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Keywords:

  • (1[RIGHTWARDS ARROW]3)-β-d-glucan;
  • acquired immunodeficiency syndrome;
  • haematological neoplasms;
  • human immunodeficiency virus;
  • pneumocystis;
  • pneumonia;
  • sensitivity and specificity

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References

Pneumocystis jirovecii pneumonia (PCP) can affect various types of immunocompromised patients. We sought to evaluate the diagnostic accuracy of (1[RIGHTWARDS ARROW]3)-β-d-glucan (BDG) for the diagnosis of PCP. We carried out a meta-analysis of relevant studies, identified through PubMed and Scopus. Eligible studies were those that reported BDG diagnostic data in cases with documented PCP and controls with other conditions. Cases of invasive fungal infections and healthy controls were excluded. We performed a bivariate meta-analysis of sensitivity and specificity and constructed a hierarchical summary receiver operating characteristics (HSROC) curve. Fourteen studies were included in the meta-analysis. BDG data were analysed for 357 PCP cases and 1723 controls. The average (95% confidence interval) sensitivity and specificity of BDG were 94.8% (90.8–97.1%) and 86.3% (81.7–89.9%), respectively. The positive and negative likelihood ratios were 6.9 (5.1–9.3) and 0.06 (0.03–0.11), respectively. The area under the HSROC curve was 0.965 (0.945–0.978). Serum BDG shows excellent sensitivity and very good specificity in the diagnosis of PCP. Still, in clinical practice the test results should be interpreted in the context of the underlying clinical characteristics of the individual patient.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References

Pneumocystis jirovecii pneumonia (PCP) is a common and potentially serious opportunistic infection affecting patients with immunosuppression due to various conditions. Such conditions include infection with human immunodeficiency virus (HIV), as well as haematological malignancies, solid organ transplantation, solid tumours, and treatment with certain immunosuppressive agents [1].Although PCP often has a characteristic clinical presentation with fever, dry cough, dyspnoea, hypoxia and diffuse ground glass opacities in the chest X-ray, the differential diagnosis of pulmonary infiltrates in patients with immunosuppression is broad [2,3].

Establishing the diagnosis of PCP requires identification of this fungal pathogen using various methods in respiratory tract samples from patients with a compatible clinical presentation. Such samples include induced sputum and bronchoalveolar lavage fluid [3].However, many patients are not able to provide an appropriate sputum specimen, while bronchoscopy might not be readily available or safe to perform in some of these patients [4]. The diagnosis of PCP can be particularly problematic for patients with immunosuppression related to conditions other than HIV infection [5]. These patients typically have PCP that progresses rapidly, which does not allow for delay in the institution of specific treatment. They also have a relatively lower burden of the pathogen in respiratory tract secretions, which is associated with lower diagnostic yield [6]. Additionally, the diagnosis can be obscured in patients who receive PCP prophylaxis, owing to altered clinical course of PCP and lower counts of the organism in respiratory specimens [7].

In routine clinical practice, some patients with suspected PCP will receive empirical treatment, mainly with trimethoprim-sulphamethoxazole. This could be associated with unnecessary drug toxicity (e.g. hypersensitivity reactions in HIV-infected patients or myelosuppression in haematological patients) [8]. An incorrect clinical diagnosis of PCP could also be associated with failure to detect and treat other potentially serious conditions [3].

Some serum or plasma biomarkers have been studied with the aim of improving the diagnosis of PCP [9]. (1[RIGHTWARDS ARROW]3)-β-d-glucan (BDG) is a candidate such marker. It is an element of the cyst wall of P. jirovecii (and of the cell wall of most other fungi), that can trigger various host inflammatory responses [10]. We sought to evaluate the diagnostic accuracy of BDG for the diagnosis of PCP with a meta-analysis of relevant studies.

Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References

Literature search

We searched in PubMed and Scopus bibliographical databases, up to August 2011, to identify studies that have evaluated the diagnostic value of serum or plasma BDG between patients with PCP and controls without PCP. The search term used for both the above databases was ‘glucan AND pneumocystis’. We additionally searched in the reference lists of relevant articles to identify eligible for inclusion studies.

