Oropharyngeal Wash Research Articles

Oropharyngeal Microbiome Analysis in Patients with Varying SARS-CoV-2 Infection Severity: A Prospective Cohort Study.

Patients with COVID-19 infection have distinct oropharyngeal microbiota composition and diversity metrics according to disease severity. However, these findings are not consistent across the literature. We conducted a multicenter, prospective study in patients with COVID-19 requiring outpatient versus inpatient management to explore the microbial abundance of taxa at the phylum, family, genus, and species level, and we utilized alpha and beta diversity indices to further describe our findings. We collected oropharyngeal washing specimens at the time of study entry, which coincided with the COVID-19 diagnosis, to conduct all analyses. We included 43 patients in the study, of whom 16 were managed as outpatients and 27 required hospitalization. Proteobacteria, Actinobacteria, Bacteroidetes, Saccharibacteria TM7, Fusobacteria, and Spirochaetes were the most abundant phyla among patients, while 61 different families were detected, of which the Streptococcaceae and Staphylococcaceae families were the most predominant. A total of 132 microbial genera were detected, with Streptococcus being the predominant genus in outpatients, in contrast to hospitalized patients, in whom the Staphylococcus genus was predominant. LeFSe analysis identified 57 microbial species in the oropharyngeal washings of study participants that could discriminate the severity of symptoms of COVID-19 infections. Alpha diversity analysis did not reveal a difference in the abundance of bacterial species between the groups, but beta diversity analysis established distinct microbial communities between inpatients and outpatients. Our study provides information on the complex association between the oropharyngeal microbiota and SARS-CoV-2 infection. Although our study cannot establish causation, knowledge of specific taxonomic changes with increasing SARS-CoV-2 infection severity can provide us with novel clues for the prognostic classification of COVID-19 patients.

Chicken dendritic cell-targeting nanobodies mediated improved protective effects against H9N2 influenza virus challenge in a homologous sequential immunization study

Global poultry production is still severely affected by H9N2 avian influenza virus (AIV), and the development of a novel universal AIV vaccine is still urgently needed. Neuraminidase (NA) has recently been shown to be an efficient conserved protective antigen.In this study, we fused the extracellular region of the NA gene with a ferritin cassette (pYL281), which resulted in self-assembled 24-mer nanoparticles with the NA protein displayed outside the nanoparticles. In addition, a chicken dendritic cell-targeting nanobody-phage74 was also inserted ahead of the NA protein to yield pYL294. Incubation with chicken bone marrow-derived dendritic cells (chBMDCs) showed that the DC-targeting nanoparticles purified from the pYL294 strain significantly increased the maturation of chBMDCs, as shown by increased levels of CCL5, CCR7, CD83 and CD86 compared with nontargeting proteins. Then, a chicken study was performed using Salmonella oral administration together with intranasal boost with purified proteins. Compared with the other groups, oral immunization with Salmonella harboring pYL294 followed by intranasal boost with purified DC-targeting nanoparticles dramatically increased the humoral IgY and mucosal IgA antibody response, as well as increased the cellular immune response, as shown by elevated splenic lymphocyte proliferation and intracellular mRNA levels of IL-4 and IFN-γ. Finally, sequential immunization with DC-targeting nanoparticles showed increased protection against G57 subtype H9N2 virus challenge compared with other groups, as shown by significantly decreased virus RNA copy numbers in oropharyngeal washes (Days 3, 5 and 7 post challenge) and cloacal washes (Day 7), significantly decreased lung virus titers on Day 5 post challenge and increased body weight gains during the challenge.

Pneumocystis jirovecii Colonization in Mexican Patients with Chronic Obstructive Pulmonary Disease.

