Distribution characteristics of human herpes viruses in the lower respiratory tract and their impact on 30-day mortality in community-acquired pneumonia patients.

Human herpes viruses (HHVs) are commonly detected in community-acquired pneumonia (CAP) patients, particularly those with complex complications, attracting increased attention from clinical practitioners. However, the significance of detecting HHVs in bronchoalveolar lavage fluid (BALF) with CAP patients is still unclear. This study retrospectively analyzed BALF samples from 64 CAP patients at the Kunming Third People's Hospital between August 2021 and December 2023. Metagenomic next generation sequencing (mNGS) was conducted on BALF samples during CAP onset. Multivariate Cox regression models were used to identify independent risk factors for 30-day all-cause mortality in CAP. HHVs were found in 84.4% of CAP patients, which were the most common pathogens (45.1%), followed by bacteria (30.2%) and fungi (11.5%). Bacterial-viral co-infections were most common, occurring in 39 patients. Notably, there was no significant difference in HHV presence between severe and non-severe CAP patients (EBV: P = 0.431, CMV: P = 0.825), except for HHV-7 (P = 0.025). In addition, there was no significant difference in the 30-day mortality between HHV positive and HHV negative groups (P = 0.470), as well as between the HHV-7 positive and HHV-7 negative groups (P = 0.910). However, neither HHVs nor HHV-7 was independent risk factors for 30-day mortality in CAP patients (HHVs: HR 1.171, P = 0.888; HHV-7: HR 1.947, P = 0.382). In summary, among the prevalent presence of multiple HHVs, EBV and CMV were the most prevalent in CAP patients. Patients with sCAP were more susceptible to HHV-7 than those with non-sCAP. These results provide valuable insights for clinicians in guiding appropriate interventions for CAP treatment.

Fungi identified via next-generation sequencing in bronchoalveolar lavage fluid among patients with COVID-19: a retrospective study.

BACKGROUND: The epidemiology of fungi identified via next-generation sequencing in bronchoalveolar lavage fluid among patients with COVID-19 is unknown. METHODS: De-identified information, including age, SARS-CoV-2 reads and fungi from bronchoalveolar lavage fluid, were used to analysis. RESULTS: A total of 960 patients with COVID-19 were included. Gender was unknown in 38 patients, and 648 (70.3%) of the rest patients were male. For 876 patients with information on age, their mean ± standard age was 63.4 ± 21.3 years, with the minimum being 0.2 years and the maximum being 101 years. For all the patients, their median [interquartile range] SARS-CoV-2 reads were 26,038 [4421.5, 44,641.5]. The Aspergilli were identified in 159 (16.6%) patients, with Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger in 103 (10.7%), 81 (8.4%) and 17 (1.8%), respectively. The Mucoraceae were identified in 14 (1.5%) patients. Pneumocystis jirovecii was identified in 65 (6.8%) patients, among whom 12 (18.5%) patients also had Aspergilli. The Cryptococcaceae and the Dematiaceae were also identified in some patients, including Cryptococcus in 11 (1.1%) patients. CONCLUSIONS: In bronchoalveolar lavage fluid among patients with COVID-19, the Aspergilli were very commonly identified, as were the Mucoraceae, Pneumocystis jirovecii and Cryptococcus via next-generation sequencing.

Comparative analysis of metagenomic and targeted next-generation sequencing for pathogens diagnosis in bronchoalveolar lavage fluid specimens.

