Study subjects
The present study was a critical re-evaluation of deceased patients by an expert team of specialists (pulmonologists, radiologists, and pathologists) routinely involved in multidisciplinary meetings of mainly DPLD [8] but also with robust experience in COVID-19 diagnosis and management [9, 10]. We retrospectively studied 27 patients who consecutively died from March to May 2020 in our hospital of severe COVID-19 pneumonia as the main diagnosis made by non-pulmonologists (i.e. emergency room clinicians, general practitioners, specialists in infectious diseases, and anaesthesiologists).
At autopsy, the whole lungs were macroscopically examined, and a small sample was taken from the most representative area of lung injury. The sample was in part preserved in RNA later and processed for molecular analyses (see below for molecular processing details) and in part fixed in formalin for routine histology. Pathological features suggestive of COVID-19 pneumonia (alveolar injury as well as vascular lesions) were quantitatively described using a scoring system, as previously reported [10]. Other associated lesions (neoplasia, infectious diseases, aspiration pneumonia, etc.) were also reported. To confirm the pathological diagnosis of COVID-19 pneumonia, the fragment preserved in RNA later was also processed by real time reverse transciptase-polymerase chain reaction (RT-PCR) for SARS-CoV2 [SARS-CoV-2 (2019-nCoV) Centers for Disease Control and Prevention (CDC) Emergency Use Authorized (EUA) Authorized qPCR probe assay primer/probe mix]. An additional fragment was analyzed by culture, as previously described [11]. In order to define the levels of certainty for a diagnosis of COVID-19 pneumonia, two expert pathologists (F.C., F.P.) scored all cases independently and blinded to clinical and autopsy data. Based on morphological/virologic evaluation, four distinct levels of diagnostic certainty were defined: 1) Definite COVID-19 pneumonia: all lung samples showing lesions typical of COVID-19 pneumonia (vascular injury and/or diffuse alveolar damage/organizing pneumonia), confirmed SARS-CoV2 lung positivity (both molecular and culture), without other lesions suggestive of alternative diagnoses, 2) Probable COVID-19 pneumonia: lung samples displaying mainly features of COVID-19 pneumonia (+/- lung SARS-CoV2 infection) with other associated lesions (i.e. foci of bacterial infection, 3) Possible COVID-19 pneumonia: lung samples showing only focal changes of COVID-19 pneumonia (+/- lung SARS-CoV2 infection, etc.) with more extensive features consistent with alternative diagnoses (i.e., lung cancer/metastasis, etc.) 4) Non-COVID-19 pneumonia: lung samples not showing any typical lesions, no evidence of SARS-CoV2 infection, and no presence of features consistent with alternative diagnoses.
During the autopsy, additional fragments were sampled from both lungs (at least 20 samples for each case) and systematically analyzed, as previously described [12].
Clinical evaluation was performed by two experienced pulmonologists (P.S., E.B.) based on the following data: past and recent medical history including comorbidities, respiratory and systemic signs and symptoms (type and duration) before hospital admission, imaging, laboratory findings, gas exchange values (FiO2, pO2, and pO2/FiO2) and their changes during hospitalization, and oxygen supplementation. Based on this data, patients were classified as follows: 1) Definite COVID-19 pneumonia: clinical findings typical of COVID-19 such as severe acute respiratory illness (i.e., fever, cough, shortness of breath, hypoxemia) in the absence of an alternative diagnosis that could explain the clinical presentation [13, 14]; 2) Probable COVID-19 pneumonia: features of COVID-19 pneumonia associated with findings suggestive of alternative diagnoses (e.g., pleural effusion, clinical and laboratory findings in keeping with heart failure, or signs of bacterial pneumonia), 3) Possible COVID-19 pneumonia: features of COVID-19 pneumonia associated with prevalent findings consistent with alternative etiologies (e.g., lung cancer, pulmonary metastases, pulmonary edema, heart failure), 4) Non-COVID-19 pneumonia: absence of typical signs/symptoms and laboratory findings of COVID-19 pneumonia in the presence of features consistent with alternative diagnoses (e.g., neoplasm, interstitial lung disease, ischemic heart disease, pulmonary edema).
With regard to the radiological assessment, all chest X-rays and, when available, chest computed tomography (CT) images were assessed by two expert thoracic radiologists (C.G., A.F.). According to the radiological findings, patients were classified as follows: 1) Definite COVID 19 pneumonia: typical findings of COVID-19 pneumonia, such as bilateral ground-glass opacities and/or consolidations [15], without any signs of alternative diagnoses; 2) Probable COVID-19 pneumonia: features of COVID-19 pneumonia associated with abnormalities such as pleural effusion, cardiomegaly, or Kerley B lines suggestive of cardiac failure, or lobar consolidation suggestive of bacterial pneumonia; 3) Possible COVID-19 pneumonia: features of COVID-19 pneumonia associated with predominant findings of alternative diagnoses (e.g., unilateral pulmonary lesions due to lung cancer, pulmonary bilateral metastatic nodules), 4) Non-COVID-19 pneumonia: no typical signs of COVID-19 with features suggestive of alternative diagnoses (e.g., unilateral pulmonary lesions due to lung cancer, reticular changes secondary to interstitial lung disease).
Data regarding demographics, smoking history, symptoms, comorbidities, treatment, disease duration, serology, radiological and pathological findings were included in a dedicated database in REDCap. Informed consent was granted by a relative/legal representative of each deceased patient. The study was approved by the local clinical institutional review Board.
Study design
To evaluate whether the contribution of pulmonologists, radiologists, and pathologists individually and/or in combination, could change the diagnosis originally made by non-pulmonologists, we planned a three-step process, modifying the methodology previously used in the evaluation of patients with DPLD [16].
Briefly, in the first step, two pulmonologists (P.S., E.B.) and two radiologists (C.G., A.F.) independently reviewed clinical and radiological data for each patient, without pathological data, and recorded their individual diagnoses and confidence levels. In the second step, pulmonologists and radiologists discussed their diagnosis and again recorded their individual or shared (in case of disagreement) confidence level. During the third step, pathologists entered the arena and reported the pathological diagnosis performed on a single lung fragment. The final diagnosis derived from the whole lung examination and full organ autopsy and was considered the diagnostic gold standard. Virtual meetings via the Zoom platform were set to allow pulmonologists, radiologists, and pathologists to discuss their interpretation with mutual collaboration (Fig. 1).