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1.
Clinica chimica acta|international journal of clinical chemistry ; 2022.
Article in English | EuropePMC | ID: covidwho-1837538

ABSTRACT

Background SARS-CoV-2 is an RNA virus that causes primarily causes respiratory disease;however, infection of other tissue has been reported. Evaluation of SARS-CoV-2 in tissue specimens may increase understanding of SARS-CoV-2 pathobiology. Materials and Methods A qualitative test for detection of SARS-CoV-2 in formalin-fixed paraffin-embedded (FFPE) tissues was developed and validated using droplet digital PCR (ddPCR), which has a lower limit of detection than reverse transcription (RT)-qPCR. After extraction of total RNA from unstained FFPE tissue, SARS-CoV-2 nucleocapsid (N1, N2) target sequences were amplified and quantified, along with human RPP30 as a control using the Bio-Rad SARS-CoV-2 ddPCR kit. Results SARS-CoV-2 was detected in all 21 known positive samples and none of the 16 negative samples. As few as approximately 5 viral copies were reliably detected. Since January 2021, many tissue types have been clinically tested. Of the 195 clinical specimens, the positivity rate was 35% with placenta and fetal tissue showing the highest percentage of positive cases. Conclusion This sensitive FFPE-based assay has broad clinical utility with applications as diverse as pregnancy loss and evaluation of liver transplant rejection. This assay will aid in understanding atypical presentations of COVID-19 as well as long-term sequelae.

2.
Arch Pathol Lab Med ; 2022 Mar 23.
Article in English | MEDLINE | ID: covidwho-1761088

ABSTRACT

CONTEXT: Studies of lungs in patients with COVID-19 have focused on early findings. OBJECTIVE: To systematically study histopathologic, imaging features and presence of SARSCoV-2 RNA in lung tissue from patients in later stages of COVID-19. DESIGN: Autopsies, explants, surgical lung biopsies; and transbronchial, cryo, and needle biopsies were studied from patients with COVID-19, whose onset of symptoms/confirmed diagnosis was more than 28 days before the procedure. Available images were reviewed. Reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) for SARS-CoV-2 RNA was performed on lung tissue. RESULTS: Forty-four specimens (43 patients, median age 59.3 years, 26 [60.5%] male) showed features of acute lung injury (ALI) in 39 (88.6%), predominantly organizing pneumonia (OP) and diffuse alveolar damage (DAD), up to 298 days after onset of COVID-19. Fibrotic changes were found in 33 specimens (75%), most commonly fibrotic DAD (N=22) and cicatricial OP (N=12). Time between acquiring COVID-19 and specimen was shorter in patients with diffuse ALI (median 61.5 days) compared to patients with focal (140 days) or no ALI (130 days) (P=.009). Sixteen (of 20, 80%) SARS-CoV-2 RT-ddPCR tests were positive, up to 174 days after COVID-19 onset. Time between COVID-19 onset and most recent CT in patients with consolidation on imaging was shorter (median 43.0 days) versus patients without consolidation (87.5 days; P=.02). Reticulations were associated with longer time after COVID-19 onset to CT (median 82 days vs 23.5 days, P=.006). CONCLUSIONS: ALI and SARS-CoV-2 RNA can be detected in patients with COVID-19 for many months. ALI may evolve into fibrotic interstitial lung disease.