The prevalence of colonization by Pneumocystis jirovecii (P. jirovecii) has not been studied in Mexico. We aimed to determine the prevalence of colonization by P. jirovecii using molecular detection in a population of Mexican patients with chronic obstructive pulmonary disease (COPD) and describe their clinical and sociodemographic profiles. We enrolled patients discharged from our hospital diagnosed with COPD and without pneumonia (n = 15). The primary outcome of this study was P. jirovecii colonization at the time of discharge, as detected by nested polymerase chain reaction (PCR) of oropharyngeal wash samples. The calculated prevalence of colonization for our study group was 26.66%. There were no statistically significant differences between COPD patients with and without colonization in our groups. Colonization of P. jirovecii in patients with COPD is frequent in the Mexican population; the clinical significance, if any, remains to be determined. Oropharyngeal wash and nested PCR are excellent cost-effective options to simplify sample collection and detection in developing countries and can be used for further studies.

Radiological Semiotics of Different Types of Nontuberculous Pulmonary Mycobacterioses

Objective: to determine, evaluate, and describe different radiological patterns of microbiologically identified nontuberculous pulmonary mycobacterioses (NTPM) based on multislice computed tomography data. Material and methods. The study included 102 patients with radiological signs of lung disease and different types of NTPM. Slowly growing types of NTPM were detected in 62 (60.8 %) patients, and rapidly growing NTPM – in 40 (39.2 %). The diagnosis was established considering a patient’s complaints, a specified case history, radiological and clinical laboratory data including microscopic studies of sputum from 63 (61.8 %) patients, bronchoalveolar lavage and different types of bronchial biopsies data from 19 (18.6 %) patients, samples of lung video-assisted thoracoscopic surgery from 17 (16.7%) patients, pleural fluid samples from 2 (1.9 %) patients, and oropharyngeal wash samples from 1 (1 %) patient. We used the Somatom Emotion 16 multislice computed tomograph (Siemens) and the high-resolution algorithm (HRCT) with 0.8 mm slice thickness and 1.5 mm slice increment. Results. The HRCT data were highly polymorphic and showed interstitial focal changes, different calibre bronchiectasis, conglomerates or cavities, involvements of vessels or pleural layers. In some patients, changes in the axial interstitium were accompanied by single small focal consolidations located either discretely or in small groups. Peribronchovascular spread of dissemination foci in NTPM was detected by HRCT as irregular infiltration of the axial interstitium (vasculitis type). Changes in the bronchial tree in NTPM were characterized by bronchiolitis symptoms (extensive thickening of bronchial walls, bronchioles) with development of the tree-in-bud sign predominantly in the subpleural lung regions. In some cases, bronchiectatic changes formed conglomerate consolidations of sublobular or lobular extent. Changes of the bronchial tree were detected by HRCT predominantly as signs of deforming bronchitis, cylindrical, varicose, or cystic bronchiectasis, either restricted or spread. Changes might be accompanied by single multi-dimensional cystic bronchiectatic cavities. Conclusion. Typical HRCT signs of NTPM are endobronchial and peribronchovascular spread of foci, development of multi-dimensional conglomerates, deforming bronchitis, bronchiectasis, and presence of single multi-dimensional cystic bronchiectatic cavities.

“A Journey of COVID 19 from Pathogenesis to Therapy”

The novel Corona virus has come up from China, the first outburst of this Pandemic disease was reported in Wuhan, China in December 2019. Hence it is named as Covid-19. The other forms such as SARS and MERS have already prevailed in the nature, but this Covid-19 is more lethal than the those. In this review the author wants to focus on the epidemiology, pathogenesis & Current treatment scenario with little impact on Ayurveda used during this outbreak. The novel Corona virus is very contagious disease which propagates rapidly via droplets of saliva and other bodily fluids. Currently, confirmation of 2019-nCoV infection is performed at CDC using the CDC real-time RT-PCR assay for 2019-nCoV on respiratory specimens (which can include nasopharyngeal or oropharyngeal aspirates or washes, nasopharyngeal or oropharyngeal swabs, bronchioalveolar lavage, tracheal aspirates, or sputum) and serum. There are some new developing approaches, for example artificial learning (AI), and high throughput screenings can possibly accelerate the revelation of treatment. During this Outburst of Corona Virus, it is necessary & need of time to form collaborations across the world to debacle this outbreak. The Researchers now re-emphasized importance in evolving the broad-spectrum Antiviral Drugs to battle the effects of this COVID-19 Virus. There are many antiViral drugs and vaccines which are under clinical trials. The rapid spread of disease permits intense surveillance and isolation protocols to prevent further transmission.

