Salivary polyreactive antibodies and Haemophilus influenzae are associated with respiratory infection severity in young children with recurrent respiratory infections.

BACKGROUND: Recurrent respiratory tract infections (rRTIs) are a common reason for immunodiagnostic testing in children, which relies on serum antibody level measurements. However, because RTIs predominantly affect the respiratory mucosa, serum antibodies may inaccurately reflect local immune defences. We investigated antibody responses in saliva and their interplay with the respiratory microbiota in relation to RTI severity and burden in young children with rRTIs. METHODS: We conducted a prospective cohort study including 100 children aged <10 years with rRTIs, their family members and healthy healthcare professionals. Total and polyreactive antibody concentrations were determined in serum and saliva (ELISA); respiratory microbiota composition (16S rRNA sequencing) and respiratory viruses (quantitative PCR) were characterised in nasopharyngeal swabs. Proteomic analysis (Olink) was performed on saliva and serum samples. RTI symptoms were monitored with a daily mobile phone application and assessed using latent class analysis and negative binomial mixed models. RESULTS: Serum antibody levels were not associated with RTI severity. Strikingly, 28% of salivary antibodies and only 2% of serum antibodies displayed polyreactivity (p<0.001). Salivary polyreactive IgA was negatively associated with recurrent lower RTIs (adjusted OR 0.80, 95% CI 0.67-0.94) and detection of multiple respiratory viruses (adjusted OR 0.76, 95% CI 0.61-0.96). Haemophilus influenzae abundance was positively associated with RTI symptom burden (regression coefficient 0.05, 95% CI 0.02-0.08). CONCLUSION: These results highlight the importance of mucosal immunity in RTI severity and burden, and suggest that the level of salivary polyreactive IgA and H. influenzae abundance may serve as indicators of infection severity and burden in young children with rRTIs.

PEGylation of Tobramycin Improves Mucus Penetration and Antimicrobial Activity against Pseudomonas aeruginosa Biofilms in Vitro.

Pseudomonas aeruginosa is the predominant pathogen in the persistent lung infections of cystic fibrosis (CF) patients among other diseases. One of the mechanisms of resistance of P. aeruginosa infections is the formation and presence of biofilms. Previously, we demonstrated that PEGylated-tobramycin (Tob-PEG) had superior antimicrobial activity against P. aeruginosa biofilms compared to tobramycin (Tob). The goal of this study was to optimize the method of PEGylation of Tob and assess its activity in an in vitro CF-like mucus barrier biofilm model. Tob was PEGylated using three separate chemical conjugation methods and analyzed by 1H NMR. A comparison of the Tob-PEG products from the different conjugation methods showed significant differences in the reduction of biofilm proliferation after 24 h of treatment. In the CF-like mucus barrier model, Tob-PEG was significantly better than Tob in reducing P. aeruginosa proliferation after only 5 h of treatment ( p < 0.01). Finally, Tob-PEG caused a reduction in the number of surviving P. aeruginosa biofilm colonies higher than that of Tob ( p < 0.0001). We demonstrate the significantly improved antimicrobial activity of Tob-PEG against P. aeruginosa biofilms compared to Tob using two PEGylation methods. Tob-PEG had better in vitro activity compared to that of Tob against P. aeruginosa biofilms growing in a CF-like mucus barrier model.

Antibiotic Prescribing for Nonbacterial Acute Upper Respiratory Infections in Elderly Persons.