3.
Mayo Clin Proc ; 96(10): 2561-2575, 2021 10.
Article in English | MEDLINE | ID: covidwho-1521396

ABSTRACT

OBJECTIVE: To compare coronavirus disease 2019 (COVID-19) acute kidney injury (AKI) to sepsis-AKI (S-AKI). The morphology and transcriptomic and proteomic characteristics of autopsy kidneys were analyzed. PATIENTS AND METHODS: Individuals 18 years of age and older who died from COVID-19 and had an autopsy performed at Mayo Clinic between April 2020 to October 2020 were included. Morphological evaluation of the kidneys of 17 individuals with COVID-19 was performed. In a subset of seven COVID-19 cases with postmortem interval of less than or equal to 20 hours, ultrastructural and molecular characteristics (targeted transcriptome and proteomics analyses of tubulointerstitium) were evaluated. Molecular characteristics were compared with archived cases of S-AKI and nonsepsis causes of AKI. RESULTS: The spectrum of COVID-19 renal pathology included macrophage-dominant microvascular inflammation (glomerulitis and peritubular capillaritis), vascular dysfunction (peritubular capillary congestion and endothelial injury), and tubular injury with ultrastructural evidence of mitochondrial damage. Investigation of the spatial architecture using a novel imaging mass cytometry revealed enrichment of CD3+CD4+ T cells in close proximity to antigen-presenting cells, and macrophage-enriched glomerular and interstitial infiltrates, suggesting an innate and adaptive immune tissue response. Coronavirus disease 2019 AKI and S-AKI, as compared to nonseptic AKI, had an enrichment of transcriptional pathways involved in inflammation (apoptosis, autophagy, major histocompatibility complex class I and II, and type 1 T helper cell differentiation). Proteomic pathway analysis showed that COVID-19 AKI and to a lesser extent S-AKI were enriched in necroptosis and sirtuin-signaling pathways, both involved in regulatory response to inflammation. Upregulation of the ceramide-signaling pathway and downregulation of oxidative phosphorylation in COVID-19 AKI were noted. CONCLUSION: This data highlights the similarities between S-AKI and COVID-19 AKI and suggests that mitochondrial dysfunction may play a pivotal role in COVID-19 AKI. This data may allow the development of novel diagnostic and therapeutic targets.


Subject(s)
Acute Kidney Injury/pathology , COVID-19/pathology , Kidney/pathology , Sepsis/pathology , Acute Kidney Injury/virology , Adult , Autopsy , Humans , Kidney Tubules, Proximal/pathology , Male , Middle Aged , Sepsis/virology
4.
Archives of Pathology & Laboratory Medicine ; 145(7):785-796, 2021.
Article in English | ProQuest Central | ID: covidwho-1338028

ABSTRACT

* Context.-Small case series have evaluated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in formalin-fixed, paraffin-embedded tissue using reverse transcription-polymerase chain reaction, immunohistochemistry (IHC), and/or RNA in situ hybridization (RNAish). [...]acute respiratory distress syndrome has been found in 3.4% of patients with COVID-19 and 72% to 93% of patients with COVID-19 who died.1-3 Furthermore, we and others have shown that acute bronchopneumonia, diffuse alveolar damage (DAD), aspiration pneumonia, and thromboemboli are common findings in the lungs of these patients at autopsy.4-9 Although evidence suggests that COVID-19 also impacts other organs such as heart,9 liver,9 and kidneys,9,10 the virus has so far been identified with certainty only in the lungs and rarely the brain, heart, liver, kidney, placenta, and blood by electron microscopy, immunohistochemistry (IHC), in situ hybridization (ISH), and reverse transcription-polymerase chain reaction (RT-PCR).8'9'11-18 Infection with SARS-CoV-2 is usually identified by the detection of viral RNA using RT-PCR on nasopharyngeal or oropharyngeal swabs. Furthermore, viral cytopathic effects and inclusions observed in cytomegalovirus, herpes simplex virus, and adenovirus infections are not evident in SARS-CoV-2 infections. [...]a formalin-fixed, paraffin-embedded (FFPE) tissue-based test is needed to establish the diagnosis of COVID-19 and guide management of the patient. Potential regional or temporal heterogeneity in viral genomic sequence due to genetic selection or drift may play a similar role, impacting the ability of existing targeted PCR assays to detect the virus in specimens from patients with COVID-19. Because formalin fixation can impact the quality and integrity of nucleic acid, sensitivity for the detection of microorganisms by PCR or other techniques may also be limited.

