key: cord-0743247-wz61p3vn authors: Schwab, Kristin; Hamidi, Sepehr; Chung, Augustine; Lim, Raymond J; Khanlou, Negar; Hoesterey, Daniel; Dumitras, Camelia; Adeyiga, Oladunni B; Phan-Tang, Michelle; Wang, Tisha S; Saggar, Rajan; Goldstein, Jeffrey; Belperio, John A; Dubinett, Steven M; Kim, Jocelyn T; Salehi-Rad, Ramin title: Occult colonic perforation in a patient with COVID-19 following interleukin-6 receptor antagonist therapy date: 2020-09-12 journal: Open Forum Infect Dis DOI: 10.1093/ofid/ofaa424 sha: a7faa723c11cbccd427c29593e0b1359e4849052 doc_id: 743247 cord_uid: wz61p3vn Interleukin-6 blockade (IL-6) has become a focus of therapeutic investigation for the coronavirus-2019 (COVID-19). We report a case of a 34 year-old with COVID-19 pneumonia receiving an IL-6 receptor antagonist (IL-6Ra) who developed spontaneous colonic perforation. This perforation occurred despite a benign abdominal exam and in the absence of other known risk factors associated with colonic perforation. Examination of the colon by electron microscopy revealed numerous intact SARS-CoV-2 virions abutting the microvilli of the colonic mucosa. Multiplex immunofluorescent staining revealed the presence of the SARS-CoV-2 spike protein on the brush borders of colonic enterocytes which expressed angiotensin-converting enzyme 2. However, no viral particles were observed within the enterocytes to suggest direct viral injury as the cause of colonic perforation. These data and absence of known risk factors for spontaneous colonic perforation implicate IL-6Ra therapy as the potential mediator of colonic injury in this case. Furthermore, this report provides the first in situ visual evidence of the virus in the colon of a patient presenting with colonic perforation adding to growing evidence that intact infectious virus can be present in the stool. A c c e p t e d M a n u s c r i p t 4 Introduction: The coronavirus disease 2019 pandemic caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a global threat to public health. Although the pathophysiology of COVID-19 has not been fully elucidated, direct viral injury and imbalanced host immune responses have been implicated in the immunopathogenesis of the disease 1 . In particular, a hyperinflammatory phenotype with elevated serum concentrations of many cytokines including interleukin-6 (IL-6) has been identified, which correlates with adverse clinical outcomes 2 . As a result, IL-6 receptor antagonist (IL-6Ra) therapy has been investigated in COVID-19 patients, with over 80 clinical trials currently underway 3 . Although the lungs are the primary organ targeted by SARS-CoV-2, gastrointestinal manifestations of the disease are common [4] [5] [6] . Here, we report a case of a patient with COVID-19 pneumonia receiving IL-6Ra who developed colonic perforation. We present the first electron microscopy (EM) image of SARS-CoV-2 virions in a human colon. Though the virus particles were not seen in enterocytes, to our knowledge this report is the first in situ visual evidence of the virus abutting the microvilli of the colonic mucosa. A 34-year-old man presented to an outside hospital with 2 days of fevers, chills, and dyspnea ( Figure S1 ). The patient was previously healthy with no medical comorbidities aside from obesity (body mass index of 36). He was diagnosed with COVID-19 pneumonia and admitted on 4 liters of supplemental oxygen. On hospital day (HD) 2, he had progressive hypoxemic respiratory failure requiring intubation. Proning and neuromuscular blockade (NMB) were initiated for severe acute A c c e p t e d M a n u s c r i p t 5 respiratory distress syndrome (ARDS), and he was transferred to our hospital on HD 6 for consideration of extracorporeal membrane oxygenation. On arrival, he received an IL-6Ra on HD 7 and HD 13 as part of a clinical trial. Lung protective ventilation, proning, and NMB were continued. The patient's procalcitonin and C-reactive protein (CRP) levels decreased with time, but he continued to have low-grade fevers on broad-spectrum antibiotic therapy. Given his otherwise negative infectious workup, the fever was attributed to COVID-19. The patient had gradual improvement of his PaO2/FiO2 ratio, and his cardiac, renal, and liver function remained intact. Despite high sedation and analgesia requirements, he tolerated enteral tube feedings with regular daily bowel movements. On HD 20, his fever increased to 41.0 o C, accompanied by an increasing white blood cell count. Figure 1D) . Intracellular viral particles were not observed. We report a case of spontaneous colonic perforation in a patient with COVID-19 receiving IL-6Ra therapy. The differential diagnosis for colonic perforation in this critically ill patient is broad, including colonic pseudo-obstruction, diverticulitis, ischemic colitis, spontaneous stercoral or idiopathic perforation, and virus-or medication-induced colonic injury. Although our patient required opiates for sedation, he had regular bowel movements with no clinical manifestations of constipation, obstruction or pseudo-obstruction. Additionally, in contrast to stercoral perforations that commonly cause an ovoid pattern of injury in the sigmoid colon or rectum with evidence of impacted stool and ischemic necrosis of the surrounding mucosa 8 , our patient had a linear tear in the cecum without feculent