BACKGROUND: Reducing inappropriate antibiotic prescribing for acute upper respiratory tract infections (AURIs) requires a better understanding of the factors associated with this practice. OBJECTIVE: To determine the prevalence of antibiotic prescribing for nonbacterial AURIs and whether prescribing rates varied by physician characteristics. DESIGN: Retrospective analysis of linked administrative health care data. SETTING: Primary care physician practices in Ontario, Canada (January-December 2012). PATIENTS: Patients aged 66 years or older with nonbacterial AURIs. Patients with cancer or immunosuppressive conditions and residents of long-term care homes were excluded. MEASUREMENTS: Antibiotic prescriptions for physician-diagnosed AURIs. A multivariable logistic regression model with generalized estimating equations was used to examine whether prescribing rates varied by physician characteristics, accounting for clustering of patients among physicians and adjusting for patient-level covariates. RESULTS: The cohort included 8990 primary care physicians and 185 014 patients who presented with a nonbacterial AURI, including the common cold (53.4%), acute bronchitis (31.3%), acute sinusitis (13.6%), or acute laryngitis (1.6%). Forty-six percent of patients received an antibiotic prescription; most prescriptions were for broad-spectrum agents (69.9% [95% CI, 69.6% to 70.2%]). Patients were more likely to receive prescriptions from mid- and late-career physicians than early-career physicians (rate difference, 5.1 percentage points [CI, 3.9 to 6.4 percentage points] and 4.6 percentage points [CI, 3.3 to 5.8 percentage points], respectively), from physicians trained outside of Canada or the United States (3.6 percentage points [CI, 2.5 to 4.6 percentage points]), and from physicians who saw 25 to 44 patients per day or 45 or more patients per day than those who saw fewer than 25 patients per day (3.1 percentage points [CI, 2.1 to 4.0 percentage points] and 4.1 percentage points [CI, 2.7 to 5.5 percentage points], respectively). LIMITATION: Physician rationale for prescribing was unknown. CONCLUSION: In this low-risk elderly cohort, 46% of patients with a nonbacterial AURI were prescribed antibiotics. Patients were more likely to receive prescriptions from mid- or late-career physicians with high patient volumes and from physicians who were trained outside of Canada or the United States. PRIMARY FUNDING SOURCE: Ontario Ministry of Health and Long-term Care, Academic Medical Organization of Southwestern Ontario, Schulich School of Medicine and Dentistry, Western University, and Lawson Health Research Institute.

Inhalable Levofloxacin Liposomes Complemented with Lysozyme for Treatment of Pulmonary Infection in Rats: Effective Antimicrobial and Antibiofilm Strategy.

Treatment of bacterial infections becomes increasingly complicated due to increasing bacterial resistance and difficulty in developing new antimicrobial agents. Emphasis should be laid on improvising the existing treatment modalities. We studied the improved antimicrobial and antibiofilm activity of levofloxacin (LFX) and lysozyme (LYS) in microbiological studies. LFX at sub-minimum inhibitory concentration with LYS eradicated > 85% of preformed biofilm. LFX was actively loaded into the liposomes using pH gradient method and was spray-dried with LYS solution. Percent entrapment of LFX in liposome was > 80% and prolonged cumulative release of 85% LFX at the end of 12 h. In vitro lung deposition study and solid-state characterization for spray dried LFX liposome in combination with LYS (LFX liposome-LYS) was performed. Co-spray dried product had mass median aerodynamic diameter ranging < 5 µm. In pharmacodynamic study, Staphylococcus aureus infected rats were treated with LFX liposome-LYS. Lungs, bronchoalveolar lavage fluid (BALF), and nasal fluid were evaluated for microbial burden. Expression of cytokine levels in BALF and serum were also studied by ELISA. In addition, mRNA expression for lung inflammatory mediators and lung myeloperoxidase activity were carried out. Further, lungs and histological changes were observed grossly. Untreated infected rat lungs demonstrated higher mRNA expression for inflammatory markers, cytokine levels, and microbial load compared to vehicle control. Conversely, LFX liposome-LYS significantly abated these adverse repercussions. Histology findings were also in agreement of above. Acute toxicity study revealed safeness of LFX liposome-LYS. Our findings confirm LFX liposome-LYS exhibited prolonged, improved antibiofilm and antimicrobial efficacy in treating S. aureus infection.

Microscopic Analysis and Quality Assessment of Induced Sputum From Children With Pneumonia in the PERCH Study.

BACKGROUND.: It is standard practice for laboratories to assess the cellular quality of expectorated sputum specimens to check that they originated from the lower respiratory tract. The presence of low numbers of squamous epithelial cells (SECs) and high numbers of polymorphonuclear (PMN) cells are regarded as indicative of a lower respiratory tract specimen. However, these quality ratings have never been evaluated for induced sputum specimens from children with suspected pneumonia. METHODS.: We evaluated induced sputum Gram stain smears and cultures from hospitalized children aged 1-59 months enrolled in a large study of community-acquired pneumonia. We hypothesized that a specimen representative of the lower respiratory tract will contain smaller quantities of oropharyngeal flora and be more likely to have a predominance of potential pathogens compared to a specimen containing mainly saliva. The prevalence of potential pathogens cultured from induced sputum specimens and quantity of oropharyngeal flora were compared for different quantities of SECs and PMNs. RESULTS.: Of 3772 induced sputum specimens, 2608 (69%) had <10 SECs per low-power field (LPF) and 2350 (62%) had >25 PMNs per LPF, measures traditionally associated with specimens from the lower respiratory tract in adults. Using isolation of low quantities of oropharyngeal flora and higher prevalence of potential pathogens as markers of higher quality, <10 SECs per LPF (but not >25 PMNs per LPF) was the microscopic variable most associated with high quality of induced sputum. CONCLUSIONS.: Quantity of SECs may be a useful quality measure of induced sputum from young children with pneumonia.

