The first case was a 58-year-old male admitted on March 19, 2020, due to abdominal pain. He had mild hypertension and dyslipemia. On physical examination, he was haemodynamically stable, with 38.5ºC fever and with a hard mass in the right iliac fossa without peritonitis. Blood tests revealed haemoglobin 11.2 g/dL, normal glucose, platelet and leucocyte counts, normal renal function, C-reactive protein (CRP) 43.3 mg/L, international normalized ratio (INR) 1.3, and D-dimer 1460 ng/mL. Basal arterial blood gas (21%) showed the following values: pH 7.41, pCO2 38 mmHg, pO2 71 mmHg, HCO3 24.1 mmHg, Sat. O2 94.2%.
An abdominal computed tomography (CT) scan showed thickened walls in the ascending colon and caecum extending to the terminal ileum and appendix that suggested a neoformative process. Several enlarged lymphadenopathies were also found (Fig. 2A). A positron emission tomography (PET) scan was performed with a staging intention, which described an intensely hypermetabolic mass with standard uptake value (SUV) max31 in the caecum, and hypermetabolic lymphadenopathies in the mesenteric area. In addition, PET scan reported a poorly defined pulmonary uptake area in the right upper and middle lobes and in the left apical region (SUVmax7.7). Chest X-ray showed local patchy shadowing in the right middle lobe. All these pulmonary findings were suggestive of SARS-CoV-2 pneumonia, which was confirmed by a nasopharyngeal smear polymerase chain reaction (PCR) test. Treatment with oxygen therapy (high-flow mask, venturi type at 28% at 5 litres) and pharmacological treatment for COVID-19 were instituted with hydroxychloroquine 200 mg/12 h orally (PO), ceftriaxone 1 g/24 h intravenous (IV), azithromycin 500 mg/24 h PO and Lopinavir/Ritonavir 400 mg/100 mg/12 h PO. The treatment schedule did not include any corticosteroid agents at any time during hospital admission. The patient underwent clinical recovery, disappearance of fever, frank reduction of the abdominal mass (Fig. 2B), blood test normalization and pulmonary infiltrate improvement. After discharge, a colonoscopy was performed on April 15, 2020. An inflammatory mass occupying most of the caecal circumference was found, and the ileocecal valve was stenotic, preventing passage to the ileum (Fig. 2C). Multiple biopsies of the ascending colon were taken, showing, on light microscopy, abundant granulation tissue with lymphoplasmocytic inflammatory infiltrates and macrophages without granuloma formation. Immunohistochemical staining ruled out the presence of either an infiltrative epithelial neoplasm or a lymphoproliferative process (Fig. 2D). Cytomegalovirus (CMV) immunostaining was negative. PCR performed on formalin-fixed/paraffin-embedded (FFPE) tissue to detect atypical mycobacterial infection and SARS-CoV-2 were also negative.
The rapid improvement in parallel with COVID-19 recovery raised the suspicion of intestinal inflammation induced by SARS-CoV-2. In order to demonstrate this, TEM of the colon tissue, retrieved from the FFPE block, was performed. On ultrastructural study, viral particles characteristic of coronavirus were found. They showed the typical outer surface with several spikes all around them; rarely were more than ten found per cell, and they were well apart from each other. They were found in the cytoplasm of endothelial cells, from capillaries and arterioles in the involved mucosa. An electron-lucid area around some of these particles made them quite visible at low power. They were usually situated in the centre of an irregular and apparently empty space resembling rough endoplasmic reticulum. This ‘halo’ apparently prevented direct contact of the viral particles with cytoplasmic organelles (Fig. 2E, F).