Study selection criteria

For a study to be eligible for inclusion in the meta-analysis, the diagnosis of PCP had to be made in patients who had relevant clinical manifestations with detection of the pathogen in sputum or bronchoalveolar lavage fluid, either by microscopy using conventional or immunofluorescent stains for P. jirovecii or with polymerase chain reaction; alternatively, the detection of P. jirovecii could have been achieved by histopathology in lung biopsy or autopsy specimens. Infants with primary P. jirovecii lung infection were also included as cases. Cases diagnosed with probable PCP based on clinical and radiological findings alone were excluded. Controls needed to be patients with diagnoses other than PCP (such as patients with other respiratory infections/diseases and patients with conditions rendering them at risk for PCP or an invasive fungal infection). Patients with documented invasive fungal infections and healthy controls were excluded. If a study did not report specific data to allow for the exclusion of the above two patient groups, it was included only if the patients in these groups constituted <25% of the total control group. Studies that provided BDG diagnostic data that could not make mathematical sense according to known mathematical formulas were excluded. Articles written in languages other than English, Spanish, French, German or Italian were also excluded.

Data extraction

From each of the included studies we extracted data on study design, the characteristics of the patient groups that were of interest to our meta-analysis, the criteria for the diagnosis of PCP, the strategy of BDG sampling, the criteria used for the definition of a positive test result, the methodology of BDG testing, and whether or not antimicrobial prophylaxis for PCP had been administered. We also extracted BDG diagnostic data, including the test cut-off level and the number of patients with true/false positive and true/false negative BDG results. If the actual numbers for any of the above patient categories were not directly provided, we calculated these numbers from data on sensitivity/specificity or positive/negative predictive value using common mathematical formulas. If diagnostic data for more than one cut-off level were reported, we included data on the cut-off level that was closer to the one approved for each BDG assay.

Data analysis

We carried out a meta-analysis of the diagnostic accuracy of BDG using a statistical model of bivariate meta-analysis of sensitivity and specificity [11,12]. We also calculated the positive/negative likelihood ratio (the ratio of the probabilities that the test will be positive/negative in cases with PCP vs. those without PCP) and the diagnostic odds ratio (which can be expressed as the positive likelihood ratio/negative likelihood ratio) [13]. We constructed a hierarchical summary receiver operating characteristics (HSROC) curve [11]. We assessed the presence of between-studies statistical heterogeneity using the chi-squared test [14]. A p value <0.1 for the chi-squared test indicated the presence of statistical heterogeneity. We also assessed the presence of between-studies heterogeneity by visual inspection of the ROC plane [15]. For all analyses performed, we used the midas module in Stata software v. 10 [16].

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References

Our bibliographical search in PubMed and Scopus yielded initially 141 and 206 articles, respectively. Fourteen studies were identified as eligible for inclusion in the meta-analysis [17–30]. Specifically, 11 eligible for inclusion studies were identified between the PubMed search results (Fig. 1) [17–21,23,24,26–30]. Four additional articles were selected for further evaluation among the Scopus search results and two of them were eligible for inclusion [17,22]. Οne additional study was included after hand-searching the bibliographical references of relevant articles [25].

image

Figure 1.  Flow diagram for the process of selection of articles for inclusion in the meta-analysis from PubMed search results.

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Characteristics of the included studies

Table 1 presents the main characteristics of the included studies, as well as the BDG diagnostic data extracted from each of the studies and analyzed. The year of publication of the included studies was 2006 or later, except for two studies that were published before 2000 [29,30].The study design was retrospective for all included studies, except for two prospective studies [17,19]. The PCP cases evaluated consisted exclusively of HIV-infected patients in two studies [24,30], of mixed HIV-infected patients and patients with other types of immunosuppression in eight studies [17,18,20,21,23,26,27,29], and of non-HIV immunosuppressed patients in two studies [19,28], while relevant data were not clearly reported in the remaining two studies [22,25]. The specimen used for the detection of P. jirovecii was bronchoalveolar lavage fluid in eight of the included studies [17,19–21,23,24,26,27], bronchoalveolar lavage fluid or other respiratory specimens in four studies [18,22,28,30], and autopsy specimens in the remaining two studies [25,29].