Nano into Micro Formulations of Tobramycin for the Treatment of Pseudomonas aeruginosa Infections in Cystic Fibrosis.

Here, nano into micro formulations (NiMs) of tobramycin for the treatment of Pseudomonas aeruginosa airway infections in cystic fibrosis (CF) are described. NiMs were produced by spray drying a solution containing polymers or sugars and a nanometric polyanion-tobramcyin complex (PTC), able to achieve a prolonged antibiotic release. NiMs properties were compared to TOBIPodhaler(Novartis), the only one commercially available dry powder inhalatory formulation based on porous microparticles. Produced NiMs showed adequate characteristics for pulmonary administration, as spherical shape, micrometric size, and high cytocompatibility toward human bronchial epithelial cells. Contrarily to TOBIPodhaler, some of produced NiMs, thanks to their specific chemical composition, are able to facilitate the drug diffusion through the mucus secretion, achieving, at the same time, a sustained tobramycin delivery. Moreover, NiMs showed pronounced antimicrobial activity against P. aeruginosa pathogens and their biofilm, if compared to free tobramycin and TOBIPodhaler, demonstrating the potential of obtained formulations as drug delivery systems for the treatment of pulmonary infections in CF patients.

Large Porous Particles for Sustained Release of a Decoy Oligonucelotide and Poly(ethylenimine): Potential for Combined Therapy of Chronic Pseudomonas aeruginosa Lung Infections.

We have recently demonstrated that the specific inhibition of nuclear factor-κB by a decoy oligonucleotide (dec-ODN) delivered through inhalable large porous particles (LPP) made of poly(lactic-co-glycolic acid) (PLGA) may be highly beneficial for long-term treatment of lung inflammation. Nevertheless, besides chronic inflammation, multifunctional systems aimed to control also infection are required in chronic lung diseases, such as cystic fibrosis (CF). In this work, we tested the hypothesis that engineering PLGA-based LPP with branched poly(ethylenimine) (PEI) may improve LPP properties for pulmonary delivery of dec-ODN, with particular regard to the treatment of Pseudomonas aeruginosa lung infections. After getting insight into the role of PEI on the technological properties of PLGA-based LPP for delivery of dec-ODN, the putative synergistic effect of PEI free or PEI released from LPP on in vitro antimicrobial activity of tobramycin (Tb) and aztreonam (AZT) against P. aeruginosa was elucidated. Meanwhile, cytotoxicity studies on A549 cells were carried out. Results clearly demonstrate that the dry powders have promising aerosolization properties and afford a prolonged in vitro release of both dec-ODN and PEI. The encapsulation of PEI into LPP results in a 2-fold reduction of the minimum inhibitory concentration of AZT, while reducing the cytotoxic effect of PEI. Of note, the developed ODN/PLGA/PEI LPP persisted at lung at least for 14 days after intratracheal administration in rats where they can provide sustained and combined release of dec-ODN and PEI. dec-ODN will likely act as an anti-inflammatory drug, while PEI may enhance the therapeutic activity of inhaled antibiotics, which are commonly employed for the treatment of concomitant lung infections.

Oral and endotracheal tubes colonization by periodontal bacteria: a case-control ICU study.

