones E7 i E8, també les més prevalents en el nostre medi. En l’estudi mitjançant microarrays de 102 d’aquestes soques es va observar un predomini del CC5 (38.2%), seguit del CC30 (21.6%). La distribució dels perfils al·lèlics del sistema regulador agr mostrava un predomini del agr II (46.1%). La mortalitat de la sèpsia per S.aureus es relaciona amb la susceptibilitat de l’hoste, característiques de la soca, característiques epidemiològiques i amb el temps transcorregut fins a l’inici d’un tractament antibiòtic adequat. Així, l’estudi de mètodes ràpids per la identificació i la detecció de sensibilitat a cloxacil·lina ( BinaxNow S.aureus / PBP 2a Test, GenomEra MRSA/SA Blood Culture, MicroPhage MRSA/MSSA Blood Culture Test) ens ha permès adequar el tractament antibiòtic en un 79.25% dels casos. L’origen de la bacterièmia en el 58’4% dels casos era nosocomial i el 45.7% d’origen en el catèter. El 80% dels malalts presentaven comorbiditats associades: 35.5% neoplàsia, 30.4% DM, 14% IRC i 13.7% tractament previ amb immunosupressor. Una quarta part van presentar bacterièmia complicada, i un 8.5% bacterièmia persistent. . D’altra banda, les infeccions de pell i parts toves per S.aureus resistents a meticil·lina d’origen comunitari (CA-SARM) representen al voltant del 20% en el nostre medi. La tipificació molecular d’aquestes soques va permetre demostrar que la majoria pertanyien a la soca USA300-ST8-SCCmec IVc i ACME negativa, variant de la clona USA300 (USA300-ST8-SCCmec IVa). No es va trobar cap associació entre la soca productora i la nacionalitat dels pacients. Les manifestacions clíniques dels pacients eren lleus i sense presència de complicacions, tant en els SASM com SARM. L’elevada prevalença de la Leucocidina de Panton Valentine (PVL) en soques SASM o SARM i la bona evolució, tot i el tractament antibiòtic inadequat, es correlaciona amb la hipòtesi que independentment de la sensibilitat antibiòtica, el drenatge de la lesió és necessari per la resolució de la infecció i posa de manifest la difícil valoració del paper de factors de virulència com la PVL en la patogènia d’aquestes infeccions.In recent decades, Staphylococcus aureus has become an important human pathogen causing a wide variety of diseases, ranging from uncomplicated infections to life-threatening septicaemia, thus leading to significant morbidity and mortality. This thesis recorded clinical and epidemiological characteristics from patients with S.aureus infections, especially bacteremia and skin-soft tissue infections as well as microorganism related factors. The second aim was to investigate the molecular typing and to characterize the virulence factors of S. aureus isolates and some phenotypic assays for its measurement. A total of 293 patients with bacteremia were included. Phage typing allowed to type the 37.5 % of MRSA strains and 43.3 % of MSSA. Regarding molecular characterization, mecA was detected in 28 strains, in accordance with the data of antibiotic susceptibility testing by microdilution method. SCCmec typing of SARM isolates showed that 27 of these were carrying SCCmecIVc and only in one case was found SCCmec IVa and ACME negative. Spa typing revealed the prevalent clonal complex CC002 (31%), enclosing two predominant spa types: t002, t067 in MSSA while the clonal complex CC067 (37.14%) including spa types t0067 and t002 was the predominant in MRSA strains. Genotyping by pulsed-field gel electrophoresis of these SARM strains grouped into two genotypes, E7 and E8. These profiles are the predominant clones in Spain. The application of microarrays for genotyping reflected that the most frequent CC was CC5 (38.2%) followed by CC30 (21.6%). Concerning the agr types, most isolates presented agr II (46.1%). Increased frequency of bacteraemia and maintenance of mortality rates are related to the susceptibility of the host, strain-specific features, epidemiological characteristics and the time from microbiological diagnosis to the starting of appropriate antibiotic treatment. The study of three diagnostic methods: BinaxNow S.aureus /MRSA PBP2, MicroPhage MRSA/ MSSA Blood Culture Test and GenomEra MRSA /SA Blood Culture for the direct identification and susceptibility S. aureus testing directly from positive blood cultures, allowed the early administration of appropriate antibiotic treatment in 79.25% of the cases. The origin of the bacteraemia was nosocomial in 58’4% of the cases and 45.7% were catheter related. Comorbidities were present in 80% of patients, being the most frequent neoplasia (35.5%), followed by diabetes mellitus (30.4%), end stage renal disease (14%) and immunosuppressive treatment (13.7%). Development of complications was present around 25% of the cases, and 8.5% of patients developed persistent bacteraemia. On the other hand, Community-Acquired methicillin resistant Staphylococcus aureus (CA-MRSA) causes approximately 20% of skin-soft tissue infections. A specific clone of CA-SARM, USA300-ST8-SCCmec IVc and characteristically lacking the ACME was the most prevalent in our environment, similar to USA300 (USA300-ST8-SCCmec Iva). We did not find any association between strain characteristics and nationality of the patients. All our patients presented an uncomplicated infection, both when MSSA and MRSA. In our study we reported a high prevalence of Panton Valentine leukocidin (PVL) producers among MSSA and MRSA and all patients presented a favourable outcome even though inappropriate antibiotic treatment was prescribed empirically. It is considered that the treatment of choice for these infections, irrespective of antibiotic susceptibility, is incision and drainage. The real pathogenic role of PVL it is still a subject of controversy, arising from results obtained from clinical studies