Table 1.   Main characteristics of the studies evaluating (1[RIGHTWARDS ARROW]3)-β-D-glucan (BDG) for the diagnosis of Pneumocystis jirovecii pneumonia, along with the BDG diagnostic data included in the meta-analysis
Author Publication yearStudy designStudy populationDiagnostic criteria for PCPStrategy of BDG sampling (sample evaluated)BDG assayUse of Pneumocystis prophylaxis prior to BDG samplingBDG cut-off value (pg/mL)Se (%)Sp (%)TPFPFNTN
  1. AIDS, acquired immunodeficiency syndrome; BAL, bronchoalveolar lavage; BDG, (1[RIGHTWARDS ARROW]3)–β-D-glucan; IFI, invasive fungal infection; FN, false negative; FP, false positive; GMS, Grocott methenamine silver; HIV, human immunodeficiency virus (infection); LDH, lactate dehydrogenase; NHL, non-Hodgkin lymphoma; NR, not reported; NPV, negative predictive value; PCP, Pneumocystis jirovecii pneumonia; PCR, polymerase chain reaction; pt(s), patient(s); PPV, positive predictive value; RA, rheumatoid arthritis, Se, sensitivity; SLE, systemic lupus erythematosus; SOT, solid organ transplantation; Sp, specificity; TMP-SMX, trimethoprim-sulph

  2. amethoxazole; TN, true negative; TP, true positive.

  3. bAll pts were tested by both tests; data in parentheses are for the β-glucan test Wako.

  4. aMain baseline characteristics for the entire cohort of 871 pts were: malignancy (73%), haematologic malignancy (60%), haematopoietic stem cell transplantation (31%), recent period of neutropenia (21%), recent use of T cell immunosuppressants or prolonged corticosteroid use (61%).

