Lavage Samples Research Articles

Association between metabolome and neutrophil count in intrauterine fluid after insemination in the mare

Assessment of post-breeding endometritis is challenging in the mare. Correlation analysis of metabolite concentration in serum and intrauterine fluid with neutrophil count in intrauterine fluid after insemination was performed in 17 Spanish mares. After body condition score (BCS) was determined, a uterine edema score and intrauterine fluid diameter (IUF) were determined by ultrasonography at insemination and 8 hours afterwards, when a blood sample and low volume uterine lavage sample were collected. Neutrophils in the recovered fluid were counted using a Neubauer camera. In lavage supernatant fraction and in serum, metabolite determination by spectrophotometry was performed. Data were analyzed with PROC CORR (SAS) after log transformation of neutrophil count, oedema score and IUF. There was no association between edema or IUF at insemination and edema/IUF at lavage. The BCS tended to be positively associated with edema score (r=0.466, p=0.059) and IUF 8 hours after insemination (r=0.4562, p=0.066), but not at the time of insemination. Correlation analysis of BCS and neutrophil count did not reach significance (r=0.3928, p=0.118). Age tended to be positively associated with neutrophil count (r=0.4112, p=0.100) and was highly associated with urea concentrations in the IUF (r=6221, p=0.008). Edema at lavage was correlated with serum globulin concentrations (r=0.50911, P=0.03). The IUF was also correlated with serum albumin (r=0.5396, p=0.02),but no association with globulin was detected; instead IUF was correlated with serum glucose concentrations (r=0.4818, p=0.05). Neutrophil count was positively correlated with specific metabolites (cholesterol, urea, globulins), but not with glucose, triglycerides or albumin concentrations in the IUF. Neutrophil count was associated with serum globulin concentration (r=0.548, p=0.02), but no other correlations were found. The only metabolite that was correlated in serum and IUF was globulin (r=0.4777, p=0.05). Glucose and urea concentrations in the IUF were negatively associated with serum albumin (r= -0.6173, p=0.008 and r= - 0.4533, p=0.06 respectively). This is probably the first report showing an association between BCS and oedema score and intrauterine fluid after insemination, suggesting that in obese mares, clearance of physiological endometritis post-breeding is altered. The finding of the association between the oedema and neutrophil count with serum globulin concentrations, and the positive correlation of globulins in serum and uterine fluid suggest a systemic pro-inflammatory state that is present locally. Moreover, serum albumin was negatively correlated with glucose and urea in the uterine fluid. Overall, neutrophil count in low volume lavage obtained after breeding is consistent with the metabolome in uterine fluid and serum, and thus neutrophil count and serum globulin determination could be used as a diagnostic tool for post-breeding endometritis.

Pneumonia panel results and antibiotic prescribing in COVID-19 patients in 2020 versus 2022

Background: Antibiotics are frequently prescribed in patients with COVID-19 infections to treat secondary bacterial pneumonia. The pneumonia panel (PNP) is a molecular diagnostic tool that rapidly detects 33 bacterial and viral targets. The utility of this panel in COVID-19 patients and how it may direct antibiotic use is unknown. We sought to understand the utilization of PNP in patients with COVID-19 pneumonia over time by comparing clinical parameters, microbiologic results, and antibiotic use between May–December 2020 and January–July 2022. Methods: We implemented the PNP in May 2020 with antimicrobial stewardship guidance, provider education, and order restriction to critical care and infectious disease clinicians. From February–July 2021 prescribers received regular structured antimicrobial stewardship feedback regarding PNP results; from August 2021 to January 2022, no antimicrobial stewardship feedback was provided; from February to July 2022, intermittent feedback was provided. We compared PNP and culture results from sputum or bronchoalveolar lavage samples and antibiotic use and modification within 24 hours of PNP result from patients with confirmed COVID-19 pneumonia between May–December 2020 and January–July 2022. Clinical data and antibiotic use were abstracted through chart review. We excluded patients who died within 72 hours of PNP, those who had concurrent nonpulmonary infections, and those whose COVID-19 test was >30 days prior. Results: We included 114 patients in 2020 and 71 patients in 2022. The overall median age was 61 years, 71% were male, and 66% were mechanically ventilated without statistical differences between the cohorts, including their comorbidities. Acute or worsening hypoxia remained the predominant indication for PNP (77% in 2020 vs 75% in 2022, NS). The median number of days between admission and PNP was 4 (IQR, 1–8) in 2020 versus 3 (IQR, 1–7), and the difference was not significant. PNP and culture results in Table 1 show that Staphylococcus aureus and Hemophilus influenzae were the pathogens most commonly identified. Table 2 describes empiric prescribing and modifications for commonly prescribed antibiotics. Prescribers used empiric cefepime and ceftriaxone more in 2020 and vancomycin more in the 2022 group; however, these were not statistically significant. Cefepime de-escalation was more common in 2022 (53% vs 28%; P = .03). Antibiotic modifications within 24 hours of PNP remained similar in 2020 vs 2022. Although vancomycin cessation was more common in 2020 (78%) versus 2022 (57%), the difference was not statistically significant. Conclusions: With ASP guidance, PNP may be a useful tool to stop or target antibiotics for secondary bacterial pneumonia in COVID-19 pneumonia. Early vancomycin cessation (prior to culture results) may be an enduring consequence of PNP implementation.Disclosures: None