Background: Although the emerging NGS-based assays, metagenomic next-generation sequencing (mNGS) and targeted next-generation sequencing (tNGS), have been extensively utilized for the identification of pathogens in pulmonary infections, there have been limited studies systematically evaluating differences in the efficacy of mNGS and multiplex PCR-based tNGS in bronchoalveolar lavage fluid (BALF) specimens. Methods: In this study, 85 suspected infectious BALF specimens were collected. Parallel mNGS and tNGS workflows to each sample were performed; then, we comparatively compared their consistency in detecting pathogens. The differential results for clinically key pathogens were confirmed using PCR. Results: The microbial detection rates of BALF specimens by the mNGS and tNGS workflows were 95.18% (79/83) and 92.77% (77/83), respectively, with no significant difference. mNGS identified 55 different microorganisms, whereas tNGS detected 49 pathogens. The comparative analysis of mNGS and tNGS revealed that 86.75% (72/83) of the specimens were complete or partial concordance. Particularly, mNGS and tNGS differed significantly in detection rates for some of the human herpesviruses only, including Human gammaherpesvirus 4 (P<0.001), Human betaherpesvirus 7 (P<0.001), Human betaherpesvirus 5 (P<0.05) and Human betaherpesvirus 6 (P<0.01), in which tNGS always had higher detection rates. Orthogonal testing of clinically critical pathogens showed a total coincidence rate of 50% for mNGS and PCR, as well as for tNGS and PCR. Conclusions: Overall, the performance of mNGS and multiplex PCR-based tNGS assays was similar for bacteria and fungi, and tNGS may be superior to mNGS for the detection of DNA viruses. No significant differences were seen between the two NGS assays compared to PCR.

SARS-CoV-2 not Detected in the Nasopharyngeal Sample but in Bronchoalveolar Lavage Fluid.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection typically relies on reverse transcription-polymerase chain reaction (RT-PCR) technology. However, there is a certain rate of missed detection of SARS-CoV-2 in nasopharyngeal samples, particularly among immunosuppressed individuals. METHODS: In this case, SARS-CoV-2 was detected in nasopharyngeal swabs and bronchoalveolar lavage fluid (BALF) using RT-PCR. Pulmonary imaging was performed using computed tomography (CT). Patient clinical data were retrieved from the Laboratory Information System (LIS). RESULTS: SARS-CoV-2 was negative in two nasopharyngeal tests of the patient, but was finally detected in BALF, confirming that the lung lesions were infected by SARS-CoV-2. CONCLUSIONS: In the post-epidemic era, it is necessary to use BALF to identify SARS-CoV-2 infection in cases where other factors have been ruled out in immunosuppressed individuals with pulmonary infections, especially when the nasopharyngeal test yields a negative result.

Metagenomic next-generation sequencing targeted and metagenomic next-generation sequencing for pulmonary infection in HIV-infected and non-HIV-infected individuals.

Background: When individuals infected with human immunodeficiency virus (HIV) experience pulmonary infections, they often exhibit severe symptoms and face a grim prognosis. Consequently, early, rapid, and accurate pathogen diagnosis is vital for informing effective treatment strategies. This study aimed to use metagenomic next-generation sequencing (mNGS) and targeted mNGS (tNGS) to elucidate the characteristics of pulmonary infections in HIV and non-HIV individuals. Methods: This study enrolled 90 patients with pulmonary infection at the Department of Infectious Diseases of The First Hospital of Jilin University from June 2022 to May 2023, and they were divided into HIV (n=46) and non-HIV (n=44) infection groups. Their bronchoalveolar lavage fluid (BALF) was collected for mNGS analysis to evaluate the differences in pulmonary infection pathogens, and tNGS detection was performed on BALF samples from 15 HIV-infected patients. Results: A total of 37 pathogens were identified in this study, including 21 bacteria, 5 fungi, 5 viruses, 5 mycobacteria, and 1 mycoplasma. The sensitivity of mNGS was 78.9% (71/90), which is significantly higher than that of conventional methods (CTM) (39/90, P=1.5E-8). The combination of mNGS with CTM can greatly enhance the sensitivity of pathogen detection. The prevalence of Pneumocystis jirovecii (82.6% vs. 9.1%), cytomegalovirus (CMV) (58.7% vs. 0%), and Epstein-Barr virus (EBV) (17.4% vs. 2.3%) was significantly higher in the HIV infection group than in the non-HIV infection group (P<0.05). Although no statistically significant difference was observed, the detection rate of Mycobacteria was higher in HIV-infected patients (17.4%) than in the non-HIV group (6.8%). Furthermore, the tNGS results of BALF from 15 HIV-infected patients were not entirely consistent with the mNGS results., and the concordance rate of tNGS for the detection of main pathogens reached 86.7% (13/15). Conclusion: Next-generation sequencing (NGS) can accurately detect pathogens in the BALF of patients with pulmonary infection. The sensitivity of tNGS is comparable to that of mNGS. Therefore, this technique should be promoted in the clinic for better patient outcomes.

