key: cord-0703720-nvdslalf
authors: Gilbert, David N.; Leggett, James E.; Wang, Lian; Ferdosian, Shirin; Gelfer, Gita D.; Johnston, Michael L.; Footer, Brent W.; Hendrickson, Kathryn W.; Park, Hiromichi S.; White, Emma E.; Heffner, John
title: Enhanced Detection of Community-Acquired Pneumonia Pathogens With the BioFire® Pneumonia FilmArray® Panel
date: 2020-10-16
journal: Diagn Microbiol Infect Dis
DOI: 10.1016/j.diagmicrobio.2020.115246
sha: 9d9c6a995dc2db085b8ab32be1146bb0a5e6ea17
doc_id: 703720
cord_uid: nvdslalf

BACKGROUND: Although most observational studies identify viral or bacterial pathogens in 50% or less of patients hospitalized with community-acquired pneumonia (CAP), we previously demonstrated that a multi-test bundle (MTB) detected a potential pathogen in 73% of patients. This study compares detection rates for potential pathogens with the MTB versus the Biofire® Pneumonia FilmArray® panel (BPFA) multiplex PCR platform and presents an approach for integrating BPFA results as a foundation for subsequent antibiotic stewardship (AS) activities. METHODS: Between January 2017 to March 2018, all patients admitted for CAP were enrolled. Patients were considered evaluable if all elements of the MTB and the BPFA were completed, and they met other a priori inclusion criteria. The primary endpoint was the percentage of potential pathogens detected using the MTB (8 viral and 6 bacterial targets) versus the BPFA (8 viral and 18 bacterial targets). Blood and sputum cultures were performed on all patients. Two or more procalcitonin (PCT) levels assisted clinical assessments as to whether detected bacteria were invading or colonizing. RESULTS: Of 585 enrolled patients, 274 were evaluable. A potential viral pathogen was detected in 40.5% with MTB versus 60.9% of patients with BPFA with an odds ratio (95% CI) of 9.00 (4.12 to 23.30) p<0.01. A potential bacterial pathogen was identified in 66.4% with the MTB vs 75.5% with the BPFA odds ratio (95% CI) of 2.09 (1.24 to 3.59), p 0.003). Low PCT levels helped identify detected bacteria as colonizers.

For hospitalized patients with CAP, the Biofire ® Pneumonia FilmArray ® multiplex platform significantly increased the number of patients with detectable viral and bacterial potential pathogens and, thereby, provided key data for AS activities.

Pneumonia is the leading cause of death due to infection in the United States with an estimated 1.2 million annual hospitalizations [1] . Management of communityacquired pneumonia (CAP) entails empiric antibiotic therapy that targets the most likely bacterial pathogens. The American Thoracic Society-Infectious Diseases Society of America CAP guidelines reserve microbiologic testing for patients with severe CAP [2] .

Further, "rapid, cost-effective, sensitive, and specific diagnostic tests to identify organisms causing CAP have potential to improve routine care by supporting the use of targeted therapy . . ." [2] A benchmark study reported that microbiologic testing identified the microbiologic etiology in only 853 of 2259 (38%) hospitalized CAP patients [3] . To improve this diagnostic yield, we reported that a multi-test bundle (MTB) consisting of sputum and blood bacterial cultures, polymerase chain reaction (PCR) testing of nasopharyngeal swabs (NP) samples by the multiplex Biofire ® (Biofire® Co., Salt Lake City, Utah) Respiratory FilmArray ® panel, anterior nasal swab PCR for Staphylococcus aureus, nasopharyngeal swab for Streptococcus pneumoniae, and urine antigen testing for S. pneumoniae and Legionella pneumophila detected a potential bacterial and/or viral pathogen in 73% of patients [4, 5] . The feasibility of this approach was hampered by long turn-around times (TAT) for multiple MTB components, and the failure of the MTB to detect common potential bacterial pathogens, such as Moraxella and Haemophilus species.