Respiratory toxicities after concurrent chemoradiotherapy and subsequent consolidation immune checkpoint inhibitors in locally advanced non-small cell lung cancer: Infectious and non-infectious subtypes by pathogen NGS testing.

e20557 Background: Consolidation immune checkpoint inhibitors (ICIs) following concurrent chemoradiotherapy (CCRT) have significantly improved survival in locally advanced non-small cell lung cancer (LA-NSCLC), while the associated respiratory toxicities were of particular concern. We aimed to investigate the > = G2 respiratory toxicities, the characteristics of concomitant infection, and provide detailed information for clinical management. Methods: We reviewed 77 LA-NSCLC patients treated with CCRT and consolidation ICIs. The details of > = G2 respiratory toxicities and concomitant infection were analyzed. Potential risk factors for > = G2 respiratory toxicities were explored by univariable and multivariable analysis. Respiratory toxicities were collected from the start of consolidation ICIs till 6 months after the last infusion of ICIs. Concomitant infection was defined as positive findings from next-generation sequencing (NGS) of bronchoalveolar lavage sampling. Results: Patients were treated with definitive radiotherapy at a median dose of 60 Gy (IQR, 60-64), concurrently with weekly docetaxel and platinum and followed by single-agent ICIs. The median duration of consolidation ICIs was 5.4 months (IQR, 2.3-8.2). Twenty-six patients (33.8%) experienced > = G2 respiratory toxicities by a median of 4.8 months (IQR 1.5-6.6) after initiation of consolidation ICIs. Pneumonitis (14, 18.2%) and tracheobronchitis (13, 16.9%) were the most common > = G2 respiratory disorders. Location of primary tumor (p = 0.027) and the proportion of CD3+ lymphocyte before consolidation ICIs (p = 0.044) were independent risk factors for the occurrence of > = G2 respiratory toxicities. Eleven patients (14.3%) had > = G2 respiratory toxicities and concomitant infection. Identified pathogens included bacteria (n = 9), Aspergillus (n = 5), Pneumocystis (n = 1) and Leptosvirus (n = 1). Concomitant infection was more commonly detected in patients with tracheobronchitis compared with those with pneumonitis ( p= 0.002), was associated with a higher grade of toxicity ( p= 0.050) and relatively poorer outcome after management ( p= 0.002). Conclusions: Respiratory toxicities after CCRT and subsequent consolidation ICIs in LA-NSCLC mainly included pneumonitis and tracheobronchitis, and could be separated into infectious and non-infectious subtypes. Bacteria and Aspergillus were the predominant pathogens. Our findings highlight the importance of the bronchoscopy and pathogen NGS testing to detect airway disease and specific infection. Long-term systemic steroids might adversely affect respiratory toxicities with concomitant opportunistic infection, which should be taken into account at the onset of management.

Quantification of Organophosphorus Insecticide Removed by Gastric Lavage in Acutely Poisoned Patients: An Observational Study.