Application of metagenomic next-generation sequencing and targeted metagenomic next-generation sequencing in diagnosing pulmonary infections in immunocompetent and immunocompromised patients.

Background: Metagenomic next-generation sequencing (mNGS) technology has been widely used to diagnose various infections. Based on the most common pathogen profiles, targeted mNGS (tNGS) using multiplex PCR has been developed to detect pathogens with predesigned primers in the panel, significantly improving sensitivity and reducing economic burden on patients. However, there are few studies on summarizing pathogen profiles of pulmonary infections in immunocompetent and immunocompromised patients in Jilin Province of China on large scale. Methods: From January 2021 to December 2023, bronchoalveolar lavage fluid (BALF) or sputum samples from 546 immunocompetent and immunocompromised patients with suspected community-acquired pneumonia were collected. Pathogen profiles in those patients on whom mNGS was performed were summarized. Additionally, we also evaluated the performance of tNGS in diagnosing pulmonary infections. Results: Combined with results of mNGS and culture, we found that the most common bacterial pathogens were Pseudomonas aeruginosa, Klebsiella pneumoniae, and Acinetobacter baumannii in both immunocompromised and immunocompetent patients with high detection rates of Staphylococcus aureus and Enterococcus faecium, respectively. For fungal pathogens, Pneumocystis jirovecii was commonly detected in patients, while fungal infections in immunocompetent patients were mainly caused by Candida albicans. Most of viral infections in patients were caused by Human betaherpesvirus 5 and Human gammaherpesvirus 4. It is worth noting that, compared with immunocompetent patients (34.9%, 76/218), more mixed infections were found in immunocompromised patients (37.8%, 14/37). Additionally, taking final comprehensive clinical diagnoses as reference standard, total coincidence rate of BALF tNGS (81.4%, 48/59) was much higher than that of BALF mNGS (40.0%, 112/280). Conclusions: Our findings supplemented and classified the pathogen profiles of pulmonary infections in immunocompetent and immunocompromised patients in Jilin Province of China. Most importantly, our findings can accelerate the development and design of tNGS specifically used for regional pulmonary infections.

Interferon-responsive neutrophils and macrophages extricate SARS-CoV-2 Omicron critical patients from the nasty fate of sepsis.

The SARS-CoV-2 Omicron variant is characterized by its high transmissibility, which has caused a worldwide epidemiological event. Yet, it turns ominous once the disease progression degenerates into severe pneumonia and sepsis, presenting a horrendous lethality. To elucidate the alveolar immune or inflammatory landscapes of Omicron critical-ill patients, we performed single-cell RNA-sequencing (scRNA-seq) of bronchoalveolar lavage fluid (BALF) from the patients with critical pneumonia caused by Omicron infection, and analyzed the correlation between the clinical severity scores and different immune cell subpopulations. In the BALF of Omicron critical patients, the alveolar violent myeloid inflammatory environment was determined. ISG15+ neutrophils and CXCL10+ macrophages, both expressed the interferon-stimulated genes (ISGs), were negatively correlated with clinical pulmonary infection score, while septic CST7+ neutrophils and inflammatory VCAN+ macrophages were positively correlated with sequential organ failure assessment. The percentages of ISG15+ neutrophils were associated with more protective alveolar epithelial cells, and may reshape CD4+ T cells to the exhaustive phenotype, thus preventing immune injuries. The CXCL10+ macrophages may promote plasmablast/plasma cell survival and activation as well as the production of specific antibodies. As compared to the previous BALF scRNA-seq data from SARS-CoV-2 wild-type/Alpha critical patients, the subsets of neutrophils and macrophages with pro-inflammatory and immunoregulatory features presented obvious distinctions, suggesting an immune disparity in Omicron variants. Overall, this study provides a BALF single-cell atlas of Omicron critical patients, and suggests that alveolar interferon-responsive neutrophils and macrophages may extricate SARS-CoV-2 Omicron critical patients from the nasty fate of sepsis.

