Adhesion of Klebsiella oxytoca to bladder or lung epithelial cells is promoted by the presence of other opportunistic pathogens.

The intestinal and respiratory tracts of healthy individuals serve as habitats for a diverse array of microorganisms, among which Klebsiella oxytoca holds significance as a causative agent in numerous community- and hospital-acquired infections, often manifesting in polymicrobial contexts. In specific circumstances, K. oxytoca, alongside other constituents of the gut microbiota, undergoes translocation to distinct physiological niches. In these new environments, it engages in close interactions with other microbial community members. As this interaction may progress to co-infection where the virulence of involved pathogens may be promoted and enhance disease severity, we investigated how K. oxytoca affects the adhesion of commonly co-isolated bacteria and vice versa during co-incubation of different biotic and abiotic surfaces. Co-incubation was beneficial for the adhesion of at least one of the two co-cultured strains. K. oxytoca enhanced the adhesion of other enterobacteria strains to polystyrene and adhered more efficiently to bladder or lung epithelial cell lines in the presence of most enterobacteria strains and S. aureus. This effect was accompanied by bacterial coaggregation mediated by carbohydrate-protein interactions occurring between bacteria. These interactions occur only in sessile, but not planktonic populations, and depend on the features of the surface. The data are of particular importance for the risk assessment of the urinary and respiratory tract infections caused by K. oxytoca, including those device-associated. In this paper, we present the first report on K. oxytoca ability to acquire increased adhesive capacities on epithelial cells through interactions with common causal agents of urinary and respiratory tract infections.

Non-classical Bordetella sp. (closely related to Bordetella hinzii and Bordetella pseudohinzii) lower respiratory tract infection in a patient with extensive bronchiectasis: a case report.

An increasing number of reports have described the pathogenic nature of several non-classical Bordetella spp. Among them, Bordetella hinzii and Bordetella pseudohinzii have been implicated in a myriad of respiratory-associated infections in humans and animals. We report the isolation of a genetically close relative of B. hinzii and B. pseudohinzii from the sputum of a woman in her early 60s with extensive bronchiectasis who presented with fever and brown colored sputum. The isolate had initially been identified as Bordetella avium by API 20NE, the identification system for non-enteric Gram-negative rod bacteria. Sequencing of the 16S rDNA, ompA, nrdA, and genes used in the Bordetella multilocus sequence typing scheme could not resolve the identity of this Bordetella isolate. Whole-genome single nucleotide polymorphism analysis positioned the isolate between B. hinzii and B. pseudohinzii in the phylogenetic tree, forming a distinct cluster. Whole-genome sequencing enabled the further identification of this rare organism, and should be considered for wider applications, especially the confirmation of organism identity in the clinical diagnostic microbiology laboratory.

Novel Approaches to Multidrug-Resistant Infections in Cystic Fibrosis.

Patients with cystic fibrosis (CF) often develop respiratory tract infections with pathogenic multidrug-resistant organisms (MDROs) such as methicillin-resistant Staphylococcus aureus, and a variety of gram-negative organisms that include Pseudomonas aeruginosa, Burkholderia sp., Stenotrophomonas maltophilia, Achromobacter xylosoxidans, and nontuberculous mycobacteria (NTM). Despite the introduction of new therapies to address underlying cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction, MDRO infections remain a problem and novel antimicrobial interventions are still needed. Therapeutic approaches include improving the efficacy of existing drugs by adjusting the dose based on differences in CF patient pharmacokinetics/pharmacodynamics, the development of inhaled formulations to reduce systemic adverse events, and the use of newer beta-lactam/beta-lactamase combinations. Alternative innovative therapeutic approaches include the use of gallium and bacteriophages to treat MDRO pulmonary infections including those with extreme antibiotic resistance. However, additional clinical trials are required to determine the optimal dosing and efficacy of these different strategies and to identify patients with CF most likely to benefit from these new treatment options.

Quantitative Value of Bacteria Associated with Leukocytes in Differential Diagnosis of Lower Respiratory Tract Infection in Children, in Comparison to Sputum Culture and Procalcitonin.

BACKGROUND: The mortality and morbidity rates in children with lower respiratory tract infection (LRTI) remain high. OBJECTIVE: To describe the number of bacteria that is associated with leukocytes in differential diagnosis of bacterial, mycoplasma, and viral LRTI in children. METHODS: Sputum smears were Gram stained for counting single-morphology bacteria associated with leukocytes. The differential diagnostic values of bacterial number were assessed in children with LRTI. RESULTS: The area under the receiver operating characteristic (ROC) curve was 0.95 for bacterial number in the differential diagnosis of bacterial infection from mycoplasma and viral infections. The area under the ROC curve was 0.62 for procalcitonin and 0.94 for bacterial number in the differential diagnosis of bacterial infection from mycoplasma infection. CONCLUSION: The number of bacteria associated with leukocytes in sputum was valuable and rapid in differential diagnosis of bacterial infection in children with suspected bacterial, mycoplasma, and viral LRTI.