5.
Arch Pathol Lab Med ; 145(7): 785-796, 2021 07 01.
Article in English | MEDLINE | ID: covidwho-1134421

ABSTRACT

CONTEXT.­: Small case series have evaluated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection in formalin-fixed, paraffin-embedded tissue using reverse transcription-polymerase chain reaction, immunohistochemistry (IHC), and/or RNA in situ hybridization (RNAish). OBJECTIVE.­: To compare droplet digital polymerase chain reaction, IHC, and RNAish to detect SARS-CoV-2 in formalin-fixed, paraffin-embedded tissue in a large series of lung specimens from coronavirus disease 2019 (COVID-19) patients. DESIGN.­: Droplet digital polymerase chain reaction and RNAish used commercially available probes; IHC used clone 1A9. Twenty-six autopsies of COVID-19 patients with formalin-fixed, paraffin-embedded tissue blocks of 62 lung specimens, 22 heart specimens, 2 brain specimens, and 1 liver, and 1 umbilical cord were included. Control cases included 9 autopsy lungs from patients with other infections/inflammation and virus-infected tissue or cell lines. RESULTS.­: Droplet digital polymerase chain reaction had the highest sensitivity for SARS-CoV-2 (96%) when compared with IHC (31%) and RNAish (36%). All 3 tests had a specificity of 100%. Agreement between droplet digital polymerase chain reaction and IHC or RNAish was fair (κ = 0.23 and κ = 0.35, respectively). Agreement between IHC and in situ hybridization was substantial (κ = 0.75). Interobserver reliability was almost perfect for IHC (κ = 0.91) and fair to moderate for RNAish (κ = 0.38-0.59). Lung tissues from patients who died earlier after onset of symptoms revealed higher copy numbers by droplet digital polymerase chain reaction (P = .03, Pearson correlation = -0.65) and were more likely to be positive by RNAish (P = .02) than lungs from patients who died later. We identified SARS-CoV-2 in hyaline membranes, in pneumocytes, and rarely in respiratory epithelium. Droplet digital polymerase chain reaction showed low copy numbers in 7 autopsy hearts from ProteoGenex Inc. All other extrapulmonary tissues were negative. CONCLUSIONS.­: Droplet digital polymerase chain reaction was the most sensitive and highly specific test to identify SARS-CoV-2 in lung specimens from COVID-19 patients.


Subject(s)
COVID-19 Testing/methods , COVID-19/diagnosis , Immunohistochemistry , In Situ Hybridization/methods , Lung/virology , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/isolation & purification , Adult , Aged , Aged, 80 and over , COVID-19/virology , Female , Humans , Male , Middle Aged , Observer Variation , Prospective Studies , RNA, Viral/isolation & purification , Reproducibility of Results , SARS-CoV-2/genetics , Sensitivity and Specificity
6.
Arch Pathol Lab Med ; 145(1): 11-21, 2021 01 01.
Article in English | MEDLINE | ID: covidwho-1067935

ABSTRACT

CONTEXT.­: Respiratory failure appears to be the ultimate mechanism of death in most patients with severe coronavirus disease 2019 (COVID-19) infection. Studies of postmortem COVID-19 lungs largely report diffuse alveolar damage and capillary fibrin thrombi, but we have also observed other patterns. OBJECTIVE.­: To report demographic and radiographic features along with macroscopic, microscopic, and microbiologic postmortem lung findings in patients with COVID-19 infections. DESIGN.­: Patients with confirmed COVID-19 infection and postmortem examination (March 2020-May 2020) were included. Clinical findings were abstracted from medical records. Lungs were microscopically reviewed independently by 4 thoracic pathologists. Imaging studies were reviewed by a thoracic radiologist. RESULTS.­: Eight patients (7 men, 87.5%; median age, 79 years; range, 69-96 years) died within a median of 17 days (range, 6-100 days) from onset of symptoms. The median lung weight was 1220 g (range, 960-1760 g); consolidations were found in 5 patients (62.5%) and gross thromboemboli were noted in 1 patient (12.5%). Histologically, all patients had acute bronchopneumonia; 6 patients (75%) also had diffuse alveolar damage. Two patients (25%) had aspiration pneumonia in addition. Thromboemboli, usually scattered and rare, were identified in 5 patients (62.5%) in small vessels and in 2 of these patients also in pulmonary arteries. Four patients (50%) had perivascular chronic inflammation. Postmortem bacterial lung cultures were positive in 4 patients (50%). Imaging studies (available in 4 patients) were typical (n = 2, 50%), indeterminate (n = 1, 25%), or negative (n = 1, 25%) for COVID-19 infection. CONCLUSIONS.­: Our study shows that patients infected with COVID-19 not only have diffuse alveolar damage but also commonly have acute bronchopneumonia and aspiration pneumonia. These findings are important for management of these patients.


