Study of CD27, CD38, HLA-DR and Ki-67 immune profiles for the characterization of active tuberculosis, latent infection and end of treatment.

Background: Current blood-based diagnostic tools for TB are insufficient to properly characterize the distinct stages of TB, from the latent infection (LTBI) to its active form (aTB); nor can they assess treatment efficacy. Several immune cell biomarkers have been proposed as potential candidates for the development of improved diagnostic tools. Objective: To compare the capacity of CD27, HLA-DR, CD38 and Ki-67 markers to characterize LTBI, active TB and patients who ended treatment and resolved TB. Methods: Blood was collected from 45 patients defined according to clinical and microbiological criteria as: LTBI, aTB with less than 1 month of treatment and aTB after completing treatment. Peripheral blood mononuclear cells were stimulated with ESAT-6/CFP-10 or PPD antigens and acquired for flow cytometry after labelling with conjugated antibodies against CD3, CD4, CD8, CD27, IFN-γ, TNF-α, CD38, HLA-DR, and Ki-67. Conventional and multiparametric analyses were done with FlowJo and OMIQ, respectively. Results: The expression of CD27, CD38, HLA-DR and Ki-67 markers was analyzed in CD4+ T-cells producing IFN-γ and/or TNF-α cytokines after ESAT-6/CFP-10 or PPD stimulation. Within antigen-responsive CD4+ T-cells, CD27- and CD38+ (ESAT-6/CFP-10-specific), and HLA-DR+ and Ki-67+ (PPD- and ESAT-6/CFP-10-specific) populations were significantly increased in aTB compared to LTBI. Ki-67 demonstrated the best discriminative performance as evaluated by ROC analyses (AUC > 0.9 after PPD stimulation). Data also points to a significant change in the expression of CD38 (ESAT-6/CFP-10-specific) and Ki-67 (PPD- and ESAT-6/CFP-10-specific) after ending the anti-TB treatment regimen. Furthermore, ratio based on the CD27 median fluorescence intensity in CD4+ T-cells over Mtb-specific CD4+ T-cells showed a positive association with aTB over LTBI (ESAT-6/CFP-10-specific). Additionally, multiparametric FlowSOM analyses revealed an increase in CD27 cell clusters and a decrease in HLA-DR cell clusters within Mtb-specific populations after the end of treatment. Conclusion: Our study independently confirms that CD27-, CD38+, HLA-DR+ and Ki-67+ populations on Mtb-specific CD4+ T-cells are increased during active TB disease. Multiparametric analyses unbiasedly identify clusters based on CD27 or HLA-DR whose abundance can be related to treatment efficacy. Further studies are necessary to pinpoint the convergence between conventional and multiparametric approaches.

Direct Quantitative Immunochemical Analysis of Autoinducer Peptide IV for Diagnosing and Stratifying Staphylococcus aureus Infections.

An immunochemical strategy to detect and quantify AIP-IV, the quorum sensing (QS) signaling molecule produced by Staphylococcus aureusagr type IV, is reported here for the first time. Theoretical calculations and molecular modeling studies have assisted on the design and synthesis of a suitable peptide hapten (AIPIVS), allowing to obtain high avidity and specific antibodies toward this peptide despite its low molecular weight. The ELISA developed achieves an IC50 value of 2.80 ± 0.17 and an LOD of 0.19 ± 0.06 nM in complex media such as 1/2 Tryptic Soy Broth. Recognition of other S. aureus AIPs (I-III) is negligible (cross-reactivity below 0.001%), regardless of the structural similarities. A pilot study with a set of clinical isolates from patients with airways infection or colonization demonstrates the potential of this ELISA to perform biomedical investigations related to the role of QS in pathogenesis and the association between dysfunctional agr or the agr type with unfavorable clinical outcomes. The AIP-IV levels could be quantified in the low nanomolar range in less than 1 h after inoculating agr IV-genotyped isolates in the culture broth, while those genotyped as I-III did not show any immunoreactivity after a 48 h growth, pointing to the possibility to use this technology for phenotyping S. aureus. The research strategy here reported can be extended to the rest of the AIP types of S. aureus, allowing the development of powerful multiplexed chips or point-of-care (PoC) diagnostic devices to unequivocally identify its presence and its agr type on samples from infected patients.