Clinical Presentation and Co-Detection of Respiratory Pathogens in Children Under 5 Years with Non-COVID-19 Bacterial and Viral Respiratory Tract Infections: A Prospective Study in Bialystok, Poland (2021-2022).

BACKGROUND Respiratory tract infections (RTIs) in children often involve a complex interplay between viruses and bacteria. This study aimed to evaluate clinical presentation in children under 5 years old diagnosed with non-COVID-19 bacterial and viral respiratory tract co-infections between October 2021 and May 2022 in Bialystok, Poland. MATERIAL AND METHODS We recruited 100 children under 5 years with RTIs who tested negative for SARS-CoV-2. Nasopharyngeal swabs were screened for 19 viruses and 7 bacterial strains using molecular assays. RESULTS Viral pathogens were detected in 71% of patients and bacterial pathogens were detected in 59%. The most common pathogens were Haemophilus influenzae (n=48), rhinoviruses (n=32), and Streptococcus pneumoniae (n=30). Single pathogens were detected in 36%, dual in 37%, triple in 15%, and quadruple in 2%. Bacterial pathogens were co-detected with viruses in 40 cases, mostly with rhinoviruses (n=15). Two different viruses were found in 14 children and the most common co-detection was adenovirus with rhinovirus (n=5); dyspnea (63% vs 11%) and wheezing (75% vs 22%) were more common in children with human bocavirus. Fever was a common symptom in children with human adenovirus (88% vs 58%). Detection of bacteria and multiple detections were more common in day-care attendees, but were not associated with clinical picture of RTI. CONCLUSIONS Consistent with previous studies, we found a high prevalence of rhinoviruses, despite ongoing implementation of non-pharmaceutical interventions to contain the COVID-19 pandemic. Co-detection of 2 different respiratory pathogens was frequent, but we found no evidence that this was associated with the severity of infections.

Evaluation of the BioFire® FilmArray® Pneumonia plus Panel for Detecting Bacterial Etiological Agents of Lower Respiratory Tract Infections in an Oncologic Hospital. Comparison with Conventional Culture Method.

Conventional methods used to determine pneumonia pathogens are characterized by low sensitivity and long turnaround times. Introducing new tests with better parameters in patients at higher risk of infections is highly anticipated. The results of the conventional quantitative culture method (CM) in determining the bacterial etiology of pneumonia were compared with the results of the Pneumonia plus Panel test (PNP; BioFire® Diagnostics, USA) in 79 samples of bronchoalveolar lavage (BAL). Materials were collected from 79 patients with suspected pneumonia treated in an oncologic hospital due to solid tumors. Only 16/79 BAL samples (20.3%) were true positive (TP) for bacterial etiology in CM vs. 27/79 samples (34.2%) true positive in the PNP test. The total agreement between methods of interpreting the result (positive or negative) was 84.8%. The most prevalent pathogens in both methods were Staphylococcus aureus, followed by Escherichia coli, Pseudomonas aeruginosa, and Haemophilus influenzae. The PNP test identified several respiratory pathogens that were not grown in culture. The semiquantitative value reported by the PNP test was higher than that reported by culture. The PNP test vs. combined test (PNP test and CM methods) demonstrated positive predictive value (PPV) and negative predictive value (NPV) values of 100.0% and 98.1%, and the sensitivity and specificity were 96.4% and 100.0%. The PNP test is a good tool for determining the etiology of bacterial pneumonia and may support the care of an oncologic patient. However, further large-sample studies are needed to research in strictly defined groups of oncologic patients.

Diagnostic value of loop-mediated isothermal amplification in detecting lower respiratory pathogens.