Is It Possible to Differentiate Pneumocystis jirovecii Pneumonia and Colonization in the Immunocompromised Patients with Pneumonia?

Respiratory sample staining is a standard tool used to diagnose Pneumocystis jirovecii pneumonia (PjP). Although molecular tests are more sensitive, their interpretation can be difficult due to the potential of colonization. We aimed to validate a Pneumocystis jirovecii (Pj) real-time PCR (qPCR) assay in bronchoscopic bronchoalveolar lavage (BAL) and oropharyngeal washes (OW). We included 158 immunosuppressed patients with pneumonia, 35 lung cancer patients who underwent BAL, and 20 healthy individuals. We used a SYBR green qPCR assay to look for a 103 bp fragment of the Pj mtLSU rRNA gene in BAL and OW. We calculated the qPCR cut-off as well as the analytical and diagnostic characteristics. The qPCR was positive in 67.8% of BAL samples from the immunocompromised patients. The established cut-off for discriminating between disease and colonization was Ct 24.53 for BAL samples. In the immunosuppressed group, qPCR detected all 25 microscopy-positive PjP cases, plus three additional cases. Pj colonization in the immunocompromised group was 66.2%, while in the cancer group, colonization rates were 48%. qPCR was ineffective at diagnosing PjP in the OW samples. This new qPCR allowed for reliable diagnosis of PjP, and differentiation between PjP disease and colonization in BAL of immunocompromised patients with pneumonia.

Sensitivity of nasopharyngeal, oropharyngeal, and nasal wash specimens for SARS-CoV-2 detection in the setting of sampling device shortage

In the context of an unprecedented shortage of nasopharyngeal swabs (NPS) or sample transport media during the coronavirus disease 2019 (COVID-19) crisis, alternative methods for sample collection are needed. To address this need, we validated a cell culture medium as a viral transport medium, and compared the analytical sensitivity of SARS-CoV-2 RT-PCR in nasal wash (NW), oropharyngeal swab (OPS), and NPS specimens. Both the clinical and analytical sensitivity were comparable in these three sample types. OPS and NW specimens may therefore represent suitable alternatives to NPS for SARS-CoV-2 detection.

Ability to biofilm formation in infectious agents isolated from patients of a large general children’s hospital

Objective: The study objective is to analyze biofilm formation in microorganisms isolated from patients of children’s hospital with severe purulent septic diseases.Methods: The ability to biofilm formation was studied in microorganisms isolated from 32 patients aged from 1 to 15 y.o. with sepsis, acute hematogenous osteomyelitis, peritonitis, and pneumonia in intensive care unit of regional general children’s hospital. Blood, phlegm, bronchial and oropharyngeal washings, wound fluid, peritoneal fluid served as specimens.Results: All tested strains have the ability to biofilm formation; moreover, the majority of them formed moderate biofilms.Conclusion: The common spread of biofilm infections can be related to the most important factors of preservation and distribution of microorganisms in health facilities which significantly limit preventive and therapeutic measures.