The effectiveness of gastric lavage in organophosphorus (OP) poisoning has not been established. We assessed the ability of gastric lavage to remove OP insecticides as a preliminary stage in assessing effectiveness. Organophosphorus poisoning patients presenting within 6 hours were included, irrespective of prior gastric lavage. A nasogastric tube was placed and gastric contents aspirated, followed by at least three cycles of gastric lavage with 200 mL of water. Samples from the initial aspirate and the first three lavage cycles were sent for identification and quantification of the OP compounds. Patients were monitored for complications of gastric lavage. Around 42 patients underwent gastric lavage. Eight (19.0%) patients were excluded from the study because of a lack of analytical standards for ingested compounds. Insecticides were detectable in the lavage samples of 24 of 34 (70.6%) patients. Lipophilic OP compounds were detected in 23 of 24 patients, while no hydrophilic OP compounds could be detected in six patients with reported ingestion of hydrophilic compounds. For chlorpyrifos poisoning (n = 10), only 0.65 mg (SD 1.2) of the estimated ingested amount (n = 5) of 8,600 mg (SD 3,200) was recovered by gastric lavage. The mean proportion of the compound removed by initial gastric aspirate was 79.4% and subsequent three cycles removed 11.5, 6.6, and 2.7%. Lipophilic OP insecticides could be quantified in the stomach contents of OP poisoning patients with the first aspiration or lavage being most effective. The amount removed was very low; hence, routine use of gastric lavage for OP poisoning patients arriving within 6 hours is unlikely to be beneficial. Mathansingh AJ, Jose A, Fleming JJ, Abhilash KPP, Chandiraseharan VK, Lenin A, et al. Quantification of Organophosphorus Insecticide Removed by Gastric Lavage in Acutely Poisoned Patients: An Observational Study. Indian J Crit Care Med 2023;27(6):397-402.

Chronic ethanol exposure impairs alveolar leukocyte infiltration during pneumococcal pneumonia, leading to an increased bacterial burden despite increased CXCL1 and nitric oxide levels.

Ethanol abuse is a risk factor for the development of pneumonia caused by Streptococcus pneumoniae, a critical pathogen for public health. The aim of this article was to investigate the inflammatory mechanisms involved in pneumococcal pneumonia that may be associated with chronic ethanol exposure. Male C57BL6/J-Unib mice were exposed to 20% (v/v) ethanol for twelve weeks and intranasally infected with 5x104 CFU of S. pneumoniae. Twenty-four hours after infection, lungs, bronchoalveolar lavage and blood samples were obtained to assess the consequences of chronic ethanol exposure during infection. Alcohol-fed mice showed increased production of nitric oxide and CXCL1 in alveoli and plasma during pneumococcal pneumonia. Beside this, ethanol-treated mice exhibited a decrease in leukocyte infiltration into the alveoli and reduced frequency of severe lung inflammation, which was associated with an increase in bacterial load. Curiously, no changes were observed in survival after infection. Taken together, these results demonstrate that chronic ethanol exposure alters the inflammatory response during S. pneumoniae lung infection in mice with a reduction in the inflammatory infiltrate even in the presence of higher levels of the chemoattractant CXCL1.

Application of metagenomic next-generation sequencing in the diagnosis and resistome analysis of community-acquired pneumonia pathogens from bronchoalveolar lavage samples.

To perform a prospective diagnostic study exploring the clinical utility of metagenomic next-generation sequencing (mNGS) in diagnosing community-acquired pneumonia (CAP), and revealing resistome differences in bronchoalveolar lavage fluid (BALF) from CAP patients with varying severity of admission base on Pneumonia Patient Outcomes Research Team (PORT) risk classes. We compared the diagnostic performances of mNGS and conventional testing for the detection of pathogens in BALF from 59 CAP patients, and performed resistome differences analysis of metagenomic data from 59 BALF samples, namely, 25 from CAP patients with PORT score I (I group), 14 from CAP patients with PORT score II (II group), 12 from CAP patients with PORT score III (III group), and 8 from CAP patients with PORT score IV (IV group). The diagnostic sensitivities of mNGS and conventional testing for the detection of pathogens in BALF in patients with CAP were 96.6% (57/59) and 30.5% (18/59), respectively. There was a significant difference in the overall relative abundance of resistance genes between the four groups (P=0.014). The results of principal coordinate analysis based on Bray-Curtis dissimilarities showed that there were significant differences in the composition of resistance genes among the I, II, III, and IV groups (P=0.007). A large number of antibiotic resistance genes, such as those affiliated with multidrug, tetracycline, aminoglycoside, and fosfomycin resistance, were enriched in the IV group. In conclusion, mNGS has a high diagnostic value in CAP. There were significant differences present in microbiota resistance to antibiotics in BALF from CAP patients in different PORT risk classes, which should attract enough attention.