Noves aproximacions als estudis d'epidemiologia molecular i al diagnòstic de les infeccions causades per Staphylococcus aureus : aportacions al coneixement dels factors de virulència i patogenicitat /

En les últimes dècades Staphylococcus aureus s'ha consolidat com un dels microorganismes amb més capacitat de causar infeccions, donant lloc des de malalties lleus i localitzades, de curs benigne, fins a infeccions greus, disseminades, amb elevada mortalitat. En aquesta tesi, s'han abordat tant aspectes clínics i epidemiològics dels pacients amb infeccions documentades per S.aureus, en especial bacterièmia i infeccions de pell i parts toves, com aspectes relacionats amb el patogen. En aquest últim apartat, s'ha estudiat l'epidemiologia molecular de les soques, factors de virulència i alguns tests fenotípics que ens ajudaran a mesurar la seva expressió. Es van incloure un total de 293 bacterièmies per S.aureus. La fagotipia va permetre tipar el 37.5% de les soques SARM i el 43.3% dels SASM. La detecció del gen mecA va resultar positiva en 28 soques, en concordança amb les dades obtingudes de la sensibilitat antibiòtica per CMI. En l'anàlisi del tipus SCCmec de les soques SARM, 27 d'elles corresponien al SCCmecIVc i només una presentava el tipus SCCmec IVa i ACME negatiu. Mitjançant spa typing es va demostrar el predomini d'un grup clonal, CC002 (31%) englobant dos spa tipus predominants: t002, t067 en soques SASM i el predomini de CC067 (37.14%) incloent els spa tipus t0067 i t002 en SARM. Comparant els patrons d'electroforesi en camp polsant es van agrupar les soques SARM en les clones E7 i E8, també les més prevalents en el nostre medi. En l'estudi mitjançant microarrays de 102 d'aquestes soques es va observar un predomini del CC5 (38.2%), seguit del CC30 (21.6%). La distribució dels perfils al·lèlics del sistema regulador agr mostrava un predomini del agr II (46.1%). La mortalitat de la sèpsia per S.aureus es relaciona amb la susceptibilitat de l'hoste, característiques de la soca, característiques epidemiològiques i amb el temps transcorregut fins a l'inici d'un tractament antibiòtic adequat. Així, l'estudi de mètodes ràpids per la identificació i la detecció de sensibilitat a cloxacil·lina ( BinaxNow S.aureus / PBP 2a Test, GenomEra MRSA/SA Blood Culture, MicroPhage MRSA/MSSA Blood Culture Test) ens ha permès adequar el tractament antibiòtic en un 79.25% dels casos. L'origen de la bacterièmia en el 58'4% dels casos era nosocomial i el 45.7% d'origen en el catèter. El 80% dels malalts presentaven comorbiditats associades: 35.5% neoplàsia, 30.4% DM, 14% IRC i 13.7% tractament previ amb immunosupressor. Una quarta part van presentar bacterièmia complicada, i un 8.5% bacterièmia persistent. . D'altra banda, les infeccions de pell i parts toves per S.aureus resistents a meticil·lina d'origen comunitari (CA-SARM) representen al voltant del 20% en el nostre medi. La tipificació molecular d'aquestes soques va permetre demostrar que la majoria pertanyien a la soca USA300-ST8-SCCmec IVc i ACME negativa, variant de la clona USA300 (USA300-ST8-SCCmec IVa). No es va trobar cap associació entre la soca productora i la nacionalitat dels pacients. Les manifestacions clíniques dels pacients eren lleus i sense presència de complicacions, tant en els SASM com SARM. L'elevada prevalença de la Leucocidina de Panton Valentine (PVL) en soques SASM o SARM i la bona evolució, tot i el tractament antibiòtic inadequat, es correlaciona amb la hipòtesi que independentment de la sensibilitat antibiòtica, el drenatge de la lesió és necessari per la resolució de la infecció i posa de manifest la difícil valoració del paper de factors de virulència com la PVL en la patogènia d'aquestes infeccions.In recent decades, Staphylococcus aureus has become an important human pathogen causing a wide variety of diseases, ranging from uncomplicated infections to life-threatening septicaemia, thus leading to significant morbidity and mortality. This thesis recorded clinical and epidemiological characteristics from patients with S.aureus infections, especially bacteremia and skin-soft tissue infections as well as microorganism related factors. The second aim was to investigate the molecular typing and to characterize the virulence factors of S. aureus isolates and some phenotypic assays for its measurement. A total of 293 patients with bacteremia were included. Phage typing allowed to type the 37.5 % of MRSA strains and 43.3 % of MSSA. Regarding molecular characterization, mecA was detected in 28 strains, in accordance with the data of antibiotic susceptibility testing by microdilution method. SCCmec typing of SARM isolates showed that 27 of these were carrying SCCmecIVc and only in one case was found SCCmec IVa and ACME negative. Spa typing revealed the prevalent clonal complex CC002 (31%), enclosing two predominant spa types: t002, t067 in MSSA while the clonal complex CC067 (37.14%) including spa types t0067 and t002 was the predominant in MRSA strains. Genotyping by pulsed-field gel electrophoresis of these SARM strains grouped into two genotypes, E7 and E8. These profiles are the predominant clones in Spain. The application of microarrays for genotyping reflected that the most frequent CC was CC5 (38.2%) followed by CC30 (21.6%). Concerning the agr types, most isolates presented agr II (46.1%). Increased frequency of bacteraemia and maintenance of mortality rates are related to the susceptibility of the host, strain-specific features, epidemiological characteristics and the time from microbiological diagnosis to the starting of appropriate antibiotic treatment. The study of three diagnostic methods: BinaxNow S.aureus /MRSA PBP2, MicroPhage MRSA/ MSSA Blood Culture Test and GenomEra MRSA /SA Blood Culture for the direct identification and susceptibility S. aureus testing directly from positive blood cultures, allowed the early administration of appropriate antibiotic treatment in 79.25% of the cases. The origin of the bacteraemia was nosocomial in 58'4% of the cases and 45.7% were catheter related. Comorbidities were present in 80% of patients, being the most frequent neoplasia (35.5%), followed by diabetes mellitus (30.4%), end stage renal disease (14%) and immunosuppressive treatment (13.7%). Development of complications was present around 25% of the cases, and 8.5% of patients developed persistent bacteraemia. On the other hand, Community-Acquired methicillin resistant Staphylococcus aureus (CA-MRSA) causes approximately 20% of skin-soft tissue infections. A specific clone of CA-SARM, USA300-ST8-SCCmec IVc and characteristically lacking the ACME was the most prevalent in our environment, similar to USA300 (USA300-ST8-SCCmec Iva). We did not find any association between strain characteristics and nationality of the patients. All our patients presented an uncomplicated infection, both when MSSA and MRSA. In our study we reported a high prevalence of Panton Valentine leukocidin (PVL) producers among MSSA and MRSA and all patients presented a favourable outcome even though inappropriate antibiotic treatment was prescribed empirically. It is considered that the treatment of choice for these infections, irrespective of antibiotic susceptibility, is incision and drainage. The real pathogenic role of PVL it is still a subject of controversy, arising from results obtained from clinical studies