Colonización por Pneumocystis jirovecii en mujeres gestantes y recién nacidos en Lima, Perú

BackgroundPneumocystisjirovecii primary infection occurs asymptomatically before 6 months of age, suggesting that the infection is acquired very early in life. Furthermore, Pneumocystis pneumonia has been described in newborns, which emphasizes the importance of studying Pneumocystis colonization in mother-infant pairs. AimsTo evaluate the prevalence of Pneumocystis colonization among pregnant women and to determine the potential transplacental transmission. MethodsA cross-sectional study was carried out on HIV-negative women over 18 years-old, and 37 or more weeks of pregnancy attending Hospital Cayetano Heredia Maternity unit during 2016-2017. Clinical and demographical information was collected on them and their newborns. Oropharyngeal washes, nasal swabs, and placenta samples were collected from women, as well as a nasopharyngeal aspirate and nasal swab from newborns. All respiratory samples were analysed by nested-PCR for the detection of Pneumocystis. Placenta samples from women with a positive PCR result in their respiratory samples were also analysed by nested-PCR. ResultsOf the 92 pregnant women included, five of them (5.43%) were colonized by Pneumocystis. Pneumocystis DNA was not found in any of the 87 available newborn samples or in the placentas of the five women who had a positive result by PCR in their upper respiratory samples. ConclusionsIt was found that 5.43% of the pregnant women were colonized by Pneumocystis, there was no evidence of any role of this colonization in the transmission to their newborns, since none of them tested positive for Pneumocystis.

SARS-CoV-2 and Saliva as a Diagnostic Tool: A Real Possibility

Covid-19 is a respiratory disease caused by the SARS-CoV-2 virus. The high rate of contagion and the spread of the virus in the population make the early detection of the pathogen the means for the adequate targeting of infection control measures. WHO directs sample collection on upper respiratory specimens, including nasopharyngeal and oropharyngeal swab or wash in ambulatory patients, as well as lower respiratory specimens: sputum and/or endotracheal aspirate or bronchoalveolar lavage, in addition to citing blood and feces. Among the various sample collection methods, saliva has been investigated and reported as a potential source for diagnosis. Thus, we propose to evaluate the current scenario, based on recent publications on the perspective of detecting SARS-CoV-2 in saliva as a diagnostic method for Covid-19. The detection of SARS-CoV-2 through saliva seems to be very promising, although obstacles such as the technique and the location of the collection and the sample size of the research carried out so far may present a limitation for its use. The current scenario presents saliva as a reliable method for the detection of SARS-CoV-2, due to the ease of obtaining the samples, the possibility of self-collection, low cost because there is no need to use specific equipment, in addition to reducing the risk of transmission for health professionals.

Airborne acquisition of Pneumocystis in bronchoscopy units: a hidden danger to healthcare workers.

The possible presence of Pneumocystis in a bronchoscopy unit of a tertiary-hospital was examined by detecting Pneumocystis-specific DNA by polymerase chain reaction in prospectively obtained samples of oropharyngeal wash from seven healthcare workers (HCWs) and air from three areas of the unit at different time points (baseline, days +15,+30,+60,+90 after initiation of the study). Positive samples were genotyped at two genetic loci: the mitochondrial large subunit ribosomal RNA (mtLSUrRNA) fragment by direct sequencing and the gene for dihydropteroate synthase (DHPS) by restriction fragment-length polymorphism. Pneumocystis DNA was identified in 13/24 samples from HCWs, in 4/14 air samples and also in two patients with Pneumocystis pneumonia (PcP) and another with a Pneumocystis-associated disease subjected to bronchoscopy on days +15 and +60 after initiation of the study. The HCWs harbored a high rate of mtLSU-rRNA genotypes 1 and 3 and samples from air and patients with only genotype 3. DHPS mutations related to sulpha resistance were detected in three samples from HCWs and in one from air; 65% of the positive samples showed genotypic concordance. The study demonstrates that HCWs of bronchoscopy units represent a new dynamic reservoir and a possible source of infection for human Pneumocystis species, including DHPS genotypes related to sulpha resistance that could be transmitted within hospitals to immunosuppressed hosts in whom a PcP can develop. The results provide the first evidence of the risk of Pneumocystis transmission in the bronchoscopy units and arguments to improve prevention and control of this infection in nosocomial setting.

Protective immunity induced by an intranasal multivalent vaccine comprising 10 Lactococcus lactis strains expressing highly prevalent M-protein antigens derived from Group A Streptococcus.