Current Utilization of Electron Microscopy in the Pediatric Pathology Setting: A Survey by the SPP Practice Committee.

Electron microscopy (EM), once an important component in diagnosing pediatric diseases, has experienced a decline in its use. To assess the impact of this, pediatric pathology practices were surveyed regarding EM services. The Society of Pediatric Pathology Practice Committee surveyed 113 society members from 74 hospitals. Settings included 36 academic tertiary, 32 free-standing children's, and 6 community hospitals. Over 60% maintained in-house EM services and had more than 2 pathologists interpreting EM while reporting a shortage of EM technologists. Freestanding children's hospitals had the most specimens (100-200 per year) and more diverse specimen types. Hospitals with fewer than 50 yearly specimens often used reference laboratories. Seventeen had terminated all in-house EM services. Challenges included decreasing caseloads due to alternative diagnostic methods, high operating costs, and shortages of EM technologists and EM-proficient pathologists. Kidney, liver, cilia, heart, and muscle biopsies most often required EM. Lung/bronchoalveolar lavage, tumor, skin, gastrointestinal, nerve, platelet, and autopsy samples less commonly needed EM. The survey revealed challenges in maintaining EM services but demonstrated its sustained value in pediatric pathology. Pediatric pathologists may need to address the centralization of services and training to preserve EM diagnostic proficiency among pathologists who perform ultrastructural interpretations.

Investigating Viral Involvement in Immunocompromised Patients Using Comprehensive Infectious Disease Testing Including FilmArray Respiratory Panel 2.1 on Bronchoscopy: A Retrospective Study.

Introduction Reports are rare on the usefulness of the FilmArray Respiratory Panel 2.1 (FARP) using lower respiratory tract specimens. This retrospective study assessed its use, as part of a comprehensive infectious disease panel, to detect the viral causes of pneumonia using bronchoalveolar lavage samples from immunosuppressed patients. Methods This study included immunocompromised patients who underwent bronchoalveolar lavage or bronchial washing by bronchoscopy between April 1, 2021, and April 30, 2022. The collected samples were submitted for comprehensive testing, including FARP test; reverse transcription polymerase chain reaction (RT-PCR) for cytomegalovirus, varicella-zoster virus DNA, and herpes simplex virus; PCR for Pneumocystis jirovecii DNA; antigen testing for Aspergillus and Cryptococcus neoformans; and loop-mediated isothermal amplification method for Legionella. Results Out of 23 patients, 16 (70%) showed bilateral infiltrative shadows on computed tomography and three (13%) were intubated. The most common causes of immunosuppression were anticancer drug use (n=12, 52%) and hematologic tumors (n=11, 48%). Only two (9%) patients tested positive for severe acute respiratory syndrome coronavirus 2 and adenovirus by FARP. Four patients (17%) tested positive for cytomegalovirus by RT-PCR, but no inclusion bodies were identified cytologically. Nine (39%) patients tested positive for Pneumocystis jirovecii by PCR, but cytology confirmed the organism in only one case. Conclusions Comprehensive infectious disease testing, performed using bronchoalveolar lavage samples collected from lung lesions in immunosuppressed patients, showed low positive detection by FARP. The viruses currently detectable by FARP may be less involved in viral pneumonia diagnosed in immunocompromised patients.

Identification of Chlamydia trachomatis genotypes in newborns with respiratory distress

