Molecular epidemiology of Pneumocystis jiroveci in human immunodeficiency virus-positive and -negative immunocompromised patients in The Netherlands

2014 ◽  
Vol 63 (10) ◽  
pp. 1294-1302 ◽  
Author(s):  
Marijke J. Vanspauwen ◽  
Vera E. J. Knops ◽  
Cathrien A. Bruggeman ◽  
Walther N. K. A. van Mook ◽  
Catharina F. M. Linssen
Keyword(s):  
The Netherlands ◽  
Clinical Relevance ◽  
Lavage Fluid ◽  
Underlying Disease ◽  
Pneumocystis Pneumonia ◽  
Surface Glycoprotein ◽  
Immunocompromised Patients ◽  
Pneumocystis Jiroveci ◽  
Major Surface ◽  
Pcr Targeting

Pneumocystis jiroveci infections can cause pneumocystis pneumonia (PCP) or lead to colonization without signs of PCP. Over the years, different genotypes of P. jiroveci have been discovered. Genomic typing of P. jiroveci in different subpopulations can contribute to unravelling the pathogenesis, transmission and spread of the different genotypes. In this study, we wanted to determine the distribution of P. jiroveci genotypes in immunocompetent and immunocompromised patients in The Netherlands and determine the clinical relevance of these detected mutations. A real-time PCR targeting the major surface glycoprotein gene (MSG) was used as a screening test for the presence of P. jiroveci DNA. Samples positive for MSG were genotyped based on the internal transcribed spacer (ITS) and dihydropteroate synthase (DHPS) genes. Of the 595 included bronchoalveolar lavage fluid samples, 116 revealed the presence of P. jiroveci DNA. A total of 52 of the 116 samples were ITS genotyped and 58 DHPS genotyped. The ITS genotyping revealed 17 ITS types, including two types that have not been described previously. There was no correlation between ITS genotype and underlying disease. All ITS- and DHPS-genotyped samples were found in immunocompromised patients. Of the 58 DHPS-genotyped samples, 50 were found to be WT. The other eight samples revealed a mixed genotype consisting of WT and type 1. The majority of the latter recovered on trimethoprim–sulfamethoxazole suggesting no clinical relevance for this mutation.

scholarly journals The Fungal PCR Initiative's evaluation of in-house and commercial Pneumocystis jirovecii qPCR assays: Toward a standard for a diagnostics assay

2019 ◽  
Vol 58 (6) ◽  
pp. 779-788 ◽  
Author(s):  
Maud Gits-Muselli ◽  
P Lewis White ◽  
Carlo Mengoli ◽  
Sharon Chen ◽  
Brendan Crowley ◽  
...  
Keyword(s):  
Target Genes ◽  
Clinical Care ◽  
Large Subunit ◽  
Reference Method ◽  
Pneumocystis Pneumonia ◽  
Surface Glycoprotein ◽  
Pneumocystis Jirovecii ◽  
Tubulin Genes ◽  
Major Surface Glycoprotein ◽  
Major Surface

Abstract Quantitative real-time PCR (qPCR) is increasingly used to detect Pneumocystis jirovecii for the diagnosis of Pneumocystis pneumonia (PCP), but there are differences in the nucleic acids targeted, DNA only versus whole nucleic acid (WNA), and also the target genes for amplification. Through the Fungal PCR Initiative, a working group of the International Society for Human and Animal Mycology, a multicenter and monocenter evaluation of PCP qPCR assays was performed. For the multicenter study, 16 reference laboratories from eight different countries, performing 20 assays analyzed a panel consisting of two negative and three PCP positive samples. Aliquots were prepared by pooling residual material from 20 negative or positive- P. jirovecii bronchoalveolar lavage fluids (BALFs). The positive pool was diluted to obtain three concentrations (pure 1:1; 1:100; and 1:1000 to mimic high, medium, and low fungal loads, respectively). The monocenter study compared five in-house and five commercial qPCR assays testing 19 individual BALFs on the same amplification platform. Across both evaluations and for all fungal loads, targeting WNA and the mitochondrial small sub-unit (mtSSU) provided the earliest Cq values, compared to only targeting DNA and the mitochondrial large subunit, the major surface glycoprotein or the beta-tubulin genes. Thus, reverse transcriptase-qPCR targeting the mtSSU gene could serve as a basis for standardizing the P. jirovecii load, which is essential if qPCR is to be incorporated into clinical care pathways as the reference method, accepting that additional parameters such as amplification platforms still need evaluation.