Macrolide resistance and molecular typing of Mycoplasma pneumoniae infections during a 4 year period in Spain

Mycoplasma pneumoniae (MP) causes community-acquired pneumonia affecting mainly children, and tends to produce cyclic outbreaks. The widespread use of macrolides is increasing resistance rates to these antibiotics. Molecular tools can help in diagnosis, typing and resistance detection, leading to better patient management. To assess the MP genotypes and resistance pattern circulating in our area while comparing serological and molecular diagnosis of MP. Molecular and serological diagnosis of MP was performed in 821 samples collected in Badalona (Barcelona, Spain) from 2013 to 2017. Multiple locus variable number tandem repeat analysis (MLVA) and macrolide resistance detection by pyrosequencing were performed in those cases positive by PCR. Presence of respiratory viruses and relevant clinical data were also recorded. MP was detected in 16.8% of cases by PCR, with an overall agreement with serology of 76%. Eleven different MLVA types were identified, with 4-5-7-2 (50.1%) and 3-5-6-2 (29.2%) being the most abundant, with the latter showing a seasonal increase during the study. A total of 8% of the strains harboured a point substitution associated with macrolide resistance, corresponding mainly to an A2063G 23S rRNA mutation and directly related to previous macrolide therapy. Analysis of respiratory viruses showed viral coinfections in most cases. Serological and molecular tools combined could improve MP diagnosis and the analysis of its infection patterns. Macrolide resistance is associated with previous therapy. Given that MP pneumonia usually resolves spontaneously, it should be reconsidered whether antibiotic treatment is suitable for all cases