Periodontal infection is a possible risk factor for respiratory disorders; however, no studies have assessed the colonization of periodontal pathogens in endotracheal tubes (ET). This case-control study analyzed whether periodontal pathogens are able to colonize ET of dentate and edentulous patients in intensive care units (ICU) and whether oral and ET periodontal pathogen profiles have any correlation between these patients. We selected 18 dentate and 18 edentulous patients from 78 eligible ICU patients. Oral clinical examination including probing depth, clinical attachment level, gingival index , and plaque index was performed by a single examiner, followed by oral and ET sampling and processing by quantitative polymerase chain reaction (total bacterial load, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Tannerella forsythia). Data were statistically analyzed by Mann-Whitney U, two-way analysis of variance (p < 0.05). Among dentate, there was no correlation between clinical parameters and ET bacterial levels. Both dentate and edentulous patients showed similar ET bacterial levels. Dentate patients showed no correlation between oral and ET bacterial levels, while edentulous patients showed positive correlations between oral and ET levels of A. actinomycetemcomitans, P. gingivalis, and T. forsythia. Periodontal pathogens can colonize ET and the oral cavity of ICU patients. Periodontal pathogen profiles tend to be similar between dentate and edentulous ICU patients. In ICU patients, oral cavity represents a source of ET contamination. Although accompanied by higher oral bacterial levels, teeth do not seem to influence ET bacterial profiles.

The Prevalence of Human Bocavirus, Human Coronavirus-NL63, Human Metapneumovirus, Human Polyomavirus KI and WU in Respiratory Tract Infections in Kuwait.

OBJECTIVE: The aim of this study was to investigate the prevalence of human coronavirus (HCoV)-NL63, human metapneumovirus (hMPV), human bocavirus (Boca), human polyomavirus KI (KIV) and human polyomavirus WU (WUV) in respiratory tract infections (RTI) in Kuwait. MATERIALS AND METHODS: Respiratory samples from 735 hospitalized patients with RTI from September 2010 to April 2013 were evaluated for the presence of HCoV-NL63, hMPV, Boca, KIV and WUV using molecular assays, polymerase chain reaction (PCR) and reverse-transcription PCR. RESULTS: Of the 735 patients, 285 (38.8%) were diagnosed with viral RTI. The distribution of respiratory viruses was hMPV: 15 (5.3%), Boca: 14 (4.9%), WUV: 10 (3.5%) and KIV: 4 (1.4%). HCoV-NL63 was not detected in any of the samples. CONCLUSIONS: These newly discovered viruses were associated with the development of RTI in Kuwait. The rapid identification of these viral infections could aid in the control of nosocomial transmission, reduce the use of antibiotics and improve treatment and management strategies.

The impact of oral diseases on respiratory health and the influence of respiratory infections on the oral microbiome.

OBJECTIVE: The objective of this review is to examine the relationship between oral diseases and respiratory health, investigating how oral microbiome disruptions contribute to respiratory tract infections. Additionally, it aims to explore the impact of respiratory disease symptoms and treatments on the oral microbiome. DATA SOURCES: The literature utilized in this review was sourced from studies focusing on the correlation between oral health and respiratory infections, spanning a period of 40 years. Various databases and scholarly sources were likely consulted to gather relevant research articles, reviews, and clinical studies. STUDY SELECTION: This review summarizes four decades-long research, providing insights into the intricate relationship between oral and respiratory health. It delves into how oral diseases influence respiratory tract conditions and vice versa. The selection process likely involved identifying studies that addressed the interaction between oral microbiome disruptions and respiratory complications. CONCLUSION: Oral diseases or poor oral habits have been known to increase the risk of getting respiratory infections. Modern techniques have demonstrated the relationship between oral disease and respiratory tract infections like influenza, chronic obstructive pulmonary diseases, asthma, and Pneumonia. Apart from that, the medications used to treat respiratory diseases affect oral physiological factors like the pH of saliva, and saliva flow rate, which can cause significant changes in the oral microbiome. This review provides regular oral hygiene and care that can prevent respiratory health and respiratory infections. CLINICAL SIGNIFICANCE: Understanding the intricate relationship between oral health and respiratory infections is crucial for healthcare providers. Implementing preventive measures and promoting good oral hygiene habits can reduce respiratory tract infections and improve overall respiratory health outcomes.

Epidemiological characteristics of common respiratory pathogens in children.