scholarly journals Additional C-type lectin receptors mediate interactions with Pneumocystis organisms and major surface glycoprotein

2021 ◽  
Vol 70 (12) ◽  
Author(s):  
Theodore J. Kottom ◽  
Eva M. Carmona ◽  
Kyle Schaefbauer ◽  
Andrew H. Limper
Keyword(s):  
Mrna Expression ◽  
Immune Cell ◽  
Expression Profiles ◽  
Pneumocystis Carinii ◽  
Pneumocystis Pneumonia ◽  
Immunofluorescence Assay ◽  
Surface Glycoprotein ◽  
Lectin Receptors ◽  
Major Surface ◽  
Surface Glycoproteins

Introduction. Pathogen-associated molecular patterns’ (PAMPs) are microbial signatures that are recognized by host myeloid C-type lectin receptors (CLRs). These CLRs interact with micro-organisms via their carbohydrate recognition domains (CRDs) and engage signalling pathways within the cell resulting in pro-inflammatory and microbicidal responses. Gap statement. In this article, we extend our laboratory study of additional CLRs that recognize fungal ligands against Pneumocystis murina and Pneumocystis carinii and their purified major surface glycoproteins (Msgs). Aim. To study the potential of newly synthesized hFc-CLR fusions on binding to Pneumocystis and its Msg. Methods. A library of new synthesized hFc-CLR fusions was screened against Pneumocystis murina and Pneumocystis carinii organisms and their purified major surface glycoproteins (Msgs) found on the respective fungi via modified ELISA. Immunofluorescence assay (IFA) was implemented and quantified to verify results. mRNA expression analysis by quantitative PCR (q-PCR) was employed to detect respective CLRs found to bind fungal organisms in the ELISA and determine their expression levels in the mouse immunosuppressed Pneumocystis pneumonia (PCP) model. Results. We detected a number of the CLR hFc-fusions displayed significant binding with P. murina and P. carinii organisms, and similarly to their respective Msgs. Significant organism and Msg binding was observed for CLR members C-type lectin domain family 12 member A (CLEC12A), Langerin, macrophage galactose-type lectin-1 (MGL-1), and specific intracellular adhesion molecule-3 grabbing non-integrin homologue-related 3 (SIGNR3). Immunofluorescence assay (IFA) with the respective CLR hFc-fusions against whole P. murina life forms corroborated these findings. Lastly, we surveyed the mRNA expression profiles of the respective CLRs tested above in the mouse immunosuppressed Pneumocystis pneumonia (PCP) model and determined that macrophage galactose type C-type lectin (Mgl-1), implicated in recognizing terminal N-acetylgalactosamine (GalNAc) found in the glycoproteins of microbial pathogens was significantly up-regulated during infection. Conclusion. The data herein add to the growing list of CLRs recognizing Pneumocystis and provide insights for further study of organism/host immune cell interactions.

scholarly journals Gomori Methenamine Silver Stain on Bronchoalveolar Lavage Fluid Is Poorly Sensitive for Diagnosis of Pneumocystis jiroveci Pneumonia in HIV-Negative Immunocompromised Patients and May Lead to Missed or Delayed Diagnoses