Acosta et al. 2011 [17]Prospective51 medical intensive care unit patients with a clinical syndrome compatible with pneumonia who required diagnostic bronchoscopy and had ≥1 of the following: cirrhosis, HIV, cancer receiving chemotherapy, haematological malignancy, steroid use, T-cell immunosuppressant use or solid organ transplantation Cases: (a) nine invasive aspergillosis cases (four proven, five probable), and one mixed case of probable invasive aspergillosis and proven PCP (we excluded them) (b) three proven PCP casesPositive microscopy stain with monoclonal antibodies in BAL, AND presence of ≥4 of the following criteria: bilateral interstitial infiltrates, hypoxia, fever, dyspnoea, cough, LDH >230 U/L, CD4 cell ≤200 cells/mm3 and appropriate response to specific treatment for PCPUpon inclusion, synchronous to BALFungitellNR801007938030
Damiani et al. 2011 [18]RetrospectiveCases: (a) 14 immunocompetent term infants with primary Pneumocystis infection (b) Six adult patients with PCP (kidney transplantation: 2 pts, HIV infection: 1 pt, bronchial carcinoma: 1 pt, colonic adenocarcinoma: 1 pt, acute alcoholic hepatitis treated with high-dose corticosteroids: 1 pt) Controls: (a) Eight patients colonized by P. jirovecii (bacterial pneumonia: 6 pts, pulmonary sarcoidosis: 1 pt, bronchial carcinoma: 1 pt) b) 14 infants hospitalized for an acute respiratory syndromeCases: (a) Acute respiratory syndrome PLUS real-time PCR on nasopharyngeal aspirate (b) Positive BAL specimens for P. jirovecii by microscopic examination with Giemsa stain AND by immunofluorescence, AND by real-time PCRCases: (a) 2 days before to 3 days after nasopharyngeal aspirate retrieval (b) On the same day as BAL fluid retrieval Controls: (a) 2 days before to 10 days after BAL fluid retrievalFungitellCases: No pt had received antibiotics809591192120
de Boer et al. 2010 [19]Prospective31 HIV-negative, immunocompromised pts (SOT: 20, haematologic malignancy: 8, immunosuppressive therapy: 3) suspected of having PCP based on clinical presentation and chest imaging Cases: 21 pts with PCP Controls: 10 pts (community-acquired pneumonia: 3, interstitial pneumonitis related to underlying disease: 5, Mycobacterium malmoense pulmonary infection: 1, pts who recovered spontaneously: 2)Direct microscopy and/or real-time PCR on BAL fluidAt enrollment and one or two more times during the 1st week of follow-up (the sample at presentation was evaluated)FungitellNo pt used PCP prophylaxis80868918138
Held et al. 2010 [20]RetrospectiveCases: 50 pts with PCP (HIV: 17, haematological malignancy: 15, SOT: 11, immunological disorders: 5, solid tumours: 3) Controls: 50 immunocompromised pts with a suspected infectious respiratory disease and whose BAL fluids were negative for P. jirovecii in immunostainingClinical picture typical of PCP (compatible pulmonary infiltrates and ≥4 of the following criteria: existing immunosuppression, fever, dyspnoea, cough, elevated LDH level and hypoxia) AND positive immunostaining and/or positive PCR findings in BAL fluid WITHOUT any positive culture of fungus from clinically relevant materials, positive serum galactomannan assay or positive serum Candida antigen assayArchived serum sample obtained around the day of bronchoscopy (±7 days)FungitellNR859894493147
Desmet et al. 2009 [21]RetrospectiveCases: all pts with HIV infection (n = 16) or haematological malignancy (n = 12) diagnosed with PCP (we excluded 4 pts who had a concurrent IFI) Controls: (a) HIV-positive pts with CD4 cell counts <200 cells/mm3 and no indication of PCP (n = 16) (b) Pts from haematology wards at risk of IFI but had no identified IFI (n = 12)Progressive dyspnoea and/or fever and/or a non-productive cough lasting <3 months AND detection of P. jirovecii DNA in BAL fluid by PCR AND (detec-tion of Pneumocystis cysts in BAL fluid with GMS stain or monoclonal antibodies OR presence of ≥4 of the following criteria: bilateral interstitial infiltrates upon chest imaging, hypoxia, elevated LDH, and appropriate response to anti-PCP therapy or a CD4 cell count <200 cells/mm3)On the same day or 1 day before the BAL fluid tested positiveFungitellNone of the PCP pts received prophylactic therapy for PCP at the time of diagnosis8010089243025
Koo et al. 2009 [22]RetrospectiveAll pts who had a BDG assay result at a cancer institutea Cases: (a) 14 pts with PCP (b) 98 pts with other proven or probable IFIs (we excluded them) Controls (n = 759): 97 (13%) pts with possible IFI and 662 (87%) pts at risk of IFICompatible clinical syndrome, underlying host factors, consistent radiology findings and cysts in induced sputum or BAL fluidAt least 1 BDG assay result during the study period (the 1st BDG assay per pt was evaluated)FungitellTMP-SMX exposure before BG assay: 5/14 (36%) PCP pts809384131241635
Nakamura et al. 2009 [23]RetrospectiveCases: 35 consecutive pts with PCP (HIV: 19, haematological malignancy: 9, solid tumour: 3, miscellaneous conditions: 4) Controls: (a) 24 pts with microbiologically diagnosed other infectious lung diseases (bacterial pneumonia: 6, tuberculosis: 6, Legionella pneumonia: 15) (b) 15 pts with asymptomatic HIV-1 infection (c) 10 healthy volunteersPositive microscopic results with staining of BAL fluid (Diff Quik, toluidine blue O, or Grocott stain) or positive PCR of BAL fluidOn admissionG testNR209488336243
Watanabe et al. 2009 [24]RetrospectiveCases: 111 HIV-infected pts with proven PCP Controls: 425 HIV infected pts without PCP (3 pts had cryptococcal infection)BAL fluid microscopic examination (Diff-Quik stain)The first measurement of BG was evaluatedFungitec G-test MKBDG was examined before treatment23.29688107524373
Obayashi et al. 2008 [25]RetrospectiveAll cases at a hospital primarily treating malignancies and infectious diseases with autopsy records available (n = 456) and serum tested for BDG within 2 weeks before death (n = 104) Cases: (a) Six cases with PCP (b) 35 cases with other IFI (we excluded them) Controls: 63 cases without IFI or PCPAutopsyBDG test at least 2 weeks before death (a single sample was evaluated)Fungitec G-Test MKNR301008669054
Persat et al. 2008 [26]Multicentre retrospectiveCases: 117 pts with IFI: (a) 20 pts with PCP (HIV: 16, cerebral lymphoma: 1, pulmonary neoplasia: 1, lung graft recipient: 1, dermatomyositis: 1) (b) 97 pts with other IFI (we excluded them) Controls: (a) 122 pts from haematology wards or intensive care units, who were at risk of IFI but had no IFI identified (b) 40 healthy blood donors (we excluded them)Identification of trophozoites or cysts in microscopic examination of BAL fluid after staining with Diff-Quick or Gomori-GMS or toluidine blueThe sample closest to the time of PCP diagnosis was evaluatedFungitellNR80100702036086
Tasaka et al. 2007 [27]Retrospective295 consecutive pts who underwent BAL for the diagnosis of PCP Cases: 57 pts with BAL positive for PCP (haematologic malignancy: 14, HIV: 13, collagen vascular disease: 11; organ transplantation: 5, lung cancer: 4) Controls: 222 pts with BAL negative for PCP (7 pts had invasive aspergillosis and were excluded)Identification of Pneumocystis in BAL fluid with GMS stain or Calcofluor white stainWithin 24 h prior to BALβ-Glucan test WakoNR31.19293227287
Iikuni et al. 2006 [28]Retrospective66 connective tissue disease pts (RA: 42, SLE: 8) who presented with symptoms, radiological findings, and/or laboratory data strongly suggesting PCP and were tested for either sputum or BAL fluid Pneumocystis PCR Cases: 21 pts Controls: 45 ptsPresence of respiratory symptoms and/or radiological findings along with a positive PCR in sputum or BAL fluidNRNRNone of the PCP cases and 11/45 (24%) of the controlsNR7877146420
Mori and Matsumura. 1999 [29]RetrospectivePatients suspected of having systemic fungal infections based on fever with no response to broad-spectrum antibiotics for a week or more Cases: (a) 4 pts with PCP (NHL: 2, AIDS: 1, SLE: 1) (b) 20 pts with other fungal infections (we excluded them) Controls: 15 pts without fungal infection (AML: 5, NHL: 3, other: 7)Findings at autopsy and compatible clinical history (fever unresponsive to broad-spectrum antibiotics, and dyspnoea, cyanosis, hypoxia with alveolar-capillary block signs, and diffuse infiltration or cloudy-glass shadows on chest X-rays) OR detection of P. carinii in the smear of sputa or BAL OR detection by PCRSix specimens for four PCP pts and 16 specimens for 15 controls (any positive sample defined a positive test)Fungitec G-test MK (β-glucan test Wako)bNR20 (11)100 (100)73 (87)4 (4)4 (2)0 (0)11 (13)
Yasuoka et al. 1996 [30]RetrospectiveCases: 7 consecutive pts with AIDS and PCP Controls: (a) Four AIDS pts with other than PCP or fungal infection lung diseases (b) Five HIV pts with oral candidiasis (c) 14 HIV pts without opportunistic infectionsMicroscopic examination of BAL fluid or percutaneous lung aspiration samplesFirst sample was evaluatedG testSamples were obtained within a week of anti-pneumocystis therapy208610060123