Evaluation of STANDARD™ M10 SARS-CoV-2 from bronchoalveolar lavage samples.

Detection of SARS-CoV-2 in bronchoalveolar lavage (BAL) is considered as a promising alternative method to detect COVID-19 infection. STANDARD™ M10 SARS-CoV-2 assay on 150 negative and 50 positives BAL samples for SARS-CoV-2 showed 96 % sensitivity, 100 % specificity compared to Allplex™ SARS-CoV-2 assay and a 31.25 genomic copies/mL limit of detection.

Respiratory Syncytial Virus Infection in an Adult with Immunodeficiency.

BACKGROUND: Respiratory syncytial virus (RSV) is a single-stranded RNA virus that commonly causes symptoms of upper respiratory tract infections in humans, with a clear seasonal trend. However, in immunocompromised and elderly patients, RSV infections still result in high rates of hospitalization and even risk of death. METHODS: We report a case of RSV infection in an adult with immunodeficiency, which initially showed only mild symptoms of upper respiratory tract infection, which did not improve after receiving empirical anti-infective treatment, and the foci of infection in the lungs continued to expand, which led to the aggravation of the disease. The diagnosis of RSV infection was finally confirmed by electron bronchoscopy and pathogenetic examination of the bronchoalveolar lavage fluid. The patient was given intravenous ribavirin treatment for one week. After one week of intravenous ribavirin treatment, the patient's symptoms improved significantly. A repeat chest CT suggested that the lung lesions were smaller than before. In order to improve clinicians' awareness of this disease, we jointly conducted a literature analysis. RESULTS: The final diagnosis of RSV was made by analyzing the patient's history, symptoms, and signs and performing relevant examinations. CONCLUSIONS: For patients with poor results of empirical application of antibiotics, electronic bronchoscopy and pathogenetic examination should be carried out at an early stage to clarify the nature of the lesions and to avoid rapid deterioration of the condition leading to life-threatening conditions in the patients. More consideration should be given to the possibility of disease diagnosis to avoid misdiagnosis and underdiagnosis, and appropriate treatment should be given at an early stage.

Next-Generation Sequencing for Characterizing Respiratory Tract Virome and Improving Detection of Viral Pathogens in Children With Pneumonia.

BACKGROUND: Pneumonia is typically caused by a variety of pathogenic microorganisms. Traditional research often focuses on the infection of a few microorganisms, whereas metagenomic studies focus on the impact of the bacteriome and mycobiome on respiratory diseases. Reports on the virome characteristics of pediatric pneumonia remain relatively scarce. METHODS: We employed de novo assembly and combined homology- and feature-based methods to characterize the respiratory virome in whole-genome DNA sequencing samples from oropharynx (OP) swabs, nasopharynx (NP) swabs, and bronchoalveolar lavage fluids (BALF) of children with pneumonia. RESULTS: Significant differences were observed in the alpha and beta diversity indexes, as well as in the composition of the oropharyngeal virome, between pneumonia cases and controls. We identified 1137 viral operational taxonomic units (vOTUs) with significant differences, indicating a preference of pneumonia-reduced vOTUs for infecting Prevotella, Neisseria, and Veillonella, whereas pneumonia-enriched vOTUs included polyomavirus, human adenovirus, and phages targeting Staphylococcus, Streptococcus, Granulicatella, and Actinomyces. Comparative analysis revealed higher relative abundances and prevalence rates of pneumonia-enriched OP vOTUs in NP and BALF samples compared to pneumonia-reduced vOTUs. Additionally, virome analysis identified six pediatric patients with severe human adenovirus or polyomavirus infections, five of whom might have been undetected by targeted polymerase chain reaction (PCR)-based testing. CONCLUSIONS: This study offers insights into pediatric pneumonia respiratory viromes, highlighting frequent transmission of potentially pathogenic viruses and demonstrating virome analysis as a valuable adjunct for pathogen detection.