INTRODUCTION: To investigate the diagnostic value of loop-mediated thermostatic amplification (LAMP) in detecting pathogenic bacteria from bronchoalveolar lavage fluid (BALF) of patients with pulmonary disorders combined with lower respiratory tract infections (LRTI). METHODOLOGY: This cross-sectional study included patients with pulmonary disorders combined with LRTI, including chronic obstructive pulmonary disease (COPD), bronchiectasis, or lung cancer, hospitalized in Meizhou People's Hospital between January 2020 and October 2021. BALF was collected using local bronchoalveolar lavage and electronic bronchoscopy. The presence of the pathogens was confirmed using the LAMP method and the bacterial culture method. RESULTS: In total, 249 patients were included (135 with COPD, 73 with bronchiectasis, and 41 with lung cancer). The proportions of Methicillin-resistant Staphylococcus aureus (4.8% vs 0.4%, p = 0.02) and Haemophilus influenzae (6.8% vs 0.4%, p < 0.001) detected by the LAMP method was higher, while the proportion of Pseudomonas aeruginosa was lower compared with that of the culture method (6.8% vs 12.4%, p = 0.034). The bacterial species with the highest agreement coefficient was Stenotrophomonas maltophilia (Kappa = 0.798, p < 0.001). Furthermore, 9 COPD patients exhibited mixed infections as determined by the LAMP method, whereas the culture method detected only 2 of these cases (1.48%) (p < 0.05). CONCLUSIONS: LAMP can detect more pathogenic bacteria, notably Haemophilus influenza, Methicillin-resistant Staphylococcus aureus, and atypical pathogens in patients with clinically common pulmonary disorders combined with LRTI. LAMP may provide etiological evidence to guide the clinical use of antibiotics in primary hospitals.

Host test based on tumor necrosis factor-related apoptosis-inducing ligand, interferon gamma-induced protein-10 and C-reactive protein for differentiating bacterial and viral respiratory tract infections in adults: diagnostic accuracy study.

OBJECTIVES: To assess the performance of a test (called BV), integrating the blood levels of three immune proteins into a score, to differentiate bacterial from viral infection among adults with suspected lower respiratory tract infection (LRTI). METHODS: Prospective diagnostic accuracy study, enrolling febrile adults >18 years with LRTI signs or symptoms for less than 7 days presenting to several hospitals' emergency departments in Israel. The main exclusion criterion was immunodeficiency. Reference standard diagnosis (bacterial/viral/indeterminate) was based on three experts independently reviewing comprehensive patient data including follow-up data. BV generated three results: viral infection or other nonbacterial condition (0 ≤ score < 35), equivocal (35 ≤ score ≤ 65) and bacterial infection including co-infection (65 < score ≤ 100). BV performance was assessed against the reference standard with indeterminate reference standard and equivocal BV cases removed. RESULTS: Of 490 enrolled patients, 415 met eligibility criteria (median age 56 years, interquartile range 35). The reference standard classified 104 patients as bacterial, 210 as viral and 101 as indeterminate. BV was equivocal in 9.6% (30/314). Excluding indeterminate reference standard diagnoses and equivocal BV results, BV's sensitivity for bacterial infection was 98.1% (101/103; 95% confidence interval 95.4-100), specificity 88.4% (160/181; 83.7-93.1) and negative predictive value 98.8% (160/162; 97.1-100). DISCUSSION: BV exhibited high diagnostic performance for febrile adults with suspected LRTI among patients with reference standard diagnoses of bacterial or viral LRTI.

The Clinical Performance of the BioCode Respiratory Pathogen Panel for the Detection of Viruses and Bacteria from Nasopharyngeal Swabs.

Early detection of microbial pathogens causing respiratory tract infection plays a crucial role in clinical management. The BioCode Respiratory Pathogen Panel (BioCode RPP) utilizes reverse transcriptase PCR (RT-PCR) in combination with barcoded magnetic beads to amplify, detect, and identify respiratory pathogens. This panel qualitatively detects and identifies 14 viruses, including influenza virus A with H1 pdm09, H1, and H3 subtyping; influenza B; respiratory syncytial virus (RSV); human metapneumovirus; parainfluenza virus 1; parainfluenza virus 2; parainfluenza virus 3; parainfluenza virus 4; coronavirus (229E, NL63, OC43, and HKU1); adenovirus; and human rhinovirus/enterovirus, and 3 bacteria, including Chlamydia pneumoniae, Mycoplasma pneumoniae, and Bordetella pertussis. Reproducibility, which was assessed with contrived specimens containing 12 targets at 3 clinical sites, with 2 operators at each site for 5 days, was 99.4% for Flu A H3 and Flu B, 98.9% for RSV, and 100% for the remaining 9 targets assayed. A multicenter clinical trial evaluated the performance of the BioCode RPP with 2,647 nasopharyngeal swab specimens from 5 geographically distinct sites and revealed comparable performance between the BioCode RPP and FilmArray Respiratory Panel (FA-RP). Specifically, the positive percent agreements (PPAs) for various pathogens ranged between 80.8% and 100% compared with the FA-RP (1.7 and 2.0). Negative percent agreement ranged from 98.4% to 100% for BioCode RPP. The BioCode RPP also offers scalable automated testing capability of up to 96 specimens in a single run with total sample-to-result time under 5 h. The invalid rate of the BioCode RPP on initial testing was 1.0% (26/2,649). IMPORTANCE Early detection of microbial pathogens causing respiratory tract infection plays a crucial role in clinical management. The BioCode Respiratory Pathogen Panel (BioCode RPP) is a high-throughput test that utilizes RT-PCR in combination with barcoded magnetic beads to amplify, detect, and identify 17 respiratory pathogens, including 14 viruses and 3 bacteria. This study summarizes data generated from a multicenter clinical trial evaluating the performance of the BioCode RPP on 2,647 nasopharyngeal swab specimens from five geographically distinct sites.