Genotypic and Phenotypic Characterization of Staphylococcus aureus Isolates from the Respiratory Tract in Mechanically-Ventilated Patients.

Staphylococcus aureus is a commensal and frequent colonizer of the upper respiratory tract. When mechanical ventilation disrupts natural defenses, S. aureus 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 S. aureus 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 (p = 0.002). Hla was detected in 71 strains (75.5%), with most of the producer strains in pneumonia and bronchial colonization group (p = 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 S. aureus 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.

Dysfunctional accessory gene regulator (agr) as a prognostic factor in invasive Staphylococcus aureus infection: a systematic review and meta-analysis.

The accessory gene regulator (agr) locus of Staphylococcus aureus is a quorum-sensing virulence regulator. Although there are many studies concerning the effect of dysfunctional agr on the outcomes of S. aureus infection, there is no systematic review to date. We systematically searched for clinical studies reporting outcomes of invasive S. aureus infections and the proportion of dysfunctional agr among their causative strains, and we performed a meta-analysis to obtain estimates of the odds of outcomes of invasive S. aureus infection with dysfunctional versus functional agr. Of 289 articles identified by our research strategy, 20 studies were meta-analysed for crude analysis of the impact of dysfunctional agr on outcomes of invasive S. aureus infection. Dysfunctional agr was generally associated with unfavourable outcomes (OR 1.32, 95% CI 1.05-1.66), and the impact of dysfunctional agr on outcome was more prominent in invasive methicillin-resistant S. aureus (MRSA) infections (OR 1.54, CI 1.20-1.97). Nine studies were meta-analysed for the impact of dysfunctional agr on the 30-day mortality of invasive S. aureus infection. Invasive MRSA infection with dysfunctional agr exhibited higher 30-day mortality (OR 1.40, CI 1.03-1.90) than that with functional agr. On the other hand, invasive MSSA infection with dysfunctional agr exhibited lower 30-day mortality (OR 0.51, CI 0.27-0.95). In the post hoc subgroup analysis by the site of MRSA infection, dysfunctional agr was associated with higher 30-day mortality in MRSA pneumonia (OR 2.48, CI 1.17-5.25). The effect of dysfunctional agr on the outcome of invasive S. aureus infection may vary depending on various conditions, such as oxacillin susceptibility and the site of infection. Dysfunctional agr was generally associated with unfavourable clinical outcomes and its effect was prominent in MRSA and pneumonia. Dysfunctional agr may be applicable for outcome prediction in cases of invasive MRSA infection with hardly eradicable foci such as pneumonia.

Efecto de la vacunación en la prevención de gripe grave en adultos atendidos en un hospital de tercer nivel durante la temporada 2017-2018 / Effect of vaccination on the prevention of influenza-related severe illness in adults attended in a third level hospital during the 2017-2018 epidemic season

OBJETIVOS: El objetivo de este trabajo fue evaluar el efecto de la vacunación antigripal en la prevención de casos graves asociados a gripe en pacientes adultos atendidos en un hospital de tercer nivel durante la temporada epidémica 2017-2018. METODOLOGÍA: Se realizó un análisis descriptivo con toda la población de sujetos con gripe confirmada por laboratorio en la temporada 2017-2018. Se definió caso grave como el ingreso en unidades de críticos o muerte. El efecto de la vacuna en la población adulta se determinó mediante análisis de regresión logística multivariante. RESULTADOS: Entre las semanas epidemiológicas 44/2017 y 19/2018, el laboratorio del hospital detectó 706 muestras positivas de virus influenza. De los 551 pacientes confirmados de 18 años o más, cuarenta y tres fueron ingresados en alguna de las unidades de críticos del hospital y 26 fallecieron durante el ingreso. El modelo multivariante explicativo mostró la vacunación de la gripe durante la temporada de estudio como factor protector del desarrollo de gravedad [OR: 0,27 (0,11-0,65), p = 0,004], ajustada por edad [1,03 (1,01-1,06), p = 0,04], sexo, tipo de virus (H1N1-pdm09, H3N2 o B) y el estar clasificado como Paciente Crónico Complejo o Enfermedad Avanzada Crónica. CONCLUSIONES: La vacuna de la gripe se muestra como factor protector frente al desarrollo de complicaciones en una temporada en la que la vacuna no contiene el virus que más ha circulado entre la población. Se debe recomendar la vacuna antigripal con periodicidad anual a los grupos de riesgo establecidos por las autoridades sanitarias