The Biofire® Pneumonia FilmArray® (BPFA) panel probes sputum samples for 8 respiratory viruses and 18 bacteria with a 1 to 2-hour TAT. However, the performance of BPFA as compared with multi-element testing, such as MTB, has neither been evaluated in large studies, nor integrated into clinical practice to identify invasive vs colonizing pathogens. Herein, we compare the detection of potential viral and bacterial pathogens with MTB as compared with BPFA. We hypothesized that the BPFA would detect more potential viral and bacterial pathogens than MTB. We also present our approach to integrating BPFA results into clinical practice by eliminating unnecessary tests and serving as the basis of antibiotic stewardship (AS) activities.

The study was registered with clinicaltrials.gov (NC02580384) as a prospective, non-blinded comparison of pathogen detection with patients as their own controls was conducted at Providence Portland Medical Center, a 480-bed community teaching hospital in Portland, Oregon between January 2017 and March 2018 and approved by the Privacy Board and the Institutional Board (IRB) that waived informed consent.

Subjects were identified from adults (over the age of 18) presenting to the Emergency Department with respiratory symptoms diagnosed by emergency physicians as CAP based on their usual clinical practice. All patients were given a consecutive study number and their clinical data were entered into an online database (FileMaker 13 ® , JEL) reviewed the database to ascertain if the patient was evaluable based on the presence of all inclusion criteria:

1.

Production of sputum, or sputum equivalent (e.g. endotracheal tube aspirate, bronchoalveolar lavage) for testing.

Availability of all elements of the MTB and BPFA and blood samples for PCT levels.

Absence of any extrapulmonary infection that required antibiotics (e.g., pyelonephritis).

No transition to comfort care within 24 hours of admission.

No history of a witnessed aspiration of gastric contents.

Diagnosis not changed by treating physicians to a non-infectious disease (e.g., pulmonary edema) within 48 hours.

The laboratory entered MTB results into the electronic medical record (EMR), and study investigators re-entered EMR data in the FileMaker ® database.

Being investigational, the BPFA results were entered by investigators into the study database. Three authors (DNG, JEL, SF) verified the accuracy of all transcribed data.

The primary objective was to determine if BPFA improved detection rates for potential viral and bacterial pathogens as compared with MTB. We also present an approach to integrating BPFA results into clinical practice aided by PCT testing. Lastly, we suggest that use of BPFA may eliminate the need for selected diagnostic tests in the current MTB. 

The FileMaker ® database calculated a pneumonia severity index (PSI) for each patient [6] . To assess the clinical importance of detected pathogens, patients were divided into four categories (Table 3) based on their clinical and laboratory presentation, PCT test results, and likelihood based on clinical judgment (adjudication) as to whether detected pathogens were colonizing or invasive.

In adults with CAP, detection of a potential viral respiratory pathogen is accepted as compatible with the etiology of the patient's invasive infection as the frequency of asymptomatic colonization is very low. In a prospective controlled detection trial in adult CAP patients, detection of influenza, hMPV, and RSV "probably indicate an etiologic role." Detection of other respiratory viruses, especially in children, requires further study [7] .

The biomarker procalcitonin (PCT) was used to assist in the clinical decision as to whether a detected virus and/or bacteria was a commensal. PCT serum levels do not increase with pure viral infection but do increase with invasive bacterial infection or the combination of viral and bacterial disease [8, 9] .

The BPFA panel includes genomic semi-quantitation of 15 of 18 bacterial targets with the premise that higher bacterial "loads" would be consistent with invasive infection. However, concordance between genomic semi-quantitation and sputum culture CFU/ml is reported between 40 and 54% and the degree of concordance varied by organism [10] [11] [12] . For these reasons, we elected to not use the BPFA semiquantitation result in the adjudication of positive pathogen detection.

We determined the costs of BPFA versus all elements in the MTB based on 2017-

The performance between MTB and the BPFA was compared based on the number of pathogen species (bacterial and/or viral) detected in each patient by the two methods, using the paired t-test and Wilcoxon signed rank test. 