Children's respiratory tract infection is a common disease affecting children's health, our purpose is to describe the epidemiological characteristics of common pathogens of children's respiratory tract infection in central Shandong, China, and compare them with those in other parts of world, so as to summarize the rules of children's respiratory tract infection in central Shandong, and provide scientific basis for health departments to prevent and treat local children's respiratory tract infection. Sputum, tracheal aspirate, alveolar lavage fluid and other samples of 4804 children admitted to wards of Zibo Maternal and Child Health Hospital for treatment of respiratory tract infection from June 2019 to December 2022 were collected, and 12 common respiratory tract pathogens were detected by PCR capillary electrophoresis fragment analysis, two bacteria (Streptococcus pneumoniae, Haemophilus influenzae), two atypical pathogens (Mycoplasma Pneumoniae, Chlamydia Pneumoniae) and eight viruses (Human rhinovirus, Respiratory Syncytial Virus, Influenza A Virus, Parainfluenza Virus, Human metapneumovirus, Human boca virus, Human coronavirus, Influenza B virus) were included, the positive detection rate of single pathogen, the proportion of each type of respiratory tract mixed infection and the positive detection rate of single pathogen in different ages and seasons were analyzed statistically. (1)Among 4804 children with respiratory tract infection, the total positive rate was 77.87% (3741/4804), the positive rate of single pathogen was 43.40% (1656/4804), Streptococcus pneumoniae, Rhinovirus and Respiratory syncytial virus were the highest, there were 2085 cases of mixed infection with two or more pathogens, the positive rate was 43.40%. (2) The positive rates of infection in infant group (0-1 years old), infant group (1-3 years old), preschool group and school age group (3 years old-) were roughly the same, the infection rates of Streptococcus pneumoniae, Respiratory syncytial virus and Parainfluenza virus in infant group, Rhinovirus in infant group, Influenza A virus, Chlamydia pneumoniae, Mycoplasma pneumoniae and Haemophilus influenzae in school age group were higher than those in other groups, the difference was statistically significant (P < 0.05). (3) The positive detection rates of spring, summer, autumn and winter groups were 43.58%, 38.64%, 33.73% and 29.27%, respectively, the positive rates of Streptococcus pneumoniae and Haemophilus influenzae in spring group, Mycoplasma pneumoniae in summer group, Rhinovirus, Respiratory syncytial virus and Influenza A virus in autumn group, Chlamydia pneumoniae, Boca virus and Influenza B virus in winter group were higher than those in other seasons, and the differences were statistically significant (P < 0.05). The pathogen detection rate of children varies with age and season, and the prevention and treatment of a certain respiratory pathogen infection must be combined with its raging season and age rule.

A comparison of the prevalence of respiratory pathogens and opportunistic respiratory pathogenic profile of 'clean' and 'unclean' removable dental prostheses.

OBJECTIVES: To determine and compare the opportunistic respiratory pathogenic index (ORPI) and prevalence of respiratory pathogens between clean and unclean removable prostheses. METHODS: A cross-sectional study was conducted among 97 removable prosthesis wearers at a teaching dental hospital. Participants' prosthesis hygiene was grouped into clean and unclean. After prosthesis plaque samples were sequenced using the Type IIB Restriction-site Associated DNA Sequencing for Microbiome method, the prevalence was assessed for the presence of respiratory pathogens on each sample. The ORPIs for clean and unclean prostheses were quantified based on the sum of the relative abundance of respiratory pathogenic bacteria in a microbiome using a reference database that contains opportunistic respiratory pathogens and disease-associated information. RESULTS: A total of 30 opportunistic respiratory pathogens were identified on the removable prostheses. Eighty-one (83.5 %) removable prostheses harboured respiratory pathogenic bacteria. Stenotrophomonas maltophilia (34.0 %), Pseudomonas aeruginosa (27.8 %), and Streptococcus agalactiae (27.8 %) were the top three prevalent respiratory pathogens detected in plaque samples. There was a significantly higher prevalence of respiratory pathogens residing on unclean than clean prostheses (P = 0.046). However, the ORPIs in both groups showed no statistically significant difference (P = 0.516). CONCLUSIONS: The ORPIs for both clean and unclean prostheses demonstrated a similar abundance of respiratory pathogens. However, the high prevalence of respiratory pathogens residing on unclean prostheses should not be underestimated. Therefore, maintaining good prosthesis hygiene is still important for overall oral and systemic health, even though the direct link between prosthesis cleanliness and reduced abundance of respiratory pathogens has not been established. CLINICAL SIGNIFICANCE: The association between the prevalence of respiratory pathogens and unclean removable prostheses has been demonstrated and might increase the theoretical risk of respiratory disease development.