2020 ◽  
Vol 144 (8) ◽  
pp. 1003-1010
Author(s):  
Marwan M. Azar ◽  
Rebecca Slotkin ◽  
Rita Abi-Raad ◽  
Yuehong Liu ◽  
Matthew H. Grant ◽  
...  
Keyword(s):  
Bronchoalveolar Lavage ◽  
Bronchoalveolar Lavage Fluid ◽  
Lavage Fluid ◽  
Hiv Positive ◽  
Immunocompromised Patients ◽  
Silver Stain ◽  
Pneumocystis Jiroveci ◽  
Pneumocystis Jiroveci Pneumonia ◽  
Methenamine Silver ◽  
Hiv Negative

Context.— Direct visualization of Pneumocystis jiroveci organisms, using Gomori methenamine silver (GMS) staining in bronchoalveolar lavage fluid (BAL), is a historical gold standard that has been widely used for the diagnosis of P jiroveci pneumonia (PJP). However, the stain may be less sensitive in human immunodeficiency virus (HIV)–negative immunocompromised patients owing to a lower burden of organisms. Objectives.— To assess the sensitivity of the GMS stain on BAL fluid for the diagnosis of PJP in HIV-negative immunocompromised patients as compared to HIV-positive patients. Design.— We conducted a retrospective review from 2012 to 2018 to identify immunocompromised patients (≥18 years old) who underwent bronchoscopy with BAL GMS staining for the diagnosis of PJP. To assess for sensitivity, we sought to identify BAL GMS-positive cases and BAL GMS-negative cases of PJP. The BAL GMS-negative cases were categorized into proven and probable PJP. Results.— We identified 45 adult immunocompromised patients with proven and probable PJP, including 24 HIV-negative (11 BAL GMS-positive and 13 BAL GMS-negative) and 21 HIV-positive cases (all were BAL GMS-positive). The sensitivity of BAL GMS for the diagnosis of PJP in HIV-negative immunocompromised patients was 11 of 24 (46%) versus 21 of 21 (100%) in HIV-positive patients (CD4: median, 10 cells/mL; range, 3–300 cells/mL). Delayed or missed diagnoses were seen in 3 cases of BAL GMS-negative PJP. Re-examination of BAL GMS slides showed rare P jiroveci cysts in 1 case. Conclusions.— BAL GMS has poor sensitivity for PJP in HIV-negative immunocompromised patients. Using BAL GMS as a sole method for PJP may result in missed or delayed diagnoses in this population.

scholarly journals Clinical Evaluation of a (1,3)-β-d-Glucan Assay for Presumptive Diagnosis of Pneumocystis jiroveci Pneumonia in Immunocompromised Patients

2009 ◽  
Vol 16 (10) ◽  
pp. 1524-1526 ◽  
Author(s):  
Valerio Del Bono ◽  
Alessandra Mularoni ◽  
Elisa Furfaro ◽  
Emanuele Delfino ◽  
Lorenzo Rosasco ◽  
...  
Keyword(s):  
Cell Wall ◽  
Clinical Evaluation ◽  
Pneumocystis Pneumonia ◽  
Immunocompromised Patients ◽  
Presumptive Diagnosis ◽  
Pneumocystis Jiroveci ◽  
Pneumocystis Jiroveci Pneumonia ◽  
Reliable Marker

ABSTRACT (1,3)-β-d-Glucan (BG) is a component of the Pneumocystis jiroveci cell wall. Thirty-one immunocompromised patients with pneumonia (16 with presumptive pneumocystis pneumonia [PCP] and 15 with non-PCP) were evaluated for serum BG levels. Serum from all 16 presumptive PCP patients and from 2/15 patients with non-PCP was positive for BG. Results indicate that BG is a reliable marker for diagnosing PCP.

scholarly journals Evaluation of the Amplex eazyplex Loop-Mediated Isothermal Amplification Assay for Rapid Diagnosis of Pneumocystis jirovecii Pneumonia

2020 ◽  
Vol 58 (12) ◽  
Author(s):  
Timo Huber ◽  
Annerose Serr ◽  
Walter Geißdörfer ◽  
Christina Hess ◽  
Christian Lynker-Aßmus ◽  
...  
Keyword(s):  
Working Hours ◽  
Lavage Fluid ◽  
Isothermal Amplification ◽  
Surface Glycoprotein ◽  
Pneumocystis Jirovecii ◽  
Molecular Testing ◽  
Pneumocystis Jirovecii Pneumonia ◽  
Content Type ◽  
Loop Mediated Isothermal Amplification ◽  
Major Surface