Streptococcus pyogenes (group A Streptococcus) causes diseases ranging from mild pharyngitis to severe invasive infections. The N-terminal fragment of streptococcal M protein elicits protective antibodies and is an attractive vaccine target. However, this N- terminal fragment is hypervariable: there are more than 200 different M types. In this study, an intranasal live bacterial vaccine comprising 10 strains of Lactococcus lactis, each expressing one N-terminal fragment of M protein, has been developed. Live bacterial-vectored vaccines cost less to manufacture because the processes involved are less complex than those required for production of protein subunit vaccines. Moreover, intranasal administration does not require syringes or specialized personnel. Evaluation of individual vaccine types (M1, M2, M3, M4, M6, M9, M12, M22, M28 and M77) showed that most of them protected mice against challenge with virulent S. pyogenes. All 10 strains combined in a 10-valent vaccine (M×10) induced serum and bronchoalveolar lavage IgG titers that ranged from three- to 10-fold those of unimmunized mice. After intranasal challenge with M28 streptococci, survival of M×10-immunized mice was significantly higher than that of unimmunized mice. In contrast, when mice were challenged with M75 streptococci, survival of M×10-immunized mice did not differ significantly from that of unimmunized mice. Mx-10 immunized mice had significantly less S. pyogenes in oropharyngeal washes and developed less severe disease symptoms after challenge than did unimmunized mice. Our L. lactis-based vaccine may provide an alternative solution to development of broadly protective group A streptococcal vaccines.

Use of Oropharyngeal Washes to Diagnose and Genotype Pneumocystis jirovecii.

Pneumocystis jirovecii is a symbiotic respiratory fungus that presents in 2 clinical forms: pneumonia in immunocompromised patients or colonization, defined by the presence of the organism without associated clinical symptoms. Currently, diagnosis requires invasive bronchoscopy, which may not be available in some settings and is inappropriate for detecting colonization in healthy individuals. Noninvasive diagnostic techniques and molecular strain typing tools that can be used on these samples are critical for conducting studies to better understand transmission. We evaluated 2 real-time polymerase chain reaction (PCR) assays targeting dihydropteroate synthase and the major surface glycoprotein for detection in 77 oropharyngeal washes (OPWs) from 43 symptomatic human immunodeficiency virus-infected patients who underwent bronchoscopy. We also evaluated the ability of a new microsatellite (MS) genotyping panel to strain type infections from these samples. Each PCR used individually provided a high sensitivity (>80%) for detection of pneumonia but a modest specificity (<70%). When used in combination, specificity was increased to 100% with a drop in sensitivity (74%). Concentration of organisms by PCR in the OPW tended to be lower in colonized individuals compared with those with pneumonia, but differences in concentration could not clearly define colonization in symptomatic individuals. Oropharyngeal wash samples were genotyped using 6 MSs with ≥4 alleles successfully genotyped in the majority of colonized patients and ≥5 alleles in patients with pneumonia. The MS profile was consistent over time within patients with serial OPWs analyzed. Microsatellite genotyping on noninvasive samples may aid in studying the molecular epidemiology of this pathogen without requiring invasive diagnostic techniques.

Pneumocystis Pneumonia in Human Immunodeficiency Virus-infected Adults and Adolescents: Current Concepts and Future Directions.

Pneumocystis jirovecii pneumonia (PCP) is one of the most common opportunistic infections in human immunodeficiency virus–infected adults. Colonization of Pneumocystis is highly prevalent among the general population and could be associated with the transmission and development of PCP in immunocompromised individuals. Although the microscopic demonstration of the organisms in respiratory specimens is still the golden standard of its diagnosis, polymerase chain reaction has been shown to have a high sensitivity, detecting Pneumocystis DNA in induced sputum or oropharyngeal wash. Serum β-D-glucan is useful as an adjunctive tool for the diagnosis of PCP. High-resolution computed tomography, which typically shows diffuse ground-glass opacities, is informative for the evaluation of immunocompromised patients with suspected PCP and normal chest radiography. Trimethoprim–sulfamethoxazole (TMP-SMX) is the first-line agent for the treatment of mild to severe PCP, although it is often complicated with various side effects. Since TMP-SMX is widely used for the prophylaxis, the putative drug resistance is an emerging concern.