Seven of the included studies used the Fungitell assay (Associates of Cape Cod Inc., East Falmouth, MA, USA) for the measurement of BDG [17–22,26]. Three studies used the Fungitec G-test MK assay and two additional studies used the G-test assay (both are developed by Seikagaku Corp, Tokyo, Japan) [23–25,29,30]. Two studies used the β-glucan test Wako (Wako Pure Chemical Industries Ltd, Osaka, Japan) [27,29]. One of the above studies evaluated both the Fungitec G test MK and the β-glucan test Wako [29]. We included diagnostic data referring to the former assay in the meta-analysis. Finally, the remaining study did not specify the BDG assay method used [28]. In all of the included studies, BDG was measured in serum samples, except for one that used plasma [29]. The definition of a positive BDG test was based on a single specified positive test result in 12 of the included studies, while relevant data were not accurately reported in the remaining two studies [28,29].

Diagnostic accuracy of BDG

The 14 analysed studies provided, in total, BDG diagnostic data for 357 patients with PCP and 1723 controls with other conditions. The bivariate meta-analysis resulted in an average sensitivity of BDG for the diagnosis of PCP of 94.8% (95% confidence interval (CI), 90.8–97.1%) and an average specificity of 86.3% (95% CI, 81.7–89.9%) (Fig. 2). Additionally, the average positive likelihood ratio for BDG was 6.9 (95% CI, 5.1–9.3) and the negative likelihood ratio was 0.06 (95% CI, 0.03–0.11). The diagnostic odds ratio was 113.7 (95% CI, 55.7– 232.3). The area under the HSROC curve was 0.965 (95% CI, 0.945– 0.978) (Fig. 3). The p value for the chi-squared test for heterogeneity was 0.31. The visual inspection of the ROC plane (Fig. 3) showed that the summary points for two studies lie clearly out of the 95% prediction ellipse (contour), which is indicative of between-studies heterogeneity [28,30]. These two outliers consisted of a study that did not specifically describe the method used [28] and the oldest of the included studies [30].