Upper respiratory Streptococcus pneumoniae colonization among working-age adults with prevalent exposure to overcrowding.

Most pneumococcal disease occurs among infants and older adults and is thought to be driven by the transmission of Streptococcus pneumoniae from young children to these vulnerable age groups. However, pneumococcal disease outbreaks also affect non-elderly adults living or working in congregate, close-contact settings. Little is known about pneumococcal carriage in such populations. From July to November 2020, we collected saliva from low-income adult farmworkers in Monterey County, California, and tested for pneumococcal carriage following culture enrichment via quantitative PCR assays targeting the pneumococcal lytA and piaB genes. Participants were considered to carry pneumococci if lytA and piaB cycle threshold values were both below 40. Among 1,283 participants enrolled in our study, 117 (9.1%) carried pneumococci. Carriers tended more often than non-carriers to be exposed to children aged <5 years [odds ratio (OR) = 1.45 (0.95-2.20)] and overcrowding [OR = 1.48 (0.96-2.30) and 2.84 (1.20-6.73), respectively, for participants in households with >2-4 and >4 persons per bedroom vs ≤2 persons per bedroom]. Household overcrowding remained associated with increased risk of carriage among participants not exposed to children aged <5 years [OR = 2.05 (1.18-3.59) for participants living in households with >2 vs ≤2 persons per bedroom]. Exposure to children aged <5 years and overcrowding were each associated with increased pneumococcal density among carriers [piaB cT difference of 2.04 (0.36-3.73) and 2.44 (0.80-4.11), respectively]. While exposure to young children was a predictor of pneumococcal carriage, associations of overcrowding with increased prevalence and density of carriage in households without young children suggest that transmission also occurs among adults in close-contact settings.IMPORTANCEAlthough infants and older adults are the groups most commonly affected by pneumococcal disease, outbreaks are known to occur among healthy, working-age populations exposed to overcrowding, including miners, shipyard workers, military recruits, and prisoners. Carriage of Streptococcus pneumoniae is the precursor to pneumococcal disease, and its relation to overcrowding in adult populations is poorly understood. We used molecular methods to characterize pneumococcal carriage in culture-enriched saliva samples from low-income adult farmworkers in Monterey County, CA. While exposure to children in the household was an important risk factor for pneumococcal carriage, living in an overcrowded household without young children was an independent predictor of carriage as well. Moreover, participants exposed to children or overcrowding carried pneumococci at higher density than those without such exposures, suggesting recent transmission. Our findings suggest that, in addition to transmission from young children, pneumococcal transmission may occur independently among adults in overcrowded settings.

Mucus polymer concentration and in vivo adaptation converge to define the antibiotic response of Pseudomonas aeruginosa during chronic lung infection.

The airway milieu of individuals with muco-obstructive airway diseases (MADs) is defined by the accumulation of dehydrated mucus due to hyperabsorption of airway surface liquid and defective mucociliary clearance. Pathological mucus becomes progressively more viscous with age and disease severity due to the concentration and overproduction of mucin and accumulation of host-derived extracellular DNA (eDNA). Respiratory mucus of MADs provides a niche for recurrent and persistent colonization by respiratory pathogens, including Pseudomonas aeruginosa, which is responsible for the majority of morbidity and mortality in MADs. Despite high concentration inhaled antibiotic therapies and the absence of antibiotic resistance, antipseudomonal treatment failure in MADs remains a significant clinical challenge. Understanding the drivers of antibiotic tolerance is essential for developing more effective treatments that eradicate persistent infections. The complex and dynamic environment of diseased airways makes it difficult to model antibiotic efficacy in vitro. We aimed to understand how mucin and eDNA concentrations, the two dominant polymers in respiratory mucus, alter the antibiotic tolerance of P. aeruginosa. Our results demonstrate that polymer concentration and molecular weight affect P. aeruginosa survival post antibiotic challenge. Polymer-driven antibiotic tolerance was not explicitly associated with reduced antibiotic diffusion. Lastly, we established a robust and standardized in vitro model for recapitulating the ex vivo antibiotic tolerance of P. aeruginosa observed in expectorated sputum across age, underlying MAD etiology, and disease severity, which revealed the inherent variability in intrinsic antibiotic tolerance of host-evolved P. aeruginosa populations. IMPORTANCE: Antibiotic treatment failure in Pseudomonas aeruginosa chronic lung infections is associated with increased morbidity and mortality, illustrating the clinical challenge of bacterial infection control. Understanding the underlying infection environment, as well as the host and bacterial factors driving antibiotic tolerance and the ability to accurately recapitulate these factors in vitro, is crucial for improving antibiotic treatment outcomes. Here, we demonstrate that increasing concentration and molecular weight of mucin and host eDNA drive increased antibiotic tolerance to tobramycin. Through systematic testing and modeling, we identified a biologically relevant in vitro condition that recapitulates antibiotic tolerance observed in ex vivo treated sputum. Ultimately, this study revealed a dominant effect of in vivo evolved bacterial populations in defining inter-subject ex vivo antibiotic tolerance and establishes a robust and translatable in vitro model for therapeutic development.