Distinct clinical characteristics of bocavirus and Mycoplasma pneumoniae infection in children plastic bronchitis.

BACKGROUND: This study investigated clinical and laboratory characteristics of human bocavirus type 1 (HBoV1)-plastic bronchiolitis (PB), Mycoplasma pneumoniae (MP)-associated plastic bronchitis (PB) and MP-NPB in children, highlighting inflammation, coagulation, and bronchoscopic needs. METHODS: Data on preschool children with PB during HBoV1 or MP infection were collected, comparing MP-PB to severe Mycoplasma pneumoniae pneumonia. RESULT: Compared with the MP-PB group, the HBoV1-PB group, with younger children, had significantly milder clinical symptoms but higher WBC counts (p = .028). The MP-PB group exhibited notably elevated Fibrinogen (p = .045) and d-dimer levels (p < .001). When contrasting the MP-PB with the MP-NPB group, children in MP-PB group still had higher levels of d-dimer and increased inflammatory indicators such as C-reactive protein, procalcitonin, lactate dehydrogenase, and interleukin-6, which were significantly elevated compared with the MP-NPB group. MP-PB showed a higher prevalence of plastic bronchial casts in lower lobes (p = .016) and a dominance of neutrophils in BALF cytology. Additionally, children in the MP-PB group tended to undergo a greater number of bronchoscopies. CONCLUSION: This study identifies key differences in plastic bronchitis in children due to HBoV1 and MP, highlighting HBoV1's milder inflammation in younger kids and MP's link to severe inflammatory and coagulation responses, guiding clinical diagnosis and treatment.

Comparison of the Results of BAL and ETA Culture in Intubated COVID-19 Patients.

BACKGROUND: The isolation of pathogens using bronchoalveolar lavage (BAL) culture or endotracheal aspirate (ETA) culture may enhance the treatment success for secondary pneumonia due to COVID-19, thereby reducing the risk of morbidity and mortality. AIM: This study aimed to retrospectively analyze the results of BAL and ETA cultures in intubated COVID-19 patients and to determine whether BAL has an advantage over ETA. METHODS: We routinely perform BAL culture via bronchoscopy or ETA culture within the first 48 h after intubation. We retrospectively reviewed cases that underwent BAL and ETA. The patients were divided into two groups: Group B (BAL) and Group E (ETA). Various parameters were evaluated and compared between the two groups. RESULTS: The demographic data and blood test results were similar between the two groups. However, ICU stay, duration of intubation, and culture positivity were significantly higher in Group B. Although not statistically significant, the mortality rate was higher in Group E. The most commonly isolated microorganisms were Candida species. CONCLUSION: The observed mortality rates were consistent with the existing literature. Since the microorganism isolation rate is higher with BAL, leading to more effective antimicrobial treatment, early deaths were prevented, and ICU stay durations were prolonged. Conversely, these durations were shorter in the ETA group due to higher mortality. In intubated COVID-19 patients, a more effective treatment process can be achieved by clearing the airway with fiberoptic bronchoscopy and tailoring the treatment based on BAL culture results. This approach may positively impact prognosis and mortality rates.

Single-cell analysis reveals lasting immunological consequences of influenza infection and respiratory immunization in the pig lung.