ABSTRACT Quantitative PCR (qPCR) assays are the gold standard for diagnosis of Pneumocystis jirovecii pneumonia (PCP). However, they are laborious and require skilled personnel. Therefore, execution outside regular working hours of the molecular biology laboratory is limited. The eazyplex P. jirovecii assay (PJA) uses loop-mediated isothermal amplification for detection of P. jirovecii. It is performed directly with respiratory specimens, without the need for special skills, and delivers a result within 3 to 25 min. The goal of our study was to compare the performance of the eazyplex PJA with that of established P. jirovecii qPCR assays. All archived bronchoalveolar lavage fluid (BALF) samples that had previously tested positive for P. jirovecii by qPCR assay and 50 control samples (retrospective part), as well as all BALF samples received for P. jirovecii analysis over a period of 4 months (prospective part), were tested. Forty-nine patients with proven PCP and 126 patients without PCP were included. The sensitivity and specificity of the eazyplex PJA (95.7% and 96.5%, respectively) were comparable to those for three different P. jirovecii qPCR assays. The detection limit of the eazyplex PJA was analogous to 103 copies of the major surface glycoprotein gene per 25 μl of BALF, corresponding to 10 to 20 P. jirovecii cells. The eazyplex PJA reliably discriminated patients with PCP from patients with P. jirovecii colonization. It delivered a positive result within a mean of 9 min 38 s and required a hands-on time of 2 min 45 s. In summary, the eazyplex PJA showed identical performance for the diagnosis of PCP, compared to qPCR assays. However, in terms of time to result, practicability, and robustness, the eazyplex PJA is clearly superior and allows for around-the-clock molecular testing.

scholarly journals Detection of Pneumocystis jirovecii in Hospitalized Children Less Than 3 Years of Age

2021 ◽  
Vol 7 (7) ◽  
pp. 546
Author(s):  
Estelle Menu ◽  
Jean-Sélim Driouich ◽  
Léa Luciani ◽  
Aurélie Morand ◽  
Stéphane Ranque ◽  
...  
Keyword(s):  
Severe Combined Immunodeficiency ◽  
Hyaline Membrane Disease ◽  
Relevant Information ◽  
Underlying Disease ◽  
Pneumocystis Pneumonia ◽  
Pneumocystis Jirovecii ◽  
Respiratory Pathogens ◽  
Respiratory Sample ◽  
Few Data ◽  
Underlying Diseases

Few data are available in the literature regarding Pneumocystis jirovecii infection in children under 3 years old. This retrospective cohort study aimed to describe medically relevant information among them. All children under 3 years old treated in the same medical units from April 2014 to August 2020 and in whom a P. jirovecii evaluation was undertaken were enrolled in the study. A positive case was defined as a child presenting at least one positive PCR for P. jirovecii in a respiratory sample. Medically relevant information such as demographical characteristics, clinical presentation, microbiological co-infections, and treatments were collected. The objectives were to describe the characteristics of these children with P. jirovecii colonization/infection to determine the key underlying diseases and risk factors, and to identify viral respiratory pathogens associated. The PCR was positive for P. jirovecii in 32 children. Cardiopulmonary pathologies (21.9%) were the most common underlying disease in them, followed by severe combined immunodeficiency (SCID) (18.8%), hyaline membrane disease (15.6%), asthma (9.4%) and acute leukaemia (6.3%). All SCID children were diagnosed with pneumocystis pneumonia. Co-infection with Pj/Rhinovirus (34.4%) was not significant. Overall mortality was 18.8%. Paediatric pneumocystis is not restricted to patients with HIV or SCID and should be considered in pneumonia in children under 3 years old.

Sign in / Sign up

Export Citation Format

Share Document