Diagnosis of invasive respiratory mycoses in the immunocompromised host.

PURPOSE OF REVIEW: The burden of invasive fungal infection is increasing worldwide, largely due to a growing population at-risk. Most serious human fungal pathogens enter the host via the respiratory tract. Early identification and treatment of invasive fungal respiratory infections (IFRIs) in the immunocompromised host saves lives. However, their accurate diagnosis is a difficult challenge for clinicians and mortality remains high. RECENT FINDINGS: This article reviews IFRIs, focussing on host susceptibility factors, clinical presentation, and mycological diagnosis. Several new diagnostic tools are coming of age including molecular diagnostics and point-of-care antigen tests. As diagnosis of IFRI relies heavily on invasive procedures like bronchoalveolar lavage and lung biopsy, several novel noninvasive diagnostic techniques are in development, such as metagenomics, 'volatilomics' and advanced imaging technologies. SUMMARY: Where IFRI cannot be proven, clinicians must employ a 'weights-of-evidence' approach to evaluate host factors, clinical and mycological data. Implementation studies are needed to understand how new diagnostic tools can be best applied within clinical pathways. Differentiating invasive infection from colonization and identifying antifungal resistance remain key challenges. As our diagnostic arsenal expands, centralized clinical mycology laboratories and efforts to ensure access to new diagnostics in low-resource settings will become increasingly important.

Advances in inhaled antibiotics for management of respiratory tract infections.

PURPOSE OF REVIEW: The incidence of bacterial respiratory tract infections is growing. In a context of increasing antibiotic resistance and lack of new classes of antibiotics, inhaled antibiotics emerge as a promising therapeutic strategy. Although they are generally used for cystic fibrosis, their use in other conditions is becoming more frequent, including no-cystic fibrosis bronchiectasis, pneumonia and mycobacterial infections. RECENT FINDINGS: Inhaled antibiotics exert beneficial microbiological effects in bronchiectasis and chronic bronchial infection. In nosocomial and ventilator-associated pneumonia, aerosolized antibiotics improve cure rates and bacterial eradication. In refractory Mycobacterium avium complex infections, amikacin liposome inhalation suspension is more effective in achieving long-lasting sputum conversion. In relation to biological inhaled antibiotics (antimicrobial peptides, interfering RNA and bacteriophages), currently in development, there is no still enough evidence that support their use in clinical practice. SUMMARY: The effective antimicrobiological activity of inhaled antibiotics, added to their potential to overcoming resistances to systemic antibiotics, make inhaled antibiotics a plausible alternative.

Nontubercular bacterial and fungal infections in patients of chronic obstructive pulmonary disease.

Introduction: Acute exacerbation of chronic obstructive pulmonary disease (COPD), frequently due to respiratory tract infection is the major cause of morbidity and mortality, and estimate suggests that it is currently the third leading cause of death worldwide. Aims and Objectives: This study aims to study the prevalence of nontubercular bacterial and fungal infections in patients of COPD. Materials and Methods: It is an observational study done for 1-year period from August 2017 to July 2018. A total of 100 COPD patients who fulfilled the inclusion and exclusion criteria were analyzed in the present study. These cases were classified according to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) combined assessment criteria and subjected to sputum or in some cases Bronchoalveolar lavage (BAL) fluid examination for nontubercular bacterial and fungal pathogens. Serum galactomannan assay, bronchoscopy, and computed tomography chest were done in selected cases. Results: The age of the study population ranged from 40 to 85 years and the mean age was 60.01 ± 9.85 years. Majority of the patients were male (81.0%) and most (78.0%) of them were smokers. Most of the patients belonged to GOLD Grades 2 and 3. Forty-six percent of the patients did show pathogenic organisms in sputum examination. Out of these, 80.4% were bacterial, mainly Gram-negative organisms (Acinetobacter, Pseudomonas, Escherichia coli, Enterobacter, Proteus, and Citrobacter) and 19.6% of cases were having fungal infections (Candida and Aspergillus). Conclusions: Increasing patient age, smoking habit, and severity of COPD were related to an increasing frequency of bacterial and fungal infections. Early detection and proper treatment could help in preventing the morbidity and mortality related to COPD.