Inhaled antibiotics: A promising drug delivery strategies for efficient treatment of lower respiratory tract infections (LRTIs) associated with antibiotic resistant biofilm-dwelling and intracellular bacterial pathogens.

Antibiotic-resistant bacteria associated with LRTIs are frequently associated with inefficient treatment outcomes. Antibiotic-resistant Streptococcus pneumoniae, Haemophilus influenzae, Pseudomonas aeruginosa, and Staphylococcus aureus, infections are strongly associated with pulmonary exacerbations and require frequent hospital admissions, usually following failed management in the community. These bacteria are difficult to treat as they demonstrate multiple adaptational mechanisms including biofilm formation to resist antibiotic threats. Currently, many patients with the genetic disease cystic fibrosis (CF), non-CF bronchiectasis (NCFB) and chronic obstructive pulmonary disease (COPD) experience exacerbations of their lung disease and require high doses of systemically administered antibiotics to achieve meaningful clinical effects, but even with high systemic doses penetration of antibiotic into the site of infection within the lung is suboptimal. Pulmonary drug delivery technology that reliably deliver antibacterials directly into the infected cells of the lungs and penetrate bacterial biofilms to provide therapeutic doses with a greatly reduced risk of systemic adverse effects. Inhaled liposomal-packaged antibiotic with biofilm-dissolving drugs offer the opportunity for targeted, and highly effective antibacterial therapeutics in the lungs. Although the challenges with development of some inhaled antibiotics and their clinicals trials have been studied; however, only few inhaled products are available on market. This review addresses the current treatment challenges of antibiotic-resistant bacteria in the lung with some clinical outcomes and provides future directions with innovative ideas on new inhaled formulations and delivery technology that promise enhanced killing of antibiotic-resistant biofilm-dwelling bacteria.

Efficacy of liposomal bismuth-ethanedithiol-loaded tobramycin after intratracheal administration in rats with pulmonary Pseudomonas aeruginosa infection.

We sought to investigate alterations in quorum-sensing signal molecule N-acyl homoserine lactone secretion and in the release of Pseudomonas aeruginosa virulence factors, as well as the in vivo antimicrobial activity of bismuth-ethanedithiol incorporated into a liposome-loaded tobramycin formulation (LipoBiEDT-TOB) administered to rats chronically infected with P. aeruginosa. The quorum-sensing signal molecule N-acyl homoserine lactone was monitored by using a biosensor organism. P. aeruginosa virulence factors were assessed spectrophotometrically. An agar beads model of chronic Pseudomonas lung infection in rats was used to evaluate the efficacy of the liposomal formulation in the reduction of bacterial count. The levels of active tobramycin in the lungs and the kidneys were evaluated by microbiological assay. LipoBiEDT-TOB was effective in disrupting both quorum-sensing signal molecules N-3-oxo-dodeccanoylhomoserine lactone and N-butanoylhomoserine lactone, as well as significantly (P < 0.05) reducing lipase, chitinase, and protease production. At 24 h after 3 treatments, the CFU counts in lungs of animals treated with LipoBiEDT-TOB were of 3 log(10) CFU/lung, comparated to 7.4 and 4.7 log(10) CFU/lung, respectively, in untreated lungs and in lungs treated with free antibiotic. The antibiotic concentration after the last dose of LipoBiEDT-TOB was 25.1 µg/lung, while no tobramycin was detected in the kidneys. As for the free antibiotic, we found 6.5 µg/kidney but could not detect any tobramycin in the lungs. Taken together, LipoBiEDT-TOB reduced the production of quorum-sensing molecules and virulence factors and could highly improve the management of chronic pulmonary infection in cystic fibrosis patients.