Of 585 enrolled patients, 274 (47.2%) were evaluable and 311 were excluded ( Figure 1 ). 

The proportion of all patients detected by MTB or BPFA with potential bacterial pathogens (± a viral pathogen) or potential viral pathogens (± a bacterial pathogen) are shown in Table 5 

The 

The superiority of the BPFA as compared with MTB in detecting co-infections is reported in Table 6 . In one extreme example, BPFA detected 1 virus (RSV) and 5

bacterial species (E.coli, P. aeruginosa, K. pneumoniae, S. marcescens, and MRSA)

while the MTB detected no virus and 3 bacteria (P. aeruginosa, E. coli, and MSSA).

Detected pathogens are either colonizing or invading and stimulating host innate immune inflammatory response. All 274 evaluable patients had a clinical syndrome consistent with pneumonia; one or more potential pathogens were detected in 261. As the clinical syndromes lack discriminating features, we assessed whether the biomarker PCT could help adjudicate whether the detected potential virus and/or bacteria were colonizing or invading.

PCT levels are a marker of activation of the host innate immune inflammatory response by invading bacterial pathogens but not by pathogenic respiratory viruses [8, 9] . PCT levels increase in the presence of concomitant invasive disease by a virus and/or bacteria.

We divided our patients into four categories depending on the pathogens detected ( Figure 5 ). By convention, a serum PCT elevation of ≥0.25 ng/ml is selected as a clinically meaningful increase. 

Our laboratory estimated the costs for MTB as compared with the projected cost of a new test bundle anchored by the BPFA, Table 7 . Results indicate cost savings for the BPFA approach partially because three nasal swab PCRs and the two urine antigen tests would no longer be needed.

Our results demonstrate, in adult patients hospitalized for CAP, that the Biofire ® Pneumonia FilmArray ® (BPFA) detected more patients with potential viral and bacterial pathogens (90.6%) than the MTB (80.9%), the latter having been our standard of practice [4] .

BPFA sputum detection of 6 of the 7 respiratory viruses was statistically more frequent than testing for the same 7 viruses with the nasopharyngeal Biofire® respiratory FilmArray® panel which is part of the MTB.

The higher yield with the BPFA may reflect higher viral loads in the lower as compared with upper airways [13, 14] . Alternatively, the higher yield may be due to differences in the viral primers used in the BPFA vs the primers in the Biofire respiratory FilmArray® 

Other studies have compared pathogen detection by BPFA with standard of care (SOC) diagnostics. The SOC is not standardized from study to study but includes some combination of cultures of sputum and blood, urine antigens and nasopharyngeal swabs for multiplex PCR panels that most often focus on seasonal respiratory viruses.

In representative studies, the BPFA panel detected a viral and/or bacterial potential pathogen in 47.4 to 72% of the CAP patients [11, 12, 20, 21] . 

Several studies report increased detection of potential viral and bacterial pathogens in lower respiratory secretions of CAP patients when SOC diagnostics are compared to use of the BPFA [11, 12, 20, 24] . The positive and negative percent agreements concur with some variation as expected with variations in the SOC tests employed.

Rapid, sensitive, and specific identification of potential bacterial pathogens is a premise of improving the care of CAP patients. In theory, identification of a pathogen will allow switching from empiric antibiotic therapy to more focused specific therapy.

Before switching, clinicians need to consider whether the detected bacteria are only colonizing and not a concern.

The BPFA platform provides a semi-quantitative number of bacterial genomes/ml present. Traditional indicators of bacterial invasion correlate with greater the numbers of bacteria, but patients with selected co-morbidities may be only colonized and yet have a high density of bacteria: e.g., cystic fibrosis, bronchiectasis, colonized tracheostomy. Growth characteristics of organisms are another variable. Antibiotic therapy can influence bacterial density. Hence, we elected to not utilize the results of bacterial quantitation in our adjudication of whether the detected bacteria are colonizing or invading.

