Scientist Wearing Respirator Mask, Coverall and Safety Glasses Looks at Petri Dishes with Bacteria

A Comprehensive Consensus on Pathogen Detection in Respiratory Infections

 

The development and clinical application of nucleic acid detection technology has significantly improved the diagnostic capability for respiratory pathogens. How to appropriately select nucleic acid testing technology based on patients' underlying conditions, types of respiratory infections, and pathogen spectrum, and correctly understand its clinical application value has become an important clinical issue.

To address this, the Chinese Laboratory Medicine Training Project Expert Committee organized multidisciplinary experts to jointly write the "Expert Consensus on Clinical Application of Nucleic Acid Detection Technology for Adult Respiratory Infection Pathogen Diagnosis (2023)." Professor Xu Yingchun from the Laboratory Department of Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, and Professor Qu Jieming from the Department of Respiratory and Critical Care Medicine, Ruijin Hospital Affiliated to Shanghai Jiao Tong University served as corresponding authors. This article introduces evidence-based guidelines for pathogen detection in respiratory infections, covering molecular and traditional testing methods.

 

I. Acute Upper Respiratory Tract Infections

 

✧ Target Population: High-risk groups for acute upper respiratory tract infections (elderly, children, pregnant women, patients with underlying conditions) and patients with tracheobronchitis.

✧ Key Recommendation: For these populations, pathogen nucleic acid testing is recommended during epidemic seasons of influenza virus, SARS-CoV-2, and in high-risk areas for differential diagnosis, reducing consultation time, and decreasing pathogen transmission risk.

✧ Consensus Rate: 100%

 

Question 1: Should routine pathogen testing be recommended for acute upper respiratory tract infection patients?

The decision should be based on epidemic situations, disease severity, symptom duration, comorbidities, immunosuppression status, relevance of other auxiliary examinations, and preliminary prognosis assessment. For example, during COVID-19 or influenza pandemics, rapid molecular testing aids in triage, reduces consultation time, and minimizes transmission risk. Testing for respiratory viruses is necessary when patients are severely immunocompromised with influenza-like symptoms, or when viral testing results might influence decisions about antiviral medication or antibiotic discontinuation. 

Question 2: What are the recommended pathogen diagnostic methods for acute upper respiratory tract infections?

During high-risk seasons, influenza virus testing for high-risk groups (elderly, children, pregnant women, patients with underlying conditions) may be more cost-effective than empirical anti-influenza treatment. Additionally, rapid result feedback (such as rapid molecular testing with results within 2 hours) can reduce outpatient consultation time, lower influenza transmission risk, and benefit infectious disease control.

 

II. Tracheobronchitis

 

✧ Target Population: Immunocompetent adult patients with tracheobronchitis at risk of hospitalization

✧ Key Recommendation: Multi-target pathogen nucleic acid detection is recommended to identify pathogens and reduce antibiotic use duration and hospital stay.

✧ Consensus Rate: 98.8%

 

Question: What are the indications for pathogen testing in tracheobronchitis patients?

For patients with recurrent acute exacerbations, fever with purulent sputum, or requiring hospitalization, bacterial smear or culture should be considered for prompt pathogen identification to guide antibiotic use.

During influenza virus and SARS-CoV-2 pandemics, outpatient antigen testing or molecular diagnostic techniques for these viruses aid in rapid triage and isolation control, preventing nosocomial transmission, though the former has lower sensitivity than the latter.

For immunocompetent adults at risk of hospitalization, early use of multiplex PCR for pathogen identification may reduce antibiotic use duration and hospital stay. However, routine viral testing is not recommended for children with bronchiolitis.

 

III. Community-Acquired Pneumonia (CAP)

 

✧ Key Recommendation: For severe CAP patients, concurrent routine pathogen examination and nucleic acid testing are recommended to improve pathogen detection rates.

✧ Consensus Rate: 100%

 

Question: What is the value of nucleic acid testing technology in CAP pathogen diagnosis?

For CAP patients, bacterial culture is time-consuming, and fastidious bacteria have low culture positivity rates, making nucleic acid testing advantageous for pathogen diagnosis. For suspected mycoplasma, chlamydia, legionella, or viral infections, serum-specific antigen/antibody testing can be combined with nucleic acid detection and isolation culture. Real-time fluorescent PCR, isothermal amplification, digital PCR, and high-throughput sequencing can simultaneously detect multiple pathogens and provide in-depth analysis of pathogen typing, virulence genes, and resistance genes. RNA high-throughput sequencing is recommended when RNA viral infection is suspected.

For critically ill patients, combining routine pathogen examination with pathogen nucleic acid detection can complement each other, enabling rapid and accurate pathogen identification for better clinical service.

 

IV. Hospital-Acquired Pneumonia (HAP) and Ventilator-Associated Pneumonia (VAP)

 

✧ Key Recommendation: Pathogen diagnosis should primarily rely on traditional culture methods, supplemented with multi-target pathogen nucleic acid detection when necessary. Routine mNGS is not required and should only be considered for immunodeficient patients or when traditional microbiological testing fails to identify pathogens within 3 days and empirical anti-infection treatment is ineffective.