OBJECTIVES: The objective of this research was to evaluate the effect of influenza vaccination on the prevention of influenza-related severe cases in adults treated in a third-level hospital during the 2017-2018 epidemic season. METHODOLOGY: A descriptive analysis was performed on the entire population of subjects with a laboratory-confirmed influenza test during the 2017-2018 season. A severe case was defined as a patient treated in one of the Intensive Care Units (ICUs) and/or death. The effect of the vaccine on the adult population was determined by multivariate logistic regression analysis. RESULTS: Between epidemiological weeks 44/2017 and 19/2018, the hospital's laboratory detected 706 positive samples for influenza virus. Of the 551 confirmed patients aged 18 years or older, forty-three were admitted to one of the ICUs, and 26 died during admission. The explanatory multivariate model has shown that flu vaccination prior to or during the epidemic season was a protective factor for the development of severity [OR:0.27 (0.11-0.65, p = 0.004)], adjusted by age [OR: 1.03 (1.01-1.06), p=.04], sex, type of virus (H1N1-pdm09, H3N2 or B virus), Chronic Complex Patient index or Advanced Chronic Disease index. Conclussions: Influenza vaccination is a protective factor against the development of severity associated with influenza infection in a season when vaccination did not contain the virus with higher epidemic circulation among the population. Flu vaccination should be recommended annually following the guidelines established by the health authorities

Diagnostic Accuracy of Interferon Gamma-Induced Protein 10 mRNA Release Assay for Tuberculosis.

Interferon gamma (IFN-γ) release assays (IGRAs) are increasingly used to test for latent tuberculosis (TB) infection. Although highly specific, IGRAs have a relatively high false-negative rate in active TB patients. A more sensitive assay is needed. IFN-γ-induced protein 10 (IP-10) is an alternative biomarker with a 100-fold-higher expression level than IFN-γ, allowing for different analysis platforms, including molecular detection. The PCR technique is already an integrated tool in most TB laboratories and, thus, an obvious platform to turn to. In this case-control study, we investigated the diagnostic sensitivity and specificity of a molecular assay detecting IP-10 mRNA expression following antigen stimulation of a blood sample. We included 89 TB patients and 99 healthy controls. Blood was drawn in QuantiFeron-TB gold in-tube (QFT) assay tubes. Eight hours poststimulation, IP-10 mRNA expression was analyzed, and 20 h poststimulation, IP-10 and IFN-γ protein plasma levels were analyzed using an in-house IP-10 enzyme-linked immunosorbent assay (ELISA) and the official QFT ELISA, respectively. The IP-10 mRNA assay provided high specificity (98%), sensitivity (80%), and area under the concentration-time curve (AUC) (0.97); however, the QFT assay provided a higher overall diagnostic potential, with specificity of 100%, sensitivity of 90%, and AUC of 0.99. The IP-10 protein assay performed on par with the QFT assay, with specificity of 98%, sensitivity of 87%, and AUC of 0.98. We have provided proof of high technical performance of a molecular assay detecting IP-10 mRNA expression. As a diagnostic tool, this assay would gain from further optimization, especially on the kinetics of IP-10 mRNA expression.

Effect of vaccination on the prevention of influenza-related severe illness in adults attended in a third level hospital during the 2017-2018 epidemic season. / Efecto de la vacunación en la prevención de gripe grave en adultos atendidos en un hospital de tercer nivel durante la temporada 2017-2018.

OBJECTIVES: The objective of this research was to evaluate the effect of influenza vaccination on the prevention of influenza-related severe cases in adults treated in a third-level hospital during the 2017-2018 epidemic season. METHODOLOGY: A descriptive analysis was performed on the entire population of subjects with a laboratory-confirmed influenza test during the 2017-2018 season. A severe case was defined as a patient treated in one of the Intensive Care Units (ICUs) and/or death. The effect of the vaccine on the adult population was determined by multivariate logistic regression analysis. RESULTS: Between epidemiological weeks 44/2017 and 19/2018, the hospital's laboratory detected 706 positive samples for influenza virus. Of the 551 confirmed patients aged 18 years or older, forty-three were admitted to one of the ICUs, and 26 died during admission. The explanatory multivariate model has shown that flu vaccination prior to or during the epidemic season was a protective factor for the development of severity [OR:0.27 (0.11-0.65, p=0.004)], adjusted by age [OR: 1.03 (1.01-1.06), p=.04], sex, type of virus (H1N1-pdm09, H3N2 or B virus), Chronic Complex Patient index or Advanced Chronic Disease index. CONCLUSSIONS: Influenza vaccination is a protective factor against the development of severity associated with influenza infection in a season when vaccination did not contain the virus with higher epidemic circulation among the population. Flu vaccination should be recommended annually following the guidelines established by the health authorities.