IntroductionOne hundred thirty million Chlamydia trachomatis infections are reported worldwide each year. Nineteen serotypes of this pathogen can cause infection in pregnant women and neonates. The distribution of these genotypes in newborns with respiratory infections in Mexico is unknown. Material and methodsWe tested 1062 bronchial lavage samples from neonates with respiratory distress syndrome for Chlamydia infection. The diagnosis of Chlamydia was made by plasmid detection with an in-house PCR assay, and genotypes were identified using a PCR-RFLP assay for the ompA gene. ResultsThe genotyping of 40 strains identified 14 as I/Ia (35%), 13 as E (32.5%), 7 as D (17.5%), 5 as F (12.5%), and 1 as L2 (2.5%). The relative risk analysis showed that genotype D was associated with neonatal sepsis (RR, 5.83; 95% confidence interval [CI], 1.51–25.985; P < .02), while the I/Ia genotype was significantly associated with chorioamnionitis in the mother (2.8; 95% CI, 1.4−5.5; P < .05). ConclusionsAlthough C. trachomatis genotypes I/Ia and E of were the strains involved most frequently in respiratory infections in Mexican neonates, 80% of patients with genotype F developed respiratory disease. In contrast, genotype D was associated with neonatal sepsis, and genotype I/Ia with chorioamnionitis.

Lung and Gut Microbiome in COPD.

Chronic obstructive pulmonary disease (COPD) is one of the leading causes of death worldwide. The association between lung and gut microbiomes in the pathogenesis of COPD has been recently uncovered. The goal of this study was to discuss the role of the lung and gut microbiomes in COPD pathophysiology. A systematic search of the PubMed database for relevant articles submitted up to June 2022 was performed. We examined the association between the lung and gut microbiome dysbiosis, reflected in bronchoalveolar lavage (BAL), lung tissue, sputum, and feces samples, and the pathogenesis and progression of COPD. It is evident that the lung and gut microbiomes affect each other and both play a vital role in the pathogenesis of COPD. However, more research needs to be carried out to find the exact associations between microbiome diversity and COPD pathophysiology and exacerbation genesis. Another field that research should focus on is the impact of treatment interventions targeting the human microbiome in preventing COPD genesis and progression.

Expedited diagnosis of pediatric tuberculosis using Truenat MTB-Rif Dx and GeneXpert MTB/RIF

Rapid, cost-effective, and sensitive diagnostic assays are essential for global tuberculosis (TB) control, especially in high TB burden, resource-limited settings. The current study was designed to evaluate diagnostic accuracy of Truenat MTB-Rif Dx (MolBio) in children less than 18 years of age, with symptoms suggestive of TB. Gastric aspirate, induced sputum, and broncho-alveolar lavage samples were subjected simultaneously to AFB-smear, GeneXpert MTB/RIF, liquid culture (MGIT-960) and Truenat MTB-Rif Dx. The index-test results were evaluated against microbiological reference standards (MRS). Truenat MTB-Rif Dx had a sensitivity of 57.1%, specificity of 92% against MRS. The sensitivity and specificity of the Truenat MTB-RIF Dx compared with liquid culture was 58.7% and 87.5% while GeneXpert MTB/RIF was 56% and 91.4%. The performance of both GeneXpert MTB/RIF and Truenat MTB-Rif Dx are comparable. Result of our study demonstrates that Truenat MTB-Rif can aid in early and efficient diagnosis of TB in children.

Tumor DNA in Peritoneal Lavage as a Novel Biomarker for Predicting Peritoneal Recurrence in Patients With Gastric Cancer.

Peritoneal lavage cytology positivity (CY1) has been identified as a prognostic factor for distant metastases that is equivalent to peritoneal dissemination in Japan. Peritoneal lavage cytology is usually diagnosed by microscopic findings; a diagnostic procedure using a liquid biopsy (LB) technique has not yet been established. We evaluated the feasibility of a LB approach using peritoneal lavage samples from 15 patients with gastric cancer. Samples were collected from both the Douglas pouch and the left subdiaphragmatic area, and cell-free DNA was extracted for analysis of TP53 mutations using droplet digital polymerase chain reaction. All 10 patients with CY1 had positive cytology for the left subdiaphragmatic specimen. However, only six out of the 10 patients had positive cytology for their Douglas pouch specimens, and these six patients had peritoneal tumor DNA (ptDNA) in these specimens. In all five patients with CY0, ptDNA was not detected. The overall survival was significantly shorter in the ptDNA-positive group than in the ptDNA-negative group. The survival of the group with a high amount of DNA from free intraperitoneal cells (ficDNA) was significantly worse than that of those with a low amount. In contrast, the group with a high amount of DNA from peritoneal cell-free DNA (pcfDNA) had significantly better survival than the group with a low amount. LB cytology showed equivalent utility to that of conventional microscopic examinations regarding its diagnostic ability. Therefore ptDNA, pcfDNA and ifcDNA are expected to be useful as prognostic factors.