Subject(s)
COVID-19/pathology , Lung/pathology , Aged , Aged, 80 and over , Autopsy , Bronchopneumonia/pathology , COVID-19/diagnostic imaging , COVID-19/mortality , Fatal Outcome , Female , Humans , Lung/diagnostic imaging , Male , Minnesota/epidemiology , Pandemics , Pneumonia, Aspiration/pathology , Pulmonary Alveoli/pathology , Pulmonary Embolism/pathology , SARS-CoV-2 , Tomography, X-Ray Computed
7.
Circulation ; 143(3): 230-243, 2021 01 19.
Article in English | MEDLINE | ID: covidwho-1039948

ABSTRACT

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its resultant clinical presentation, coronavirus disease 2019 (COVID-19), is an emergent cause of mortality worldwide. Cardiac complications secondary to this infection are common; however, the underlying mechanisms of such remain unclear. A detailed cardiac evaluation of a series of individuals with COVID-19 undergoing postmortem evaluation is provided, with 4 aims: (1) describe the pathological spectrum of the myocardium; (2) compare with an alternate viral illness; (3) investigate angiotensin-converting enzyme 2 expression; and (4) provide the first description of the cardiac findings in patients with cleared infection. METHODS: Study cases were identified from institutional files and included COVID-19 (n=15: 12 active, 3 cleared), influenza A/B (n=6), and nonvirally mediated deaths (n=6). Salient information was abstracted from the medical record. Light microscopic findings were recorded. An angiotensin-converting enzyme 2 immunohistochemical H-score was compared across cases. Viral detection encompassed SARS-CoV-2 immunohistochemistry, ultrastructural examination, and droplet digital polymerase chain reaction. RESULTS: Male sex was more common in the COVID-19 group (P=0.05). Nonocclusive fibrin microthrombi (without ischemic injury) were identified in 16 cases (12 COVID-19, 2 influenza, and 2 controls) and were more common in the active COVID-19 cohort (P=0.006). Four active COVID-19 cases showed focal myocarditis, whereas 1 case of cleared COVID-19 showed extensive disease. Arteriolar angiotensin-converting enzyme 2 endothelial expression was lower in COVID-19 cases than in controls (P=0.004). Angiotensin-converting enzyme 2 myocardial expression did not differ by disease category, sex, age, or number of patient comorbidities (P=0.69, P=1.00, P=0.46, P=0.65, respectively). SARS-CoV-2 immunohistochemistry showed nonspecific staining, whereas ultrastructural examination and droplet digital polymerase chain reaction were negative for viral presence. Four patients (26.7%) with COVID-19 had underlying cardiac amyloidosis. Cases with cleared infection had variable presentations. CONCLUSIONS: This detailed histopathologic, immunohistochemical, ultrastructural, and molecular cardiac series showed no definitive evidence of direct myocardial infection. COVID-19 cases frequently have cardiac fibrin microthrombi, without universal acute ischemic injury. Moreover, myocarditis is present in 33.3% of patients with active and cleared COVID-19 but is usually limited in extent. Histological features of resolved infection are variable. Cardiac amyloidosis may be an additional risk factor for severe disease.


Subject(s)
COVID-19 , Coronary Thrombosis , Fibrin/metabolism , Myocardium , SARS-CoV-2/metabolism , Adolescent , Adult , Aged , Aged, 80 and over , Angiotensin-Converting Enzyme 2/biosynthesis , COVID-19/metabolism , COVID-19/mortality , COVID-19/pathology , Child , Child, Preschool , Coronary Thrombosis/metabolism , Coronary Thrombosis/mortality , Coronary Thrombosis/pathology , Female , Gene Expression Regulation, Enzymologic , Humans , Immunohistochemistry , Infant , Male , Middle Aged , Myocardium/metabolism , Myocardium/pathology
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