Development and Evaluation of a Microarray Platform for Detection of Serum Antibodies Against Streptococcus pneumoniae Capsular Polysaccharides.

Streptococcus pneumoniae is responsible for severe infections, causing millions of deaths yearly. Immunoglobulin G (IgG) antibodies against the capsular polysaccharide (CPS) offer S. pneumoniae serotype-specific protection. In this work, we examined the applicability of the microarray technology to detect CPS type-specific IgGs in serum, using a collection of 22 microarray-printed S. pneumoniae CPSs. First, printing of five CPSs onto nitrocellulose-coated glass slides was tested. Successful printing was only achieved for certain CPS types and concentrations. This behavior was tentatively related with diverse viscosities of the CPS solutions. Measurement of dynamic viscosities fully supported this assumption and helped to establish suitable CPS type- and concentration-dependent printing conditions. Next, the potential of CPS microarrays for detecting recognition by anti-CPS IgGs was examined using well-defined rabbit pneumococcal antisera. In all cases, the expected antiserum-CPS binding signals were detected, prompting a proof-of-concept analysis of human serum samples. Clearly distinct serum- and CPS-specific binding patterns and intensities were observed, evidencing selective detection of CPS type-specific IgGs. Compared to the ELISA assay commonly used to quantitate CPS type-specific IgGs in serum, the newly developed S. pneumoniae CPS microarrays offer the advantage of enabling the simultaneous analysis of multiple CPS-serum interactions using minute CPS amounts and significantly reduced serum volumes. Therefore, the approach could be particularly valuable for gauging the presence of CPS type-specific IgGs in human serum when sample volumes are limited and/or numerous serum samples are being examined.

Novel intracellular antibiotic delivery system against Staphylococcus aureus: cloxacillin-loaded poly(d,l-lactide-co-glycolide) acid nanoparticles.

Aim: First, to compare in vitro minimum inhibitory concentrations (MIC) of free cloxacillin and cloxacillin-containing nanoparticles (NP) against methicillin-susceptible (MSSA) and resistant Staphylococcus aureus (MRSA) and second, to assess NP antimicrobial activity against intracellular S. aureus. Methods: Poly(d,l-lactide-co-glycolide) acid (PLGA)-NP were loaded with cloxacillin and physico-chemically characterized. MICs were determined for reference strains Newman-(MSSA) and USA300-(MRSA). Murine alveolar macrophages were infected, and bacterial intracellular survival was assessed after incubating with free-cloxacillin or PLGA-cloxacillin-NP. Results & conclusion: For both isolates, MICs for antibiotic-loaded-NP were lower than those obtained with free cloxacillin, indicating that the drug encapsulation improves antimicrobial activity. A sustained antibiotic release was demonstrated when using the PLGA-cloxacillin-NP. When considering the lowest concentrations, the use of drug-loaded NP enabled a higher reduction of intracellular bacterial load.

Cell-Mediated Immune Responses to in vivo-Expressed and Stage-Specific Mycobacterium tuberculosis Antigens in Latent and Active Tuberculosis Across Different Age Groups.