Accessory gene regulator (Agr) functionality in <i>Staphylococcus aureus</i> derived from lower respiratory tract infections

<div><p>Objective</p><p>Characterization of <i>Staphylococcus aureus</i> clinical isolates derived from lower respiratory tract infections (LRTIs), and correlation between the functionality of the accessory gene regulator (Agr) and genotypic and phenotypic characteristics, clinical variables and clinical outcome.</p><p>Methods</p><p><i>S aureus</i> isolates derived from LRTIs and control groups (nasal carriage and bacteraemia) were genotyped using StaphyType DNA microarray. Agr activity was evaluated using the CAMP synergistic haemolysis assay and the Vesicle Lysis Test (VLT). Discordant strains were analysed by quantitative reverse-transcriptase real-time PCR (qRT-PCR).</p><p>Results</p><p>Agr was functional in 79.7% and 84.5% of strains according to the CAMP and VLT assays respectively. Higher concordance with RNAIII expression measured by qRT-PCR was observed with the VLT assay (76.2%) compared with the CAMP assay (23.8%). No statistically significant differences were observed in Agr functionality between the study groups, nor the phenotypical/genotypical bacterial characteristics. No association between increased mortality/respiratory complications and Agr function was observed.</p><p>Conclusions</p><p>Agr activity was high (82.2%) in isolates from LRTIs suggesting the importance of this global regulator in lower respiratory tract colonisation and infection. However, equally high Agr activity was observed in isolates derived from nasal carriage and bacteraemia, contradictory to previous observations. Agr functionality measured by the VLT assay was superior to CAMP assay.</p></div

Accessory gene regulator (Agr) functionality in Staphylococcus aureus derived from lower respiratory tract infections

Altres ajuts: This work has been funded by the project PI13/01418 which is part of "Plan Nacional de I+D+I" and co-funded by ISCIII-Subdirección General de Evaluacioón and "Fondo Europeo de Desarrollo Regional"(FEDER).D. Domínguez-Villanueva is funded by "Plan Nacional de I+D+I" and co-funded by ISCIII-Subdirección General de Evaluación and "Fondo Europeo de Desarrollo Regional"(FEDER). M. Gomes-Fernandes is funded by CAPES Foundation, Ministry of Education of Brazil (Brasılia, Brazil). Maisem Laabei was supported by a joint ERS/SEPAR fellowship (LTRF2015).This work also received a grant from the Spanish Society of Pneumology and Thoracic Surgery (SEPAR054/2011).Objective. Characterization of Staphylococcus aureus clinical isolates derived from lower respiratory tract infections (LRTIs), and correlation between the functionality of the accessory gene regulator (Agr) and genotypic and phenotypic characteristics, clinical variables and clinical outcome. Methods. S aureus isolates derived from LRTIs and control groups (nasal carriage and bacteraemia) were genotyped using StaphyType DNA microarray. Agr activity was evaluated using the CAMP synergistic haemolysis assay and the Vesicle Lysis Test (VLT). Discordant strains were analysed by quantitative reverse- transcriptase real-time PCR (qRT-PCR). Results. Agr was functional in 79.7% and 84.5% of strains according to the CAMP and VLT assays respectively. Higher concordance with RNAIII expression measured by qRT-PCR was observed with the VLT assay (76.2%) compared with the CAMP assay (23.8%). No statistically significant differences were observed in Agr functionality between the study groups, nor the phenotypical/genotypical bacterial characteristics. No association between increased mortality/respiratory complications and Agr function was observed. Conclusions. Agr activity was high (82.2%) in isolates from LRTIs suggesting the importance of this global regulator in lower respiratory tract colonisation and infection. However, equally high Agr activity was observed in isolates derived from nasal carriage and bacteraemia, contradictory to previous observations. Agr functionality measured by the VLT assay was superior to CAMP assay

Correlation between Agr functionality determined through CAMP, VLT and qRT-PCR and genotypic characteristics, defined as presence (Pos) or absence (Neg) of specific virulence factor gene probes.

<p>Correlation between Agr functionality determined through CAMP, VLT and qRT-PCR and genotypic characteristics, defined as presence (Pos) or absence (Neg) of specific virulence factor gene probes.</p