✧ Consensus Rate: 96.4%

 

Question 1: Is routine microbiological specimen collection recommended for HAP/VAP patients?

After clinical diagnosis of HAP/VAP, respiratory, blood, and pleural effusion specimens should be actively collected for microbiological examination. Respiratory specimens should undergo smear microscopy first, followed by pathogen culture, antigen, and nucleic acid testing.

Question 2: How should we understand the value of mNGS in diagnosing HAP/VAP patients?

Common pathogens in HAP/VAP patients are bacteria, and pathological diagnosis primarily relies on traditional culture, supplemented with multi-target pathogen nucleic acid detection such as multiplex PCR and microfluidic chips when necessary. Routine mNGS is not required. DNA-based mNGS should only be considered for immunodeficient patients or when traditional microbiological testing fails to provide clear pathological evidence within 3 days and empirical anti-infection treatment is ineffective.

 

V. Tuberculosis Patients

 

✧ Key Recommendations

Table 1. Recommendations for Tuberculosis Testing in Different Clinical Settings

Clinical Scenario Specific Recommendations Consensus Rate
Patients with Pulmonary Tuberculosis Routine acid-fast bacilli smear, culture and nucleic acid testing are recommended. mNGS and TB-specific PCR have similar diagnostic capabilities for M. tuberculosis 97.60%
Patients with Tuberculous Pleuritis M. tuberculosis culture and nucleic acid testing of pleural fluid and pleural tissue are recommended 97.60%
Patients with Suspected Active Tuberculosis For acid-fast staining negative patients, at least one respiratory specimen should undergo M. tuberculosis nucleic acid testing; for acid-fast staining positive patients, nucleic acid amplification technology can differentiate between M. tuberculosis and non-tuberculous mycobacteria 94.00%

 

Question 1: Is Mycobacterium tuberculosis nucleic acid testing necessary?

Nucleic acid amplification technology can serve as a rapid diagnostic method for M. tuberculosis. The CDC recommends at least one respiratory specimen M. tuberculosis nucleic acid test for suspected active tuberculosis patients with negative acid-fast staining. For acid-fast staining positive patients, nucleic acid amplification technology can help differentiate between M. tuberculosis and non-tuberculous mycobacteria.

Question 2: How to choose between TB-specific PCR or tNGS/mNGS for M. tuberculosis testing?

TB-specific PCR and tNGS/mNGS have similar diagnostic capabilities for M. tuberculosis. Compared to traditional methods, tNGS/mNGS shows more advantages than combined TB-specific PCR testing, significantly improving pathological diagnosis of tuberculosis and other pathogens. tNGS testing can provide support for tuberculosis drug resistance genes.

 

VI. Immunocompromised Patients with Pulmonary Infections

 

✧ Key Recommendation: Concurrent blood and bronchoalveolar lavage fluid mNGS is recommended, with tissue specimen mNGS when necessary.

✧ Consensus Rate: 97.6%

 

Question 1: Is BALF nucleic acid testing recommended for immunocompromised hosts?

For acid-fast staining positive patients, nucleic acid amplification technology can help differentiate M. tuberculosis from non-tuberculous mycobacteria. While initiating empirical treatment, early bronchoscopy and bronchoalveolar lavage should be performed for pathogen detection, including PCR or mNGS, to minimize the impact of initial empirical treatment on pathological test results.

Other microbiological examinations of BALF samples depend on the patient's clinical presentation, imaging features, and risk factors for specific pathogen infections, including bacterial smear and culture, fungal smear and culture, cytological analysis, and histopathology.

Question 2: What techniques should be used when fungal infection is suspected in immunocompromised hosts?

For patients at high risk of invasive fungal infections, fungal fluorescence staining, fungal culture, serology, molecular biology, and even histopathology testing are recommended. Particularly, combining fungal fluorescence staining, 1,3-β-D-glucan test, cryptococcal capsular polysaccharide antigen, blood and BALF galactomannan test with nucleic acid detection can improve fungal detection rates. mNGS can be performed for patients with available histopathological specimens. For suspected Pneumocystis infection, respiratory specimen hexamine silver staining or PCR combined with serum G test can be used for definitive diagnosis. BALF and blood specimen mNGS also aids diagnosis, but positive PCR or mNGS results for Pneumocystis cannot distinguish between colonization and infection, requiring comprehensive clinical judgment.

Question 3: Is blood specimen mNGS necessary for immunocompromised hosts with pulmonary infections?

Immunocompromised hosts with pulmonary infections have a 5-10% chance of concurrent bacteremia, significantly higher than immunocompetent hosts. For such patients with severe CAP, concurrent BALF and blood specimen mNGS can help identify potential pathogenic bacteria. Blood specimen mNGS has slightly higher specificity but lower sensitivity than BALF.

 

Experts point out: "Different pathogen diagnostic methods each have their own applicable scenarios and prerequisites. The selection of suitable testing methods should be based on patient population, disease characteristics, suspected pathogen types, and local laboratory capabilities."

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