Résumé Introduction: L'exacerbation aiguë de la maladie pulmonaire obstructive chronique (MPOC), souvent en raison de l'infection des voies respiratoires, est la principale cause de morbidité et de mortalité, et l'estimation suggère qu'il s'agit actuellement de la troisième cause de décès dans le monde. Objectifs et objectifs: Cette étude vise à étudier la prévalence des infections bactériennes et fongiques non tubulaires chez les patients de la MPOC. Matériaux et méthodes: Il s'agit d'une étude d'observation réalisée pour une période de 1 an d'août 2017 à juillet 2018. Un total de 100 patients atteints de MPOC qui remplissaient les critères d'inclusion et d'exclusion ont été analysés dans la présente étude. Ces cas ont été classés selon l'initiative globale des critères d'évaluation combinés chroniques obstructifs (OR) et soumis à des expectorations ou dans certains cas examen des liquides de lavage bronchoalvéolaire (BAL) pour les agents pathogènes bactéries et fongiques non tubulaires. Le test de galactomannane sérique, la bronchoscopie et le poitrine de tomodensitométrie ont été effectués dans certains cas. Résultats: L'âge de la population d'étude variait de 40 à 85 ans et l'âge moyen était de 60,01 ± 9,85 ans. La majorité des patients étaient des hommes (81,0%) et la plupart (78,0%) d'entre eux étaient des fumeurs. La plupart des patients appartenaient à GOLD GRADES 2 et 3. Quarante-six pour cent des patients ont montré des organismes pathogènes à l'examen des expectorations. Parmi ceux-ci, 80,4% étaient des organismes bactériens, principalement à Gram - négatifs (Acinetobacter, Pseudomonas, Escherichia coli, Enterobacter, Proteus et Citrobacter) et 19,6% des cas avaient des infections fongiques (Candida et 23 aspergillus). Conclusions: L'âge croissant du patient, l'habitude du tabagisme et la gravité de la MPOC étaient liés à une fréquence croissante des infections bactériennes et fongiques. La détection précoce et le traitement approprié pourraient aider à prévenir la morbidité et la mortalité liées à la MPOC. Mots-clés: Maladie pulmonaire obstructive chronique, infection fongique, initiative mondiale pour la maladie pulmonaire obstructive chronique, infection bactérienne non tuberculeuse.

Clinical outcomes associated with Achromobacter species infection in people with cystic fibrosis.

BACKGROUND: Achromobacter species are emerging pathogens isolated from respiratory samples of Patients with cystic fibrosis (pwCF) causing growing concerns in the CF community. The epidemiology and the clinical impact of Achromobacter in CF is unclear since data are restricted to small case control studies or selected populations. AIM: To characterize the effect of Achromobacter respiratory infection on CF lung disease. METHODS: European CF Society Patient Registry data was analysed for association between Achromobacter infection and demographic/clinical characteristics and outcomes of pwCF. RESULTS: Of eligible 38,795 patients, Achromobacter infection was reported in 2,093 (prevalence (95% CI) of 5.40% (5.17 - 5.62). The prevalence varied significantly between the countries and increased with age peaking at the age 20-30. Achromobacter infection was more prevalent in pwCF carrying class minimal function mutations, having worse nutrition or lower pulmonary function, and more patients inhaled antibiotics against P. aeruginosa. Patient infected with Achromobacter had similar pulmonary function and BMI to patients infected with P. aeruginosa at all age groups. Being infected with both bacteria was associated with significantly lower pulmonary function and BMI at all age groups. CONCLUSIONS: Achromobacter infection was associated with disease severity similar to infection with P. aeruginosa. Being infected with both bacteria is associated with even more severe disease. This suggests to study if eradication will improve the outcome of pwCF.

Combination of Cefditoren and N-acetyl-l-Cysteine Shows a Synergistic Effect against Multidrug-Resistant Streptococcus pneumoniae Biofilms.