A quarter of the global human population is estimated to be latently infected by Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). TB remains the global leading cause of death by a single pathogen and ranks among the top-10 causes of overall global mortality. Current immunodiagnostic tests cannot discriminate between latent, active and past TB, nor predict progression of latent infection to active disease. The only registered TB vaccine, Bacillus Calmette-Guérin (BCG), does not adequately prevent pulmonary TB in adolescents and adults, thus permitting continued TB-transmission. Several Mtb proteins, mostly discovered through IFN-γ centered approaches, have been proposed as targets for new TB-diagnostic tests or -vaccines. Recently, however, we identified novel Mtb antigens capable of eliciting multiple cytokines, including antigens that did not induce IFN-γ but several other cytokines. These antigens had been selected based on high Mtb gene-expression in the lung in vivo, and have been termed in vivo expressed (IVE-TB) antigens. Here, we extend and validate our previous findings in an independent Southern European cohort, consisting of adults and adolescents with either LTBI or TB. Our results confirm that responses to IVE-TB antigens, and also DosR-regulon and Rpf stage-specific Mtb antigens are marked by multiple cytokines, including strong responses, such as for TNF-α, in the absence of detectable IFN-γ production. Except for TNF-α, the magnitude of those responses were significantly higher in LTBI subjects. Additional unbiased analyses of high dimensional flow-cytometry data revealed that TNF-α+ cells responding to Mtb antigens comprised 17 highly heterogeneous cell types. Among these 17 TNF-α+ cells clusters identified, those with CD8+TEMRA or CD8+CD4+ phenotypes, defined by the expression of multiple intracellular markers, were the most prominent in adult LTBI, while CD14+ TNF-α+ myeloid-like clusters were mostly abundant in adolescent LTBI. Our findings, although limited to a small cohort, stress the importance of assessing broader immune responses than IFN-γ alone in Mtb antigen discovery as well as the importance of screening individuals of different age groups. In addition, our results provide proof of concept showing how unbiased multidimensional multiparametric cell subset analysis can identify unanticipated blood cell subsets that could play a role in the immune response against Mtb.

Multidrug- and Extensively Drug-Resistant Mycobacterium tuberculosis Beijing Clades, Ukraine, 2015.

Multidrug-resistant (MDR) and extensively drug-resistant (XDR) tuberculosis (TB) is an emerging threat to TB control in Ukraine, a country with the third highest XDR TB burden globally. We used whole-genome sequencing of a convenience sample to identify bacterial genetic and patient-related factors associated with MDR/XDR TB in this country. MDR/XDR TB was associated with 3 distinct Mycobacterium tuberculosis complex lineage 2 (Beijing) clades, Europe/Russia W148 outbreak, Central Asia outbreak, and Ukraine outbreak, which comprised 68.9% of all MDR/XDR TB strains from southern Ukraine. MDR/XDR TB was also associated with previous treatment for TB and urban residence. The circulation of Beijing outbreak strains harboring broad drug resistance, coupled with constraints in drug supply and limited availability of phenotypic drug susceptibility testing, needs to be considered when new TB management strategies are implemented in Ukraine.

A multicentre analysis of Nocardia pneumonia in Spain: 2010-2016.

OBJECTIVE: To analyse all cases of Nocardia pneumonia occurring between 2010 and 2016 in five Spanish hospitals. METHODS: This was a retrospective observational analysis of clinical and microbiological data collected from 55 cases of Nocardia pneumonia. RESULTS: There were one to 20 cases per hospital and six to nine cases per year. Chronic obstructive pulmonary disease, bronchiectasis, and asthma were the main predisposing underlying respiratory conditions. Thirty-four patients were receiving systemic and/or inhaled corticosteroids prior to infection, eight had neoplasia, and six had haematological malignancies. Clinical and radiological findings were common to pneumonia of other infectious aetiologies, except for the frequent presence of nodules and cavitation. Overall, the 1-year mortality was high (38.2%), and mortality was directly related to the pulmonary disease in 15 patients (27.3%). The most frequently identified species were N. cyriacigeorgica (n=21), N. abscessus (n=8), and N. farcinica (n=5). All Nocardia isolates were susceptible to linezolid and all but two were susceptible to amikacin and trimethoprim-sulfamethoxazole. CONCLUSIONS: Nocardia pneumonia-associated mortality remains high, probably because of the debilitated status of patients in whom this pathogen is able to cause pulmonary infection.

International prevalence and risk factors evaluation for drug-resistant Streptococcus pneumoniae pneumonia.