Antibacterial activity of the human host defence peptide LL-37 and selected synthetic cationic lipids against bacteria associated with oral and upper respiratory tract infections.

OBJECTIVES: We aim to develop antibacterial peptide mimics resistant to protease degradation, with broad-spectrum activity at sites of infection. METHODS: The bactericidal activities of LL-37, ceragenins CSA-13, CSA-90 and CSA-92 and the spermine-conjugated dexamethasone derivative D2S were evaluated using MIC and MBC measurements. Gingival fibroblast counting, interleukin-8 (IL-8) and lactate dehydrogenase (LDH) release from keratinocytes (HaCat) were used to determine effects on cell growth, pro-inflammatory response and toxicity. RESULTS: All tested cationic lipids showed stronger bactericidal activity than LL-37. Incubation of Staphylococcus aureus with half the MIC of LL-37 led to the appearance of bacteria resistant to its bactericidal effects, but identical incubations with CSA-13 or D2S did not produce resistant bacteria. Cathelicidin LL-37 significantly increased the total number of gingival fibroblasts, but ceragenins and D2S did not alter gingival fibroblast growth. Cationic lipids showed no toxicity to HaCat cells at concentrations resulting in bacterial killing. CONCLUSIONS: These data suggest that cationic lipids such as ceragenins warrant further testing as potential novel antibacterial agents.

Novel diagnostics: progress toward a breath test for invasive Aspergillus fumigatus.

In this paper we will briefly review some of the possible techniques for the development of a breath test for aspergillosis and describe progress made toward validating 2-Pentyl furan (2PF) as a marker of Aspergillus infection. Breath testing to diagnose pulmonary aspergillosis is attractive because of the proximity of the lesion to the sample and the simplicity of obtaining diagnostic specimens. Techniques to detect volatile organic compounds (VOCs) in breath discussed include the electronic nose, selective ion flow mass spectrometry (SIFT), ion mobility spectrometry (IMS) and gas chromatography with mass spectrometry (GC/MS). We have used GC/MS to identify Aspergillus-derived VOC's in the head space of cultures. A promising diagnostic marker molecule is 2PF, which was not detectable from cultures of bacterial respiratory pathogens with the possible exception of Streptococcus pneumoniae. 2-Pentylfuran is not known to be produced by mammalian metabolism and was not detectable in the breath of healthy controls, or neutropenic subjects. In contrast, 2PF was found in the breath of patients with lung disease who were colonized or infected with A. fumigatus. Case reports are presented of two severely immune compromised patients with invasive aspergillosis from whom 2PF was detected in multiple breath samples but became undetectable with effective treatment. A well conducted prospective trial is needed to validate the clinical usefulness of this marker for diagnosis and monitoring of invasive aspergillosis. Unanswered questions remaining include how much 2PF, if any, is produced by extensive lung inflammation, and whether food containing high levels of 2PF can cause false positive breath tests either from contamination in the mouth or gastrointestinal absorption and subsequent excretion in the breath.

A comparison of cotton-tipped and nylon flocked swabs for culture of Neisseria gonorrhoeae from oropharyngeal samples.

Our aim was to determine if there was a difference in culture positivity for oropharyngeal gonorrhoea when sampling using a nylon-flocked versus cotton-tipped swab. We collected FLOQSwabs and cotton-tipped swabs from individuals aged ≥ 18 years who had untreated oropharyngeal gonorrhoea detected by NAAT between November 2019-June 2020.Of 78 participants, 32 (41.0%) were culture-positive for N. gonorrhoeae from either swab. Of these 32, 29 (90.6%, 95%CI: 75.0%-98.0%) were positive on both swabs, one (3.1%, 95%CI: 0.0%-16.2%) tested positive on FLOQSwab only and two (6.2%, 95%CI: 0.1%-20.8%) tested positive on cotton-tipped swabs only. There was moderate agreement between the swabs in the amount of bacterial growth (Cohen's Kappa (k)=0.745; 95%CI: 0.622-0.868, p<0.001). Our results showed that the proportion of positive results was comparable using the FLOQSwabs versus the cotton-tipped swabs for oropharyngeal gonorrhoea culture.