The pig is a natural host for influenza viruses and integrally involved in virus evolution through interspecies transmissions between humans and swine. Swine have many physiological, anatomical, and immunological similarities to humans, and are an excellent model for human influenza. Here, we employed single cell RNA-sequencing (scRNA-seq) and flow cytometry to characterize the major leukocyte subsets in bronchoalveolar lavage (BAL), twenty-one days after H1N1pdm09 infection or respiratory immunization with an adenoviral vector vaccine expressing hemagglutinin and nucleoprotein with or without IL-1ß. Mapping scRNA-seq clusters from BAL onto those previously described in peripheral blood facilitated annotation and highlighted differences between tissue resident and circulating immune cells. ScRNA-seq data and functional assays revealed lasting impacts of immune challenge on BAL populations. First, mucosal administration of IL-1ß reduced the number of functionally active Treg cells. Second, influenza infection upregulated IFI6 in BAL cells and decreased their susceptibility to virus replication in vitro. Our data provide a reference map of porcine BAL cells and reveal lasting immunological consequences of influenza infection and respiratory immunization in a highly relevant large animal model for respiratory virus infection.

Detection and Genomic Characterization of Torque Teno Virus in Pneumoconiosis Patients in China.

Pneumoconiosis is a common occupational disease that can worsen with accompanying infection. Torque teno virus (TTV) is a prevalent human virus with multiple genotypes that can chronically and persistently infect individuals. However, the prevalence of TTV in pneumoconiosis patients is still unclear. This research aims to detect the presence and prevalence of TTV in the alveolar lavage fluid of pneumoconiosis patients in the Hunan Province of China using PCR. As a result, a 65.5% positive rate (19 out of 29) of TTV was detected. The TTV detection rate varies among different stages of silicosis and different pneumoconiosis patient ages. Nine novel TTV genomes ranging in size from 3719 to 3908 nt, named TTV HNPP1, HNPP2, HNPP3, HNPP4, HNPP5, HNPP6-1, HNPP6-2, HNPP7-1 and HNPP7-2, were identified. A genomic comparison and phylogenetic analysis indicated that these nine TTVs represent five different species with high genetic diversity which belong to the genus Alphatorquevirus. HNPP6-1 and HNPP6-2 belong to TTV3, HNPP5 belongs to TTV13, HNPP1 belongs to TTV24, HNPP4 belongs to TTV20, and the others belong to TTV19. The genomes of TTV HNPP1, HNPP6-1, and HNPP6-2 contain three putative open reading frames (ORFs) coding for proteins, ORF1, ORF2, and ORF3, while the other six TTV genomes contain two ORFs coding for proteins, ORF1 and ORF2. These results provide the first description of TTV epidemiology in pneumoconiosis patients in China. The newly identified TTV genome sequences reveal the high genetic diversity of TTV in pneumoconiosis patients and could contribute to a deeper understanding of TTV retention and infection in humans.

The bacterial and fungal profiles of patients hospitalized with non-COVID-19 lower respiratory tract infections in Wuhan, China, 2019-2021.

AIMS: A severe lockdown occurred in Wuhan during the COVID-19 pandemic, followed by a remission phase in the pandemic's aftermath. This study analyzed the bacterial and fungal profiles of respiratory pathogens in patients hospitalized with non-COVID-19 lower respiratory tract infections (LRTIs) during this period to determine the pathogen profile distributions in different age groups and hospital departments in Wuhan. METHODS AND RESULTS: We collected reports of pathogen testing in the medical records of patients hospitalized with non-COVID-19 LRTI between 2019 and 2021. These cases were tested for bacterial and fungal pathogens using 16S and internal transcribed spacer sequencing methods on bronchoalveolar lavage fluid samples. The study included 1368 cases. The bacteria most commonly identified were Streptococcus pneumoniae (12.50%) and Mycoplasma pneumoniae (8.33%). The most commonly identified fungi were Aspergillus fumigatus (2.49%) and Pneumocystis jirovecii (1.75%). Compared to 2019, the S. pneumoniae detection rates increased significantly in 2021, and those of M. pneumoniae decreased. Streptococcus pneumoniae was detected mainly in children. The detection rates of almost all fungi were greater in the respiratory Intensive Care Unit compared to respiratory medicine. Streptococcus pneumoniae and M. pneumoniae were detected more frequently in the pediatric department. CONCLUSIONS: Before and after the COVID-19 outbreak, a change in the common pathogen spectrum was detected in patients with non-COVID-19 in Wuhan, with the greatest change occurring among children. The major pathogens varied by the patient's age and the hospital department.