OBJECTIVE: Streptococcus pneumoniae is the most frequent bacterial pathogen isolated in subjects with Community-acquired pneumonia (CAP) worldwide. Limited data are available regarding the current global burden and risk factors associated with drug-resistant Streptococcus pneumoniae (DRSP) in CAP subjects. We assessed the multinational prevalence and risk factors for DRSP-CAP in a multinational point-prevalence study. DESIGN: The prevalence of DRSP-CAP was assessed by identification of DRSP in blood or respiratory samples among adults hospitalized with CAP in 54 countries. Prevalence and risk factors were compared among subjects that had microbiological testing and antibiotic susceptibility data. Multivariate logistic regressions were used to identify risk factors independently associated with DRSP-CAP. RESULTS: 3,193 subjects were included in the study. The global prevalence of DRSP-CAP was 1.3% and continental prevalence rates were 7.0% in Africa, 1.2% in Asia, and 1.0% in South America, Europe, and North America, respectively. Macrolide resistance was most frequently identified in subjects with DRSP-CAP (0.6%) followed by penicillin resistance (0.5%). Subjects in Africa were more likely to have DRSP-CAP (OR: 7.6; 95%CI: 3.34-15.35, p<0.001) when compared to centres representing other continents. CONCLUSIONS: This multinational point-prevalence study found a low global prevalence of DRSP-CAP that may impact guideline development and antimicrobial policies.

Cigarette smoke exposure redirects Staphylococcus aureus to a virulence profile associated with persistent infection.

Tobacco smoking represents the leading preventable cause of death worldwide. Smoking is a recognised risk factor for several pathologies and is detrimental to host immune surveillance and defence. However, the impact of smoking on microbial residents of the nasopharyngeal cavity, in contact with cigarette smoke (CS), is lacking. Staphylococcus aureus is a major human pathogen that colonises the human nasopharynx and causes a wide range of infections. We investigated the impact of CS on specific virulence phenotypes important in S aureus pathogenesis. We observed strain-dependent differences following exposure to CS, namely growth inhibition, augmented biofilm formation, increased invasion of, and persistence within, bronchial alveolar epithelial cells. Additionally, we confirm the critical role of a functional accessory gene regulator (Agr) system in mediating increased biofilm development and host cell invasion and persistence following CS exposure. Furthermore, CS exposure resulted in reduced toxin production. Importantly, exposure of S aureus to CS accelerated the frequency of mutations and resulted in a significant increase in gentamicin-resistant small colony variant (SCV) formation. Mutational analysis revealed that CS induced SCVs emerge via the SOS response DNA mutagenic repair system. Taken together, our results suggest that CS redirects certain S aureus strains to a virulence profile associated with persistence.

Impact of Host Genetics and Biological Response Modifiers on Respiratory Tract Infections.

Host susceptibility to respiratory tract infections (RTI) is dependent on both genetic and acquired risk factors. Repeated bacterial and viral RTI, such as pneumonia from encapsulated microorganisms, respiratory tract infections related to respiratory syncytial virus or influenza, and even the development of bronchiectasis and asthma, are often reported as the first symptom of primary immunodeficiencies. In the same way, neutropenia is a well-known risk factor for invasive aspergillosis, as well as lymphopenia for Pneumocystis, and mycobacterial infections. However, in the last decades a better knowledge of immune signaling networks and the introduction of next generation sequencing have increased the number and diversity of known inborn errors of immunity. On the other hand, the use of monoclonal antibodies targeting cytokines, such as tumor necrosis factor alpha has revealed new risk groups for infections, such as tuberculosis. The use of biological response modifiers has spread to almost all medical specialties, including inflammatory diseases and neoplasia, and are being used to target different signaling networks that may mirror some of the known immune deficiencies. From a clinical perspective, the individual contribution of genetics, and/or targeted treatments, to immune dysregulation is difficult to assess. The aim of this article is to review the known and newly described mechanisms of impaired immune signaling that predispose to RTI, including new insights into host genetics and the impact of biological response modifiers, and to summarize clinical recommendations regarding vaccines and prophylactic treatments in order to prevent infections.

Persistent Isolation of Staphylococcus aureus in Mechanically-ventilated Patients: Impact of Host-Pathogen Factors on Outcome / Aislamiento persistente de Staphylococcus aureus en pacientes sometidos a ventilación mecánica: impacto de factores del huésped y del patógeno en la evolución clínica

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Matryoshka-type gastro-resistant microparticles for the oral treatment of Mycobacterium tuberculosis.