material. An ischemic insult was also an unlikely cause of perforation in this patient, as he was on minimal to no vasopressors throughout his hospitalization leading up to the perforation with no signs of hypoperfusion in other organs and no features of ischemia on microscopic examination. Furthermore, the patient did not receive glucocorticoids or non-steroidal anti-inflammatory medications, and had no pathological evidence of diverticuli, tumor, or appendicitis. Intestinal infarct or perforation can also occur due to thromboses of the intestinal vascular bed, and COVID-19-associated coagulopathy can result in venous, arterial, and microvascular thrombotic events [9] [10] [11] . However, we did not see histopathological evidence of thrombosis in the adjacent colonic vasculature to indicate that this patient's intestinal perforation occurred due to thrombosis. Although ACE2 expression has been identified in many organs, a recent study revealed that coexpression of ACE2 and TMPRSS2 is highly enriched on type II pneumocytes in the lung, nasal goblet secretory cells, and absorptive enterocytes in the terminal ileum, but not colonic enterocytes 13 . While previous reports have implicated SARS-CoV-2 as a cause of colitis and enteritis, evidence for natural infection of human colonic enterocytes by SARS-CoV-2 has not been reported 4, 14 , although colonic organoids can be infected in vitro 15, 16 . Similarly, high concentrations of viral genome as determined by polymerase chain reaction have been detected in stool samples, but efforts to culture the virus from these samples have been largely unsuccessful 17 , except in one isolated case 16 , thus suggesting viral particles in the gastrointestinal tract may not be intact in most cases. To assess if direct viral injury contributed to colonic perforation, we evaluated the resected bowel for the presence of SARS-CoV-2 by MIF and EM. Although MIF revealed limited staining of ACE2 on colonic enterocytes, which was confined to the brush borders 7 , we did not detect any viral particles within the enterocytes by EM and MIF to suggest direct viral injury to the mucosa. Surprisingly, we observed numerous spheroidal viral particles abutting the microvilli of the colonic mucosa on EM, which was corroborated by MIF staining of the viral S protein. We also found numerous viral particles abutting the microvillii of adjacent healthy tissue (data not shown), which is consistent with another recent report in which SARS-CoV-2 virions were detected in healthy rectal tissue of a patient with COVID-19 18 . To our knowledge, this is the first reported evidence of in situ SARS-CoV-2 virions on colonic enterocytes at the site of intestinal injury. Although we cannot definitively rule out direct viral injury as the cause, exclusion of other potential etiologies suggests IL-6Ra as a possible cause of spontaneous colonic perforation in this patient. Colonic perforation is a known complication of IL-6 inhibition with tocilizumab and sarilumab 19,20 . A c c e p t e d M a n u s c r i p t 8 This is partly due to the anti-apoptotic and proliferative properties of IL-6 signaling in the colon, which prolongs enterocyte lifespan and stimulates intestinal epithelial regeneration at the onset of colonic injury [21] [22] [23] . IL-6 inhibition also results in decreased levels of vascular endothelial growth factor (VEGF), which plays a key role in maintaining vascular integrity via angiogenesis 24 . The importance of VEGF in maintaining mucosal integrity is further suggested by the fact that VEGF-inhibitors are associated with gastrointestinal perforation 25 . Of note, the reported IL-6Ra-associated perforations often present silently without abdominal pain 26 , as was the case in our patient. Furthermore, while the literature suggests that these perforations occur in patients with other predisposing factors such as diverticulitis, glucocorticoid use, or older age, the increased risk of gastrointestinal perforation with IL-6Ra therapy remains after multivariate adjustment for these factors 26, 27 . This is particularly relevant in our case, as our patient had none of these predisposing factors. These data suggest that the gastrointestinal tract is likely subject to frequent erosions that must be countered by cell proliferation and vascular regeneration, which rely partially on IL-6 signaling. Although the Food and Drug Association cautions against using tocilizumab and sarilumab in patients at-risk for gastrointestinal perforation, our report suggests that the at-risk population may be potentially larger 28, 29 . In summary, we report a case of colonic perforation in a patient with COVID-19 who received IL-6Ra therapy. Our patient's perforation occurred in the absence of known risk factors or clear evidence of direct viral injury to the colon, implicating IL-6Ra therapy as the potential mediator of colonic injury. This case also represents the first reported evidence of in situ SARS-CoV-2 virions in the colon of a patient presenting with colonic perforation. Additional studies in larger cohorts of COVID-19 patients, such as tissue examinations from biopsies or autopsies, are needed to validate our findings and elucidate the role of the colon as a potential target and source of SARS-CoV-2 replication. 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