Pseudo-outbreak of Mycobacterium lentiflavum at a general hospital in Japan.

Mycobacterium lentiflavum is a slow-growing nontuberculous mycobacterium that is widely distributed in soil and water systems, but it is sometimes pathogenic to humans. Although cases of M. lentiflavum infections are rare, 22 isolates of M. lentiflavum were identified at a single hospital in Japan. We suspected a nosocomial outbreak; thus, we conducted transmission pattern and genotype analyses. Cases of M. lentiflavum isolated at Kushiro City General Hospital in Japan between May 2020 and April 2021 were analyzed. The patient samples and environmental culture specimens underwent whole-genome sequencing (WGS). Additionally, we retrospectively collected clinical data from patient medical records. Altogether, 22 isolates of M. lentiflavum were identified from sputum and bronchoalveolar lavage samples. Clinically, the instances with M. lentiflavum isolates were considered contaminants. In the WGS analysis, 19 specimens, including 18 patient samples and 1 environmental culture from the hospital's faucet, showed genetic similarity. The frequency of M. lentiflavum isolation decreased after we prohibited the use of taps where M. lentiflavum was isolated. WGS analysis identified that the cause of M. lentiflavum pseudo-outbreak was the water used for patient examinations, including bronchoscopy.

Identificación de genotipos de Chlamydia trachomatis en neonatos con distrés respiratorio

IntroductionOne hundred thirty million Chlamydia trachomatis infections are reported worldwide each year. Nineteen serotypes of this pathogen can cause infection in pregnant women and neonates. The distribution of these genotypes in newborns with respiratory infections in Mexico is unknown. Material and methodsWe tested 1062 bronchial lavage samples from neonates with respiratory distress syndrome for Chlamydia infection. The diagnosis of Chlamydia was made by plasmid detection with an in-house PCR assay, and genotypes were identified using a PCR-RFLP assay for the ompA gene. ResultsThe genotyping of 40 strains identified 14 as I/Ia (35%), 13 as E (32.5%), 7 as D (17.5%), 5 as F (12.5%), and 1 as L2 (2.5%). The relative risk analysis showed that genotype D was associated with neonatal sepsis (RR, 5.83; 95% confidence interval [CI], 1.51-25.985; P<.02), while the I/Ia genotype was significantly associated with chorioamnionitis in the mother (2.8; 95% CI, 1.4–5.5; P<.05). ConclusionsAlthough Chlamydia trachomatis genotypes I/Ia and E of were the strains involved most frequently in respiratory infections in Mexican neonates, 80% of patients with genotype F developed respiratory disease. In contrast, genotype D was associated with neonatal sepsis, and genotype I/Ia with chorioamnionitis.

Evaluation of BioFire® FilmArray® Pneumonia Panel in Bronchoalveolar Lavage Samples From Immunocompromised Patients With Suspected Pneumonia.

Objectives Immunocompromised patients, specifically those with solid organ transplants or cancer on chemotherapy, are at particularly high risk of severe pneumonia and opportunistic infections. In select patients, bronchoalveolar lavage (BAL) is performed to provide high-quality samples for analysis. We compare BioFire® FilmArray® Pneumonia Panel (BioFire Diagnostics,Salt Lake City, Utah, United States), a multiplex polymerase chain reaction (PCR) assay, with standard of care diagnostics in BAL samples from immunocompromised patients to identify opportunities for this test to affect clinical decision making. Methods Patients hospitalized with pneumonia based on clinical and radiographic findings who underwent evaluation with bronchoscopybetween May 2019 to January 2020 were reviewed. Among those patients undergoing bronchoscopy, those who were immunocompromised were selected for inclusion in the study. BAL specimens submitted to the microbiology laboratory were chosen based onas part of the internal validation of the panel in comparison with sputum culture at our hospitals.We compared the outcomes of the multiplex PCR assay with traditional culture methods and evaluated the role of PCR assay in de-escalating antimicrobial therapy. Results Twenty-four patients were identified for testing with the multiplex PCR assay. Of the 24 patients, 16 were immunocompromised, all with solid or hematological malignancy or a history of organ transplant. Seventeen individual BAL samples from the 16 patients were reviewed. BAL culture results and the multiplex PCR assay were in agreement in 13 samples (76.5%). In four cases, the multiplex PCR assay identified a possible causative pathogen not detected by standard workup. The median time to de-escalation of antimicrobials was three days (interquartile range (IQR) 2-4) from the day of collection of the BAL samples. Conclusions Studies have established the additive role of multiplex PCR testing in addition to traditional diagnostic tools like sputum culture in diagnosing the etiology of pneumonia. Limited data exist specifically looking at immunocompromised patients, in whom a timely and accurate diagnosis is particularly important. There is a potential benefit for performing multiplex PCR assays as an additive diagnostic tool in BAL samples for these patients.