Bidirectional transfer of human cytomegalovirus strains in donor and recipient seropositive lung transplant patients.

Donor and recipient human cytomegalovirus (HCMV) seropositive (D+R+) lung transplant recipients (LTRs) often harbor multiple strains of HCMV, likely due to transmitted donor (D) strains and reactivated recipient (R) strains. To date, the extent and timely occurrence of each likely source in shaping the post-transplantation (post-Tx) strain population is unknown. Here, we deciphered the D and R origin of the post-Tx HCMV strain composition in blood, bronchoalveolar lavage (BAL), and CD45+ BAL cell subsets. We investigated either D and/or R formalin-fixed paraffin-embedded blocks or fresh D lung tissue from four D+R+ LTRs obtained before transplantation. HCMV strains were characterized by short amplicon deep sequencing. In two LTRs, we show that the transplanted lung is reseeded by R strains within the first 6 months after transplantation, likely by infiltrating CD14+ CD163+/- alveolar macrophages. In three LTRs, we demonstrate both rapid D-strain dissemination and persistence in the transplanted lung for >1 year post-Tx. Broad inter-host diversity contrasts with intra-host genotype sequence stability upon transmission, during follow-up and across compartments. In D+R+ LTRs, HCMV strains of both, D and R origin can emerge first and dominate long-term in subsequent episodes of infection, indicating replication of both sources despite pre-existing immunity.

Epstein-Barr virus qPCR testing on bronchoalveolar lavage fluid from immunocompromised patients.

BACKGROUND: Epstein-Barr Virus (EBV) is associated with lung disease in immunocompromised patients, particularly transplant recipients. EBV DNA testing of lower respiratory tract specimens may have diagnostic utility. METHODS: This was a retrospective, observational study of all patients with bronchoalveolar lavage (BAL) fluids submitted for EBV qPCR testing from February 2016 to June 2022 at the Stanford Clinical Virology Laboratory. RESULTS: There were 140 patients that underwent 251 EBV qPCR BAL tests (median 1; range 1 - 10). These patients had a mean age of 15.9 years (standard deviation, 15.1 years) and 50 % were female. Transplant recipients accounted for 67.1 % (94/140) of patients, including 67.0 % (63/94) solid organ transplant (SOT) and 33.0 % (31/94) hematopoietic cell transplant. Diagnostic testing was performed more commonly than surveillance testing [57.0 % (143/251) v. 43.0 % (108/251)]; 96.2 % (104/108) of surveillance samples were from lung transplant recipients. Excluding internal control failures, 34.7 % (83/239) of BAL had detectable EBV DNA, encompassing a wide range of viral loads (median=3.03 log10 IU/mL, range 1.44 to 6.06). Overall agreement of EBV DNA in BAL compared to plasma was 74.1 % [117/158; 95 % confidence interval (CI): 66.5 % to 80.7 %], with a kappa coefficient of 0.44 (95 % CI: 0.30 to 0.57). Only 20.1 % (48/239) of results were discussed in a subsequent clinical note, and one result (0.4 %; 1/239) changed clinical management. CONCLUSIONS: EBV qPCR testing on BAL offers limited clinical impact. Additional biomarkers are required to improve the diagnosis of EBV-associated lung diseases.

Comparative single-cell analysis reveals IFN-γ as a driver of respiratory sequelae after acute COVID-19.

Postacute sequelae of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (PASC) represent an urgent public health challenge and are estimated to affect more than 60 million individuals globally. Although a growing body of evidence suggests that dysregulated immune reactions may be linked with PASC symptoms, most investigations have primarily centered around blood-based studies, with few focusing on samples derived from affected tissues. Furthermore, clinical studies alone often provide correlative insights rather than causal mechanisms. Thus, it is essential to compare clinical samples with relevant animal models and conduct functional experiments to understand the etiology of PASC. In this study, we comprehensively compared bronchoalveolar lavage fluid single-cell RNA sequencing data derived from clinical PASC samples and a mouse model of PASC. This revealed a pro-fibrotic monocyte-derived macrophage response in respiratory PASC, as well as abnormal interactions between pulmonary macrophages and respiratory resident T cells, in both humans and mice. Interferon-γ (IFN-γ) emerged as a key node mediating the immune anomalies in respiratory PASC. Neutralizing IFN-γ after the resolution of acute SARS-CoV-2 infection reduced lung inflammation and tissue fibrosis in mice. Together, our study underscores the importance of performing comparative analysis to understand the cause of PASC and suggests that the IFN-γ signaling axis might represent a therapeutic target.