During the next 3 months the patient remained afebrile and in overall good condition, but he presented self-limited episodes of abdominal pain. Two more CT scans and colonoscopies showed progressive decrease in the size of the inflammatory mass and of the lymph nodes, but persistence of predominantly fibrotic stenosis of the terminal ileum including the ileocecal valve. Ileocaecal resection was indicated that was performed on August 12, 2020 (Fig. 3A). Microscopic examination of a surgical specimen showed infiltration of the caecum and ileum by a neoplastic lesion with a diffuse growth pattern, formed by atypical, medium to large-sized lymphocytes, with scant cytoplasm, irregular nuclei, vesicular chromatin and conspicuous nucleoli. These morphological features were suggestive of lymphoma. Immunohistochemistry study was performed, showing expression for B-Cell markers (CD20, CD79a), as well as expression for bcl2, CD10, bcl6, LMO2, CD21, MUM1, and c-MYC in the neoplastic cells. Staining for CD23 ruled out the presence of a dendritic cell network. A high proliferative index (Ki67) was observed (90% in the hotspot areas). ISH technique for Epstein-Barr virus was negative. The tumour was classified as a diffuse large B-cell lymphoma (DLBCL), germinal center subtype (according to Hans’ algorithm) (Fig. 3B–D). No component of low-grade lymphoma was identified. Rearrangements of MYC, BCL2 and BCL6 genes were studied by FISH, detecting translocations in BCL2 and BCL6, but not in MYC. TEM of the lymphoma showed very scanty but convincing inclusions. In the endothelial cells of the lymphoma blood vessels, after very careful study, particles with coronavirus morphology were detected (Fig. 3E, F). Finally, an RNA in situ hybridization (RNA-ISH) assay of lymphoma and endothelial cells with SARS-CoV-2 was performed with negative results (Fig. 4).
The second case was a 38-year-old male patient, without relevant medical history, was admitted on March 18, 2020 for fever, dry cough, dyspnoea, and non-bloody diarrhoea. On physical examination he was haemodynamically stable and he had fever (39.2ºC) and bilateral rhoncus. Initial blood analysis showed normal haemogram, normal glucose, proteins, renal and liver function, INR 1.2, and D-Dimer 217 ng/mL. Arterial blood gas analysis performed with a high-flow mask (50% venturi type at 10 litres) showed pH 7.45, pCO2 21 mmHg, pO2 95 mmHg, HCO3 19.7 mmHg, and Sat.O2 97%. Chest X-ray showed right peripheral patchy shadowing due to COVID-19 (Fig. 5A) that was confirmed by positive SARS-CoV-2 nasopharyngeal PCR.
Treatment was started with ceftriaxone 1 g/24 h IV, azithromycin 500 mg/24 h PO, lopinavir/ritonavir 400 mg/100 mg/12 h PO, hydroxychloroquine 200 mg/12 h PO, Interferonβ1b 0.25 mg/mL subcutaneously every 48 h and dexamethasone 20 mg/24 IV. The patient was transferred to the intensive care unit (ICU) due to acute respiratory failure for orotracheal intubation and norepinephrine perfusion. He developed acute renal failure with oliguria of multifactorial cause (hypotension, diarrhoea, and rhabdomyolysis). After 1-month, mechanical ventilation was withdrawn and he was transferred to a conventional ward where oral diet was restarted with good tolerance. Control chest X-ray showed almost complete disappearance of alveolar infiltrates. On May 6, 2020, the patient suddenly presented abdominal pain, predominantly in the right hemiabdomen with signs of peritonitis, diaphoresis, and fever (38.5ºC). His status worsened, with haemodynamic instability, and respiratory and renal failure. An abdominal CT scan was performed, showing a stenosis in the transverse colon and a marked thickening of the right colon wall, surrounded by fat stranding (Fig. 5B). An emergency right hemicolectomy with terminal ileostomy was performed. Gross examination showed the colonic mucosa to have a ground cobblestone appearance, focal ulceration, and deposition of fibrinoid material. Histologically, mucosal necrosis, submucosal oedema, and haemorrhage were seen. The transmural ulcer showed granulation tissue, chronic inflammation, fibrosis, and steatonecrosis consistent with intestinal ischaemia. The adjacent mucosa showed regenerative changes, distorted architectural pattern, and mucinous hyperplasia. There was no evidence of CMV infection, nor of an infiltrative lesion (Fig. 5C, D). PCR for Clostridium difficile (CD) was positive in colon samples. A TEM of the colon tissue was performed, showing viral particles with a morphologic distribution essentially similar to those of case 1 (Fig. 5E, F).
After surgery, the patient was readmitted to the ICU with septic shock and multiorgan failure. Antibiotic treatment was started with wide antibiotic coverage (meropenem 500 mg/8 h IV + linezolid 600 mg/12 h IV + metronidazole 500 mg/8 h IV + Vancomycin 125 mg/6 h via nasogastric route + rectal vancomycin enemas). Norepinephrine, dobutamine, and hydrocortisone treatment were administered for 96 h. Finally, the patient was extubated, and then transferred to a conventional ward to continue rehabilitation until June 4, 2020, when he was discharged from hospital. The final diagnoses were bilateral pneumonia secondary to COVID-19 and colonic ischaemia with endothelial damage related to SARS-CoV-2 (Fig. 6).