AIM: Production of Matryoshka-type gastroresistant microparticles containing antibiotic-loaded poly lactic-co-glycolic acid (PLGA) nanoparticles (NP) against Mycobacterium tuberculosis. MATERIALS & METHODS: The emulsification and evaporation methods were followed for the synthesis of PLGA-NPs and methacrylic acid-ethyl acrylate-based coatings to protect rifampicin from degradation under simulated gastric conditions. RESULTS & CONCLUSION: The inner antibiotic-loaded NPs here reported can be released under simulated intestinal conditions whereas their coating protects them from degradation under simulated gastric conditions. The encapsulation does not hinder the antituberculosis action of the encapsulated antibiotic rifampicin. A sustained antibiotic release could be obtained when using the drug-loaded encapsulated NPs. Compared with the administration of the free drug, a more effective elimination of M. tuberculosis was observed when applying the NPs against infected macrophages. The antibiotic-loaded PLGA-NPs were also able to cross an in vitro model of intestinal barrier.

Microbiological Progress in Patients with Bronchial Infection with Pseudomonas aeruginosa Treated with Nebulised Colistin.

BACKGROUND: Inhaled antibiotics allow the delivery of higher drug concentrations at the site of infection without the systemic adverse effects observed with the use of parenteral or oral antibiotics. These antibiotics have shown to decrease the number of exacerbations, reduce bacterial load or improve pulmonary function in several chronic respiratory conditions. OBJECTIVES: The aim of this study was to describe changes in the bacteriology of sputum in patients with chronic bronchial infection with Pseudomonas aeruginosa treated with nebulised colistin. MATERIAL AND METHODS: All patients with chronical infection with P. aeruginosa treated with nebulised colistin attending a day care unit during a 5-year (January 2010 to December 2014) period were included. Repeated-measures t tests were used to assess whether the introduction of colistin was associated with changes in the number of exacerbations or the length of the hospitalisations. RESULTS: Treatment with colistin was associated with a decrease in the number of ambulatory exacerbations (1.87-1.1, p = 0.007), of hospital exacerbations (1.3-0.7, p = 0.010) and of length of stay (15.7-8.6 days, p = 0.005). There was no linear trend in the proportion of isolate Enterobacteriaceae, gram-positive cocci, Haemophilus influenzae or fungi. Isolation of Enterobacteriaceae within 1 year after the beginning of the treatment with nebulised colistin was associated with an increase in the number of ambulatory exacerbations (incidence rate ratio 1.99, 95% CI 1.05-3.79). CONCLUSIONS: Nebulised colistin was effective in the treatment of chronic infection with P. aeruginosa, and no significant changes in the microbiological evolution were observed. Isolation of Enterobacteriaceae within 1 year after the beginning of the treatment with nebulised colistin was associated with an increase in the number of exacerbations.

Microbiological testing of adults hospitalised with community-acquired pneumonia: an international study.

This study aimed to describe real-life microbiological testing of adults hospitalised with community-acquired pneumonia (CAP) and to assess concordance with the 2007 Infectious Diseases Society of America (IDSA)/American Thoracic Society (ATS) and 2011 European Respiratory Society (ERS) CAP guidelines. This was a cohort study based on the Global Initiative for Methicillin-resistant Staphylococcus aureus Pneumonia (GLIMP) database, which contains point-prevalence data on adults hospitalised with CAP across 54 countries during 2015. In total, 3702 patients were included. Testing was performed in 3217 patients, and included blood culture (71.1%), sputum culture (61.8%), Legionella urinary antigen test (30.1%), pneumococcal urinary antigen test (30.0%), viral testing (14.9%), acute-phase serology (8.8%), bronchoalveolar lavage culture (8.4%) and pleural fluid culture (3.2%). A pathogen was detected in 1173 (36.5%) patients. Testing attitudes varied significantly according to geography and disease severity. Testing was concordant with IDSA/ATS and ERS guidelines in 16.7% and 23.9% of patients, respectively. IDSA/ATS concordance was higher in Europe than in North America (21.5% versus 9.8%; p<0.01), while ERS concordance was higher in North America than in Europe (33.5% versus 19.5%; p<0.01). Testing practices of adults hospitalised with CAP varied significantly by geography and disease severity. There was a wide discordance between real-life testing practices and IDSA/ATS/ERS guideline recommendations.