Rapid Diagnosis of Recurrent Paucibacillary Tuberculosis.

The rapid diagnosis of tuberculosis recurrence can be challenging due to persistently positive detection of Mycobacterium tuberculosis-specific DNA from sputum and bronchopulmonary samples in the absence of active disease. We compared the diagnostic accuracy of the detection of M. tuberculosis-specific DNA by either Xpert (January 2010-June 2018) or Xpert Ultra (July 2018-June 2020) and M. tuberculosis-specific ELISPOT in bronchoalveolar lavage (BAL) samples with M. tuberculosis culture results from sputum or bronchopulmonary samples in patients with suspected recurrence of pulmonary tuberculosis. Among 44 individuals with previous tuberculosis and a presumptive diagnosis of recurrent pulmonary tuberculosis, 4/44 (9.1%) were diagnosed with recurrent tuberculosis by culture. DNA of M. tuberculosis was detected by Xpert in BAL fluid in 1/4 (25%) individuals with recurrent tuberculosis and in 2/40 (5%) cases with past tuberculosis without recurrence, while BAL-ELISPOT with a cut-off of >4,000 early secretory antigenic target-6-specific or culture filtrate protein-10-specific interferon-γ-producing lymphocytes per 1 million BAL-lymphocytes was positive in 4/4 (100%) individuals with recurrent tuberculosis and in 2/40 (5%) cases of past tuberculosis without recurrence. M. tuberculosis-specific BAL-ELISPOT is more accurate than BAL-Xpert for the diagnosis of paucibacillary tuberculosis recurrence.

Bacterial Species and Inflammatory Cell Variability in Respiratory Tracts of Patients with Chronic Obstructive Pulmonary Disease Exacerbation: A Multicentric Study.

Acute exacerbation of chronic obstructive pulmonary disease (AECOPD) has profound effects on disease progression and patients' quality of life. Emerging evidence suggests an association between alterations in the respiratory microbiome flora species and airway inflammation in patients with AECOPD. The present study aimed to describe the inflammatory cells and bacterial microbiome distributions in respiratory tract in Egyptian patients with AECOPD. The present cross-sectional study included 208 patients with AECOPD. Sputum and broncho-alveolar lavage samples from the studied patients were submitted to microbial cultures using appropriate media. Total and differential leukocytic counts and were done via automated cell counter. The present study included 208 AECOPD patients. They comprised 167 males (80.3%) and 41 females (19.7%) with an age of 57.9 ± 4.9 years. AECOPD was categorized as mild, moderate and severe in 30.8%, 43.3% and 26%, respectively. Sputum samples had significantly higher TLC, neutrophil percent and eosinophil percent when compared with BAL samples. In contrast, lymphocyte percent was significantly higher in BAL samples. Sputum specimens had significantly lower frequency of positive growths (70.2% versus 86.5%, p = 0.001). Among the identified organisms, sputum specimens had significantly lower frequency of Strept. pneumoniae (14.4% versus 30.3%, p = 0.001), Klebsiella pneumoniae (19.7% versus 31.7%, p = 0.024), Haemophilus influenzae (12.5% versus 26.9%, p = 0.011), Pseudomonas aeruginosa (2.9% versus 10%, p = 0.019) and Acinetobacter spp. (1.9% versus 7.2%, p = 0.012) growths when compared with BAL samples. The present study could identify a distinctive pattern of inflammatory cell distribution in sputum and BAL samples of AECOPD patients. The most commonly isolated organisms were Klebsiella pneumoniae and Strept. pneumoniae.