We elected to use the serum biomarker procalcitonin (PCT) to assist in the assessment of whether detected bacteria were colonizing or invading. Elevations of serum PCT indicate activation of innate immunity [8, 9] . PCT gene activation does not occur with pure viral infection but promptly increases in response to invasion by bacteria to include dual infection with a virus [25] [26] [27] . The other variables that influence PCT serum levels; e.g., impaired renal function, are known [8] .

Jain and Pavia point out that the "holy grail" would apply molecular diagnostics to lung tissue [28] . In the absence of lung tissue, we integrated BPFA results with the clinical presentation and PCT test results to characterize with an a priori classification system the likelihood of CAP due to bacterial and/or viral pathogens (Table 3 ). This approach allows clinicians to focus the antimicrobial regimen and promote antibiotic stewardship.

Based on previous research, we selected PCT serum levels of ≥0.25 ng/ml as consistent with activation of an innate immune response by invasive bacteria [25, 26] .

PCT levels of <0.25 ng/ml were interpreted as consistent with colonization (negative predictive value for invasive bacterial infection is >90%) [1, 9, 17] . The potential value of pathogen detection coupled to PCT levels is acknowledged in multiple publications [25] [26] [27] [28] [29] . A recent economic model supported use of PCT serum levels as a way to reduce hospital costs, decrease the risk of antibiotic resistance and Clostridium difficile infections, and enhance clinical decision-making beyond clinical judgment and heuristic approaches [30] .

The diagnostic yield of BPFA creates an opportunity to simplify diagnostic testing for CAP. The BPFA has PCR primers for all Legionella species causing disease in humans suggesting elimination of the urine antigen test for L. pneumophila. Also, the urine antigen test and NP PCR for S. pneumoniae may be eliminated because BPFA outperformed both. The low yield of blood cultures compared to BPFA suggests they can be reserved for ICU patients [2] . Of note, nasal swabs for S. aureus PCR may remain an optional test added to BPFA as nasal samples were 9% more sensitive than sputum samples submitted to BPFA. Sputum cultures remain necessary to provide antibiotic susceptibility testing of detected bacteria.

A new test bundle would include BPFA, sputum culture and sensitivity, blood cultures for the critically ill, and perhaps nasal S. aureus PCR. Two (2) PCT levels 4-6 hours apart for their high negative predictive value and to help discriminate between infection and colonization. This bundle replaces traditional testing, which may take 1-2 days to complete vs a bundle anchored by BPFA that takes less than 2 hours. Moreover, this proposed bundle decreases the cost of testing (Table 7) and, in theory, can reduce antibiotic consumption, and shorten length of hospitalization [21] .

Patients inability to provide sputum and the difficulty of ED physicians in this single center study to make an accurate diagnosis of CAP excluded a majority of patients from the study. To avoid excluding patients from BPFA testing, studies are needed to assess the potential value of oropharyngeal swab specimens in lieu of sputum for testing, as has been investigated for children [31] . Only 13 patients were excluded because of incomplete test results beyond failure to collect sputum. Second, despite the large panel size, a failure of the PCR panels to detect bacteria or virus does not exclude the presence of an undetected virus and/or bacteria. Third, without systematic antibody seroconversion testing, we could not provide definite differentiation between invasive viral disease vs colonization. However, colonization of adults is rare.

Lastly, we did not document the influence of rapid pathogen detection on antibiotic therapy, antibiotic adverse events to include C. difficle infection, antibiotic resistance, or need for patient isolation.

For hospitalized adult patients with CAP, the BPFA multiplex platform detected significantly more common viral and bacterial potential pathogens as compared to a MTB. Serum PCT levels can help in the judgement as to whether detected bacteria are colonizing or causing infection.

Future studies are needed to critically assess the influence of enhanced pathogen detection on antibiotic usage, prevalence of antibiotic adverse effects and antibiotic resistance, and other facets of antimicrobial stewardship

This study was funded by Biofire® Diagnostics, Salt Lake City, Utah Table 2 . Some patients were detected with more than one bacterial species. 

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