Biofilm formation by Streptococcus pneumoniae is associated with colonization of the upper respiratory tract, including the carrier state, and with chronic respiratory infections in patients suffering from chronic obstructive pulmonary disease (COPD). The use of antibiotics alone to treat recalcitrant infections caused by biofilms is insufficient in many cases, requiring novel strategies based on a combination of antibiotics with other agents, including antibodies, enzybiotics, and antioxidants. In this work, we demonstrate that the third-generation oral cephalosporin cefditoren (CDN) and the antioxidant N-acetyl-l-cysteine (NAC) are synergistic against pneumococcal biofilms. Additionally, the combination of CDN and NAC resulted in the inhibition of bacterial growth (planktonic and biofilm cells) and destruction of the biofilm biomass. This marked antimicrobial effect was also observed in terms of viability in both inhibition (prevention) and disaggregation (treatment) assays. Moreover, the use of CDN and NAC reduced bacterial adhesion to human lung epithelial cells, confirming that this strategy of combining these two compounds is effective against resistant pneumococcal strains colonizing the lung epithelium. Finally, administration of CDN and NAC in mice suffering acute pneumococcal pneumonia caused by a multidrug-resistant strain was effective in clearing the bacteria from the respiratory tract in comparison to treatment with either compound alone. Overall, these results demonstrate that the combination of oral cephalosporins and antioxidants, such as CDN and NAC, respectively, is a promising strategy against respiratory biofilms caused by S. pneumoniae. IMPORTANCE Streptococcus pneumoniae is one of the deadliest bacterial pathogens, accounting for up to 2 million deaths annually prior to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Vaccines have decreased the burden of diseases produced by S. pneumoniae, but the rise of antibiotic-resistant strains and nonvaccine serotypes is worrisome. Pneumococcal biofilms are associated with chronic respiratory infections, and treatment is challenging, making the search for new antibiofilm therapies a priority as biofilms become resistant to traditional antibiotics. In this work, we used the combination of an antibiotic (CDN) and an antioxidant (NAC) to treat the pneumococcal biofilms of relevant clinical isolates. We demonstrated a synergy between CDN and NAC that inhibited and treated pneumococcal biofilms, impaired pneumococcal adherence to the lung epithelium, and treated pneumonia in a mouse pneumonia model. We propose the widely used cephalosporin CDN and the repurposed drug NAC as a new antibiofilm therapy against S. pneumoniae biofilms, including those formed by antibiotic-resistant clinical isolates.

Urinary tract infection caused by bacterial pathogens of the respiratory tract in children.

BACKGROUND: Urinary tract infections (UTIs) caused by bacterial pathogens of the respiratory tract such as Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis are rare and little is known about their clinical features and potential host risk factors. The aim of this study is to reveal their clinical characteristics. METHODS: We conducted a retrospective descriptive study on pediatric UTI due to S. pneumoniae, Haemophilus spp., or M. catarrhalis at a tertiary-care pediatric hospital. Pediatric patients diagnosed with UTI between 2002 and 2020 were included. Patient demographics, laboratory data, and microbiological findings were extracted from their electronic medical records and the infectious disease surveillance system. RESULTS: Among 46,332 urine samples, 76 bacteriuria (0.16%) and 22 UTI (0.05%) events due to the targeted species were identified (S. pneumoniae, n = 7, and Haemophilus spp., n = 15). Of the patients, 17 (85%) had underlying urinary tract abnormalities and 13 (60%) had vesicocutaneous fistula. All the UTI episodes caused by S. pneumoniae and Haemophilus spp. occurred after cystostomy. All the patients had satisfactory clinical outcomes. CONCLUSIONS: Although S. pneumoniae and Haemophilus spp. are rare causes of UTIs in children, they could be the true causative bacteria of UTI, particularly in the patients with urinary tract abnormalities and vesicocutaneous fistulas. Thus, clinicians should not ignore these pathogens as contaminations in special populations.

Adaptation and Evolution of Pathogens in the Cystic Fibrosis Lung.

As opposed to acute respiratory infections, the persistent bacterial infections of the lung that characterize cystic fibrosis (CF) provide ample time for bacteria to evolve and adapt. The process of adaptation is recorded in mutations that accumulate over time in the genomes of the infecting bacteria. Some of these mutations lead to obvious phenotypic differences such as antibiotic resistance or the well-known mucoid phenotype of Pseudomonas aeruginosa. Other mutations may be just as important but harder to detect such as increased mutation rates, cell surface changes, and shifts in metabolism and nutrient acquisition. Remarkably, many of the adaptations occur again and again in different patients, signaling that bacteria are adapting to solve specific challenges in the CF respiratory tract. This parallel evolution even extends across distinct bacterial species. This review addresses the bacterial systems that are known to change in long-term CF infections with a special emphasis on cross-species comparisons. Consideration is given to how adaptation may impact health in CF, and the possible evolutionary mechanisms that lead to the repeated parallel adaptations.

Development of a three-panel multiplex real-time PCR assay for simultaneous detection of nine canine respiratory pathogens.