Gaceta Médica de México in SARS-CoV-2 pandemic times

December 2019 ended with the identification and development of an atypical pneumonia epidemic caused by an unknown pathogen in Wuhan, China. At the dawn of 2020, at first week, on January 7, the causative agent was isolated in bronchial lavage sam-ples from a critically ill patient. Five days later, on January 12, the Chinese health authorities made the whole genome sequence of a new strain of the Coro-naviridae family public: SARS-CoV-2, among whose relevant characteristics a high mortality rate stood out, as a consequence of the development of acute respi-ratory failure, and easy transmission.

Integrated Analysis of Tracheobronchial Fluid from Before and After Cardiopulmonary Bypass Reveals Activation of the Integrated Stress Response and Altered Pulmonary Microvascular Permeability.

Objective: We aim to comprehensively describe the transcriptional activity and signaling of pulmonary parenchymal and immune cells before and after cardiopulmonary bypass (CPB) by using a multi-omic approach coupled with functional cellular assays. We hypothesize that key signaling pathways from specific cells within the lung alter pulmonary endothelial cell function resulting in worsening or improving disease. Methods: We collected serial tracheobronchial lavage samples from intubated patients less than 2-years-old undergoing surgery with CPB. Samples were immediately processed for single cell RNA sequencing (10x Genomics). Cell clustering, cell-type annotation, and visualization were performed, and differentially expressed genes (DEG) between serial samples were identified. Metabolomic and proteomic analyses were performed on the supernatant using mass spectrometry and a multiplex assay (SomaScan) respectively. Functional assays were done using electric cell-substrate impedance sensing to measure resistance across human pulmonary microvascular endothelial cells (HPMECs). Results: Analysis of eight patients showed a heterogeneous mixture of pulmonary parenchymal and immune cells. Cell clustering demonstrated time-dependent changes in the transcriptomic signature indicating altered cellular phenotypes after CPB. DEG analysis was represented by genes involved in host defense, innate immunity, and the mitochondrial respiratory transport chain. Ingenuity pathway analysis showed upregulation of the integrated stress response across all cell types after CPB. Metabolomic analysis demonstrated upregulation of ascorbate and aldarate metabolism. Unbiased proteomic analysis revealed upregulation of proteins involved in cytokine and chemokine pathways. Post-CPB patient supernatant improved HMPEC barrier function, suggesting a protective cellular response to CPB. Conclusion: Children who undergo CPB for cardiac surgery have distinct cell populations, transcriptional activity, and metabolism that change over time. The response to ischemia-reperfusion injury in the lower airway of children appears to be protective, with the need to identify potential targets through future investigations.

Lung microhaemorrhage drives oxidative/inflammatory damage in α1-antitrypsin deficiency.

Animal models using intratracheal instillation show that elastase, unopposed by α1-antitrypsin (AAT), causes alveolar damage and haemorrhage associated with emphysematous changes. The aim of the present study was to characterise any relationship between alveolar haemorrhage and human AAT deficiency (AATD) using bronchoalveolar lavage (BAL) and lung explant samples from AATD subjects. BAL samples (17 patients, 15 controls) were evaluated for free haem (iron protoporphyrin IX) and total iron concentrations. Alveolar macrophage activation patterns were assessed using RNA sequencing and validated in vitro using haem-stimulated, monocyte-derived macrophages. Lung explants (seven patients, four controls) were assessed for iron sequestration protein expression patterns using Prussian blue stain and ferritin immunohistochemistry, as well as ferritin iron imaging and elemental analysis by transmission electron microscopy. Tissue oxidative damage was assessed using 8-hydroxy-2'-deoxyguanosine immunohistochemistry. BAL collected from AATD patients showed significantly elevated free haem and total iron concentrations. Alveolar and interstitial macrophages in AATD explants showed elevated iron and ferritin accumulation in large lysosomes packed by iron oxide cores with degraded ferritin protein cages. BAL macrophage RNA sequencing showed innate pro-inflammatory activation, replicated in vitro by haemin exposure, which also triggered reactive oxygen species generation. AATD explants showed massive oxidative DNA damage in both lung epithelial cells and macrophages. BAL and tissue markers of alveolar haemorrhage, together with molecular and cellular evidence of macrophage innate pro-inflammatory activation and oxidative damage, are consistent with free haem stimulation. Overall, this initial study provides evidence for a pathogenetic role of elastase-induced alveolar haemorrhage in AATD emphysema.