Clinical significance of lower respiratory tract culture within 48 h of admission in patients with viral pneumonia: an observational study.

BACKGROUND: The aim of this retrospective study was to examine the risk factors of positive lower respiratory tract cultures and to investigate whether nosocomial infections are common in patients with positive lower respiratory tract cultures. METHODS: We enrolled 86 patients diagnosed with influenza A-related critical illness who were treated at Fuzhou Pulmonary Hospital of Fujian in China between 1st October 2013 and 31st March 2019. The of admission were used to divide the enrolled patients into two groups. Sputum and bronchoalveolar lavage fluid specimens were collected within 48 h after admission for culture. All samples were cultured immediately after sampling. Nosocomial infections are defined as any symptom or sign of pulmonary infiltration, confirmed by X-ray, after 5 days of admission and positive results from one or more cultures. RESULTS: The average age of this cohort was (54.13 ± 16.52) years. Based on the culture results, Staphylococcus aureus and Candida albicans had the highest positive rates (3.40% (3/86) and 20.90% (18/86), respectively). In patients with positive lower respiratory tract cultures, the incidence of nosocomial infection was 73.30% (22/30) five days after admission. However, the incidence of nosocomial infection was lower (42.80%, 24/56) in patients with negative lower respiratory tract cultures. Hemoptysis, systolic pressure at admission, and blood urea nitrogen level at admission were all independent risk factors for positive lower respiratory tract cultures within 48 h of admission. CONCLUSION: Our data showed that a significant proportion of patients with pneumonia exhibited co-infections with bacteria or fungi within five days of hospital admission. Hemoptysis, systolic pressure, and blood urea nitrogen levels at admission emerged as the key risk factors. These findings underscore the necessity of closely monitoring patients with influenza infection, particularly for positive bacterial or fungal cultures within the initial 48 h of admission.

Bacteriophages isolated from mouse feces attenuates pneumonia mice caused by Pseudomonas aeruginosa.

BACKGROUND: Most of the current bacteriophages (phages) are mostly isolated from environments. However, phages isolated from feces might be more specific to the bacteria that are harmful to the host. Meanwhile, some phages from the environment might affect non-pathogenic bacteria for the host. METHODS: Here, bacteriophages isolated from mouse feces were intratracheally (IT) or intravenously (IV) administered in pneumonia mice caused by Pseudomonas aeruginosa at 2 hours post-intratracheal bacterial administration. As such, the mice with phage treatment, using either IT or IV administration, demonstrated less severe pneumonia as indicated by mortality, serum cytokines, bacteremia, bacterial abundance in bronchoalveolar lavage fluid (BALF), and neutrophil extracellular traps (NETs) in lung tissue (immunofluorescence of neutrophil elastase and myeloperoxidase). RESULTS: Interestingly, the abundance of phages in BALF from the IT and IV injections was similar, supporting a flexible route of phage administration. With the incubation of bacteria with neutrophils, the presence of bacteriophages significantly improved bactericidal activity, but not NETs formation, with the elevated supernatant IL-6 and TNF-α, but not IL-1ß. In conclusion, our findings suggest that bacteriophages against Pseudomonas aeruginosa can be discovered from feces of the host. CONCLUSIONS: The phages attenuate pneumonia partly through an enhanced neutrophil bactericidal activity, but not via inducing NETs formation. The isolation of phages from the infected hosts themselves might be practically useful for future treatment. More studies are warranted.