Use of Oropharyngeal Washes to Diagnose and Genotype Pneumocystis jirovecii

Use of Oropharyngeal Washes to Diagnose and Genotype Pneumocystis jirovecii

Epstein-Barr Virus Latent Membrane Protein 1 Genetic Variability in Peripheral Blood B Cells and Oropharyngeal Fluids

We report the diversity of latent membrane protein 1 (LMP1) gene founder sequences and the level of Epstein-Barr virus (EBV) genome variability over time and across anatomic compartments by using virus genomes amplified directly from oropharyngeal wash specimens and peripheral blood B cells during acute infection and convalescence. The intrahost nucleotide variability of the founder virus was 0.02% across the region sequences, and diversity increased significantly over time in the oropharyngeal compartment (P = 0.004). The LMP1 region showing the greatest level of variability in both compartments, and over time, was concentrated within the functional carboxyl-terminal activating regions 2 and 3 (CTAR2 and CTAR3). Interestingly, a deletion in a proline-rich repeat region (amino acids 274 to 289) of EBV commonly reported in EBV sequenced from cancer specimens was not observed in acute infectious mononucleosis (AIM) patients. Taken together, these data highlight the diversity in circulating EBV genomes and its potential importance in disease pathogenesis and vaccine design. This study is among the first to leverage an improved high-throughput deep-sequencing methodology to investigate directly from patient samples the degree of diversity in Epstein-Barr virus (EBV) populations and the extent to which viral genome diversity develops over time in the infected host. Significant variability of circulating EBV latent membrane protein 1 (LMP1) gene sequences was observed between cellular and oral wash samples, and this variability increased over time in oral wash samples. The significance of EBV genetic diversity in transmission and disease pathogenesis are discussed.

Improved characterization of medically relevant fungi in the human respiratory tract using next-generation sequencing

BackgroundFungi are important pathogens but challenging to enumerate using next-generation sequencing because of low absolute abundance in many samples and high levels of fungal DNA from contaminating sources.ResultsHere, we analyze fungal lineages present in the human airway using an improved method for contamination filtering. We use DNA quantification data, which are routinely acquired during DNA library preparation, to annotate output sequence data, and improve the identification and filtering of contaminants. We compare fungal communities and bacterial communities from healthy subjects, HIV+ subjects, and lung transplant recipients, providing a gradient of increasing lung impairment for comparison. We use deep sequencing to characterize ribosomal rRNA gene segments from fungi and bacteria in DNA extracted from bronchiolar lavage samples and oropharyngeal wash. Comparison to clinical culture data documents improved detection after applying the filtering procedure.ConclusionsWe find increased representation of medically relevant organisms, including Candida, Cryptococcus, and Aspergillus, in subjects with increasingly severe pulmonary and immunologic deficits. We analyze covariation of fungal and bacterial taxa, and find that oropharyngeal communities rich in Candida are also rich in mitis group Streptococci, a community pattern associated with pathogenic polymicrobial biofilms. Thus, using this approach, it is possible to characterize fungal communities in the human respiratory tract more accurately and explore their interactions with bacterial communities in health and disease.