image

Figure 2.  Forest plot of the bivariate meta-analysis of the sensitivity and specificity of (1[RIGHTWARDS ARROW]3)-β-D-glucan for the diagnosis of Pneumocystis jirovecii pneumonia. The circles in squares and horizontal lines represent the point estimate and 95% confidence interval, respectively, for each included study; the dotted line represents the average point estimate; the diamond shape represents the 95% confidence interval of the average estimate.

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image

Figure 3.  Summary hierarchical receiver operating characteristics (HSROC) curve of the sensitivity vs. specificity of (1[RIGHTWARDS ARROW]3)-β-D-glucan for the diagnosis of Pneumocystis jirovecii pneumonia. The straight line represents the HSROC curve; the shaded diamond shape represents the point of the curve that corresponds to the average point estimates of sensitivity and specificity; the dashed line represents the 95% confidence area for this point; the numbered circles represent the data from each of the included studies; and the dotted line (the 95% prediction contour) represents the 95% confidence area in which a new relevant study will be located.

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Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References

In this meta-analysis, serum or plasma BDG had an excellent sensitivity and very good specificity for the diagnosis of documented PCP in patients with HIV infection, haematological malignancies or other underlying risk factors. The negative likelihood ratio of BDG was very low, indicating that a negative test is much less likely to be observed in patients with PCP compared with patients without PCP. The positive likelihood ratio was moderately high (7.1), reflecting the fact that BDG might be falsely positive in some patients without PCP. The area under the HSROC curve was more than 0.90, indicating an overall very high diagnostic accuracy of the test. The presence of PCP was documented with polymerase chain reaction or microscopic examination of respiratory tract specimens (bronchoalveolar lavage fluid or sputum) or with histological examination of autopsy specimens. However, the analysed studies differed in many of their characteristics, including the study design, the characteristics of the study population, the reference diagnostic method, and the type of the BDG assay used.

The sensitivity of BDG (94.7%) for the diagnosis of PCP found in this meta-analysis appears to be greater than the sensitivity of BDG for the diagnosis of invasive fungal infections (mainly, invasive aspergillosis or systemic candidiasis) found in another meta-analysis (average sensitivity of 76.8%) [31]. Very high sensitivity of BDG in different groups of patients with PCP has been noted in several studies that have not included a control group and, therefore, they were not eligible for inclusion in this meta-analysis. For example, one study has found that serum or plasma BDG was positive in 33/34 (97.1%) cases of PCP in patients with acquired immunodeficiency syndrome (AIDS) [32]. Another study has found that serum BDG was positive in 15/16 (93.8%) patients with PCP related to various underlying conditions [33]. A third study has reported that BDG was positive in 13/15 (86.7%) patients with PCP and underlying connective tissue diseases [34]. In a larger study that retrospectively evaluated a clinical-trial cohort, the sensitivity of BDG for the detection of 173 cases with confirmed or probable PCP was 92% [35].

The high sensitivity of BDG in the diagnosis of PCP suggests that a negative serum BDG result could reasonably exclude PCP in patients with a low or moderate pretest probability for the disease. For example, if the pretest probability of PCP (which can be estimated subjectively by clinical judgement or can be considered equal to the prevalence of the disease in a specific patient population) is 30%, the probability that PCP will be present after a negative BDG test will be <5%, even if the lowest bound of the 95% interval for sensitivity (90.2%) is used in the calculation.

The specificity of BDG found in this meta-analysis (86.3%) appears similar to the specificity of BDG in patients at risk of invasive fungal infections found in another relevant meta-analysis (average sensitivity of 85.3%) [31]. In clinical practice, care must be taken when evaluating BDG in patients with factors that could account for a false positive BDG result, such as haemodialysis through cellulose membranes, application of certain types of gauzes on mucosal or serosal surfaces, administration of blood products produced through cellulose filters, Gram-negative endotoxinemia, use of certain antimicrobial agents, renal failure, or severe mucositis [36–38]. One or more of the above characteristics are likely to be present in certain patient groups, such as critically ill patients, where the use of the BDG test warrants caution [39].