Infectious respiratory disease is one of the most common diseases in dogs worldwide. Several bacterial and viral pathogens can serve as causative agents of canine infectious respiratory disease (CIRD), including Mycoplasma cynos, Mycoplasma canis, Bordetella bronchiseptica, canine adenovirus type 2 (CAdV-2), canine herpesvirus 1 (CHV-1), canine parainfluenza virus (CPIV), canine distemper virus (CDV), canine influenza virus (CIA) and canine respiratory coronavirus (CRCoV). Since these organisms cause similar clinical symptoms, disease diagnosis based on symptoms alone can be difficult. Therefore, a quick and accurate test is necessary to rapidly identify the presence and relative concentrations of causative CIRD agents. In this study, a multiplex real-time PCR panel assay was developed and composed of three subpanels for detection of the aforementioned pathogens. Correlation coefficients (R2) were >0.993 for all singleplex and multiplex real-time PCR assays with the exception of one that was 0.988; PCR amplification efficiencies (E) were between 92.1% and 107.8% for plasmid DNA, and 90.6-103.9% for RNA templates. In comparing singular and multiplex PCR assays, the three multiplex reactions generated similar R2 and E values to those by corresponding singular reactions, suggesting that multiplexing did not interfere with the detection sensitivities. The limit of detection (LOD) of the multiplex real-time PCR for DNA templates was 5, 2, 3, 1, 1, 1, 4, 24 and 10 copies per microliter for M. cynos, M. canis, B. brochiseptica, CAdV-2, CHV-1, CPIV, CDV, CIA and CRCoV, respectively; and 3, 2, 6, 17, 4 and 8 copies per microliter for CAdV-2, CHV-1, CPIV, CDV, CIA and CRCoV, respectively, when RNA templates were used for the four RNA viruses. No cross-detection was observed among the nine pathogens. For the 740 clinical samples tested, the newly designed PCR assay showed higher diagnostic sensitivity compared to an older panel assay; pathogen identities from selected samples positive by the new assay but undetected by the older assay were confirmed by Sanger sequencing. Our data showed that the new assay has higher diagnostic sensitivity while maintaining the assay's specificity, as compared to the older version of the panel assay.

Causes of Lower Respiratory Tract Infections and the Use of Diagnostic Biomarkers in Blood Samples from Children in Hohhot, Inner Mongolia, China, Between July 2019 and June 2020.

BACKGROUND Lower respiratory tract infection (LRTI) in children is due to various pathogens. Appropriate diagnosis and early treatment are important for reducing the mortality rate of LRTI. Data on the epidemiology profiles of LRTI are scarce in northern China. The aim of this study was to provide data on the pathogen pattern of LRTI in hospitalized children in Hohhot, Inner Mongolia, China. MATERIAL AND METHODS From July 2019 to June 2020, nasopharyngeal swabs were collected from 265 children in Hohhot with LRTI, and pathogens were detected with RT-PCR and PCR. The correlations among procalcitonin (PCT), C-reactive protein (CRP), and white blood cells (WBC) with acute respiratory infections were evaluated. RESULTS The highest prevalence of LRTI was detected in 2- to 6-year-old children (149, 56.2%) in winter. Eleven respiratory pathogens were evaluated, and respiratory syncytial virus, Streptococcus pneumoniae and Haemophilus influenza were the most common pathogens in this region. Single viruses, bacteria, mycoplasma, and multiple pathogens were identified in 24.2, 15.8, 5.3, and 54.7% of patients, respectively. The mean blood biomarker values of patients with LRTI were significantly different from those of healthy children. Furthermore, The AUCs were 0.90, 0.74, and 0.84 for bacteria, virus, and mycoplasma PCT values, which were significantly higher than that of WBC and CRP. CONCLUSIONS This evaluation of the regional pattern of pathogens in children with acute respiratory infections and the correlation with blood biomarkers provides valuable information for the prevention and treatment of LRTI in children.

Cytopathological analysis of bronchoalveolar lavage fluid in patients with and without HIV infection.