Detection of Pneumocystis in the nasal swabs of immune-suppressed rats by use of PCR and microscopy

BackgroundDetection of Pneumocystis jiroveci colonization in lungs or oral samples due to high sensitivity of PCR methods results in undue treatment of patients without any symptoms of Pneumocystis pneumonia. The aim of the present study is to demonstrate Pneumocystis carinii in rats, immune suppressed by oral and subcutaneous administration of dexamethasone.Material/MethodsBlood, oral, nasal and eye swabs were collected prior to immune suppression and 2, 6, 12 weeks after administration of dexamethasone. Also, samples were collected from lung, heart, liver, kidney, diaphragm, brain, spleen, tongue, muscle, eye, intestine, and feces. Cysts and trophozoites were investigated in stained slides and MSG gene was detected by PCR.ResultsThe results showed that weight loss is significantly higher in rats administered oral dexamethasone (P<0.05). Microscopy was positive only in lungs of rats orally administered dexamethasone. PCR was positive in lungs and oral swabs of rats prior to the administration of dexamethasone. After the administration of dexamethasone, the MSG gene was detected in oral swabs, lungs, spleen, kidney and (for the first time) in nasal swabs. PCR was positive in nasal swabs during the second and sixth weeks of oral and subcutaneous administration of dexamethasone, respectively.ConclusionsPresence of P. jiroveci in nasopharyngeal aspirate, oropharyngeal wash, oral swab, induced sputum or BAL, and absence in nasal swab in a patient without symptoms of PCP may support clinician’s decision regarding colonization. Overall, detection of P. carinii in nasal swabs of rats by PCR demonstrated that nasal sampling can be used for the diagnosis of Pneumocystis pneumonia.

Recent advances in the diagnosis ofPneumocystis jiroveciipneumonia in HIV-infected adults

Pneumocystis jirovecii pneumonia (PCP) is one of the most common opportunistic infections in HIV-infected adults. Although the microscopic demonstration of the organisms in respiratory specimens is still the golden standard of its diagnosis, recent advances in the diagnostic tools have been changing the situation. Colonization of Pneumocystis is highly prevalent among the general population and could be associated with the transmission and development of PCP in immunocompromised individuals. Nested or conventional polymerase chain reaction (PCR) has a high sensitivity, detecting Pneumocystis DNA in induced sputum or oropharyngeal wash, but often produces false positives. Although quantitative real-time PCR is promising for discriminating colonization from PCP, the targeted DNA sequences and the cut-off values remain to be standardized. Serum β-D-glucan is useful as an adjunctive tool for the diagnosis of PCP. High-resolution computed tomography, which typically shows diffuse ground-glass opacities, is informative for evaluation of immunocompromised patients with suspected PCP and normal chest radiography. Although these new tools have been making the diagnosis of PCP less invasive and more accurate, any one of them can not make a definitive diagnosis by itself. The diagnostic criteria based on the combination of the testing ought to be established.

Lung-enriched Organisms and Aberrant Bacterial and Fungal Respiratory Microbiota after Lung Transplant

Long-term survival after lung transplantation is limited by infectious complications and by bronchiolitis obliterans syndrome (BOS), a form of chronic rejection linked in part to microbial triggers. To define microbial populations in the respiratory tract of transplant patients comprehensively using unbiased high-density sequencing. Lung was sampled by bronchoalveolar lavage (BAL) and upper respiratory tract by oropharyngeal wash (OW). Bacterial 16S rDNA and fungal internal transcribed spacer sequencing was used to profile organisms present. Outlier analysis plots defining taxa enriched in lung relative to OW were used to identify bacteria enriched in lung against a background of oropharyngeal carryover. Lung transplant recipients had higher bacterial burden in BAL than control subjects, frequent appearance of dominant organisms, greater distance between communities in BAL and OW indicating more distinct populations, and decreased respiratory tract microbial richness and diversity. Fungal populations were typically dominated by Candida in both sites or by Aspergillus in BAL but not OW. 16S outlier analysis identified lung-enriched taxa indicating bacteria replicating in the lower respiratory tract. In some cases this confirmed respiratory cultures but in others revealed enrichment by anaerobic organisms or mixed outgrowth of upper respiratory flora and provided quantitative data on relative abundances of bacteria found by culture. Respiratory tract microbial communities in lung transplant recipients differ in structure and composition from healthy subjects. Outlier analysis can identify specific bacteria replicating in lung. These findings provide novel approaches to address the relationship between microbial communities and transplant outcome and aid in assessing lung infections.