Most importantly, the presence of an invasive fungal infection as the cause of a positive BDG test, instead of PCP, must be carefully considered for every individual patient. For example, in patients with HIV infection, an elevated BDG could represent underlying histoplasmosis, crytpococcosis or, even probably oesophageal candidiasis, rather than PCP [35]. Likewise, in patients with haematological malignancies, invasive aspergillosis or systemic candidiasis are common causes of an elevated BDG [40]. In our meta-analysis, we excluded cases with invasive fungal infections from the control group, because the aim was to evaluate the diagnostic performance of the test against well-defined reference patient groups. The combined evaluation of BDG with specific diagnostic tests for suspected invasive fungal infections (e.g. serum fungal antigens or galactomannan) has been proposed as a diagnostic strategy to overcome the above limitation [35,41].

Some studies have found very high BDG serum values in the majority of patients with PCP [42]. This has been mostly observed for HIV-infected patients, in whom the counts of the infecting organisms can be particularly high, but also for patients with other types of immunosuppression, such as haematological malignancies [21–23,26]. The above findings, if confirmed, might suggest that the optimal cut-off level of BDG for the diagnosis of PCP could be different to the one used for invasive fungal infections. The HSROC curve we created shows the expected variation in sensitivity and specificity with the variation in the cut-off level. If the aim is to increase the specificity of the test, a slightly higher cut-off level than the one used for the diagnosis of invasive fungal infections could potentially provide higher specificity without unacceptable loss in sensitivity [21].

An important consideration in the evaluation of the utility of any diagnostic test is the accuracy of the reference standard. The sensitivity of the routine diagnostic methods for PCP is not optimal, particularly for certain patient groups [43]. For example, PCP can be strongly suspected clinically and radiographically or on the basis of response to empirical therapy, but the pathogen might be detected with usual microscopic methods. Some such cases in which BDG has been found positive have been reported, suggesting that BDG testing could augment our ability to diagnose PCP [22,44]. Furthermore, the detection of P. jirovecii DNA in sputum samples from immunosuppressed patients with pulmonary infiltrates can sometimes merely represent colonization [45]. Serum BDG could potentially be of value in differentiating between colonization and infection in such cases [46].

Although our meta-analysis shows very high diagnostic accuracy of BDG for PCP, certain limitations should be considered. The main one is that almost all of the included studies were retrospective in nature. Moreover, specific and clinically relevant diagnostic strategies of serial BDG measurements in high-risk patients for PCP were not evaluated in the included studies. A large prospective study of high methodological quality would be useful to corroborate the findings of this meta-analysis. Notably, a study that evaluated a twice-weekly BDG screening strategy for early diagnosis of invasive fungal infections in patients with haematological malignancies has found a high rate of false-positive test results [40].

Secondly, the included studies differed in many of their characteristics, which is commonly observed in diagnostic test accuracy reviews. It cannot be excluded that the accuracy of BDG testing for the diagnosis of PCP may differ between different patient groups. For example, haematological neutropenic patients are at high risk of developing invasive fungal infections, which can be the cause of a positive BDG result. The role of BDG testing in patients receiving PCP prophylaxis has also not been adequately evaluated. Finally, different BDG assays were used in the studies analysed, with potential differences in their diagnostic performance, although there are many similarities between the Fungitell and the Fungitec assays, which were used in the majority of the included studies [25,47]. It should be mentioned, however, that the methods we used for this meta-analysis are valid in the presence of heterogeneity in the characteristics of the analysed studies, including explicit or implicit differences in the diagnostic threshold [48].

In conclusion, this meta-analysis shows a very high accuracy of the serum or plasma BDG assay for the diagnosis of PCP in patients with documented PCP vs. those with other conditions. The very high sensitivity of BDG suggests that a negative result in patients without a high pretest probability of PCP should lead to the consideration of an alternative diagnosis. In addition, a positive BDG result can be a strong indicator for the presence of PCP in patients with compatible clinical and radiological findings. In patients with low pretest probability for PCP, however, a positive BDG should be interpreted in the context of the presence or not of other factors that can increase BDG. An appropriately designed prospective study would be useful to further corroborate the above findings. Certain factors regarding the optimal application of BDG testing for the diagnosis of PCP warrant further evaluation.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Funding
  8. Transparency Declaration
  9. References
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