BACKGROUND: Human immunodeficiency virus/acquired immunodeficiency syndrome (HIV/AIDS) infection can lead to a broad spectrum of lung diseases, including infectious diseases and tumors. Recently, with the wide application of bronchoscopes and cytopathology of bronchoalveolar lavage fluid (BALF), the diagnostic efficiency of lung diseases has improved. The present study focuses on analyzing the cytopathologic characteristics of BALF in the diagnosis of HIV/AIDS-related lung disease and comparing the lung disease spectrum between HIV and HIV-uninfected patients. METHODS: BALF specimens were collected from 2211 patients. Using ThinPrep liquid-based technology, the cytologic smears were prepared by staining with Hematoxylin and Eosin (HE), Gomori's methenamine silver (GMS), and Periodic Acid Schiff (PAS), acid-fast and immunocytochemical (ICC) staining. Real-time PCR was used to detect cytomegalovirus (CMV) and Mycobacterium tuberculosis (M. tuberculosis) in the remaining BALF. PCR-reverse dot hybridization was used for mycobacterial species identification. RESULTS: From the 2211 BALF specimens, 1768 (79.96%) were specimens from HIV-infected patients, and 443 (20.04%) were speciments from HIV-uninfected patients. The HIV-infected patients with a median age of 38.5 ± 11.3 years were markedly younger than the HIV-uninfected patients (52.9 ± 14.9 years) (p < 0.01). We found that 1635 (92.5%) HIV-infected patients were males, showing a prominently higher proportion than those without HIV infection (71.1%) (p < 0.01). Meanwhile, 1045 specific lesions were found in 1768 HIV-infected patients (59.1%), including 1034 cases of infectious diseases and 11 neoplastic lesions, also exhibiting a distinctly higher proportion compared to the HIV-uninfected patients (12.2%) (p < 0.001). For the HIV-infected group, a distinctly higher proportion of single infection lesions (724/1768, 41%) was noted than the HIV-uninfected group (14/443, 3.2%) (p < 0.001). Among single infection lesions, the most common was Cytomegalovirus(CMV) infection (20.9%) for the HIV-infected group, followed by Pneumocystis jiroveci(PJ) (13.0%), Fungal (3.5%), and Mycobacterial infections (3.4%), of which M. tuberculosis infection accounted for 3.1%. Double infections (300/1768, 17.0%) and Triple infections (10/1768, 0.6%) were found only among the patients with HIV. The malignancies among HIV-infected patients included adenocarcinomas (0.22%), small cell carcinomas (0.2%), squamous cell carcinomas (0.1%), and diffuse large B-cell lymphoma (0.1%). HIV-infected patients exhibited a significantly lower incidence of neoplastic lesions (0.6% vs. 9.0%) than the HIV-uninfected patients (p < 0.001). CONCLUSIONS: There was a significant difference in the spectrum of lung diseases between HIV-infected and non-infected patients diagnosed by BALF cytopathology.

Comparison of next-generation sequencing with traditional methods for pathogen detection in cases of lower respiratory tract infection at a community hospital in Eastern China.

Lower respiratory tract infection (LRTI) is still a threat to human health. Metagenomics next-generation sequencing (NGS) provides an efficient and unbiased way to identify LRTI pathogens, and has been shown to have several advantages over traditional methods. However, its application is currently limited in low-resource settings. Our aim was to collect and analyze data on LRTI cases at a county-level community hospital in Eastern China over one year, in order to compare the efficiency of NGS and traditional methods including culture, nucleic acid amplification and antibody techniques. We performed NGS of bronchoalveolar lavage fluid (BALF) for pathogen identification in 71 patients with LRTI. We compared the detection rates, identified pathogens, and turnaround time of NGS with traditional methods. Pathogens were detected using traditional methods in 19 cases, and the results were compared with those obtained with the NGS technique in 60 cases. The pathogen detection rate of NGS (84.5%) was much higher than that of the traditional methods (26.8%). Moreover, with the traditional methods considered the gold standard, the consistency rate between NGS and traditional methods was 68.4%. For the 19 cases in which the traditional method was used, the main pathogens included invasive Aspergillus (5 cases), Pseudomonas aeruginosa (3 cases), Candida albicans (3 cases), and Staphylococcus aureus (2 cases). Among the 60 cases detected by NGS, the main pathogens included Mycobacterium (12 cases), Streptococcus pneumoniae (5 cases), Klebsiella pneumoniae (3 cases), P. aeruginosa (3 cases), Haemophilus influenzae (3 cases), and S. aureus (3 cases), Aspergillus (9 cases), Pneumocystis jiroveci (5 cases), C. albicans (3 cases), Human Papilloma Virus (9 cases), Epstein-Barr virus (8 cases), and parvovirus (6 cases). In addition, 2 cases of chlamydia and 1 case of mycoplasma infection were detected by NGS. The time taken to perform the NGS tests was significantly shorter than that taken with the traditional method. NGS analysis of bronchoalveolar lavage fluid, in combination with traditional pathogen detection methods, can improve the efficiency of pathogen detection. More attention should be paid to the regional epidemic characteristics of infectious pathogens in LRTI.