Персистирующая инфекция

chuka_lis — 15.01.2022 Статья от ученых из Института Здоровья США, в которой разбираются данные по 44 аутопсиям тех, кто болел ковидом (большинство умерло от ковида, а не с, и у большинства была тяжелая форма болезни, включая и критическую), и у кого была разная продолжительность инфекции до смерти (они выделили группу раннюю, до 2 недель, среднюю, до месяца, и позднюю, более 31 дня). Ткани умерших быстро, интенсивно и экстенсивно проверялись на гистологические изменения, тип пораженных клеток, и наличие живого вируса, его частей и генетических следов (и в нескольких случаях- ген. модификаций).
Авторы приходят к заключению, что еще на ранних стадиях заражения вирус идет не только в легкие, а и в другие органы и ткани (где его обнаружили после смерти). Причем даже у тех, кто имеет умеренную форму болезни (те, немногие, кто умер "с" ковидом).
В 79 из 85 тканевых локаций у них выделялся или обнаруживался вирус. Больше всего вируса было в респираторной системе (и легких), особенно на ранних стадиях болезни. Но так же вирус был обнаружен в тканях сердца, жкт, лимофидных органов,  мозга, почек, печени, эндокринных желез, жировой ткани, кожи, периферической нервной системы, кожи, глаз, репродуктивной системы.
На ранних стадиях болезни,  если пациент умер, то вирус культивировался из всех  тканей, где обнаруживался, до 7 дня (после постановки диагноза). Однако, реплицироваться мог дольше- у некоторых до 2-3 месяцев, после диагноза, тк  в их тканях и клетках обнаруживалась анти-с-РНК (копя  соригинальной, необходимая для того, чтоб вирус мог воспроизвестись). Со временем после болезни, количество вирусного материала в тканях, преимущественно, уменьшалось. В одном из поздних случаях вирусная РНК обнаруживалась спустя 7 месяцев после ковида, в мозге. Авторы думают, что в некоторых случаях, могут формироваться неполноценные вирусные частицы, в результате борьбы, которые могут быть ответственны за нарушения в клетках.
Корнавирус, таким образом,  заражает весь организм, хотя его в других тканях значительно меньше (на порядки) чем  респираторной системе и в легких, и так же, нет обычно такого выраженного патологического воспаления, и повреждений такого уровня, как  в легких. Тем не менее, вирус там есть, и в некоторых случаях может оставаться там долго (персистировать, без особых мутаций)- и это может пояснять, отчасти, наличие лонгковида у некоторых переболевших ковидом.
Статья интересная, ниже выдержки, а таблицы и более подробностей по ссылке.
В общем- на мой взгляд, аутопсии показали, что как и у хомячков, на модели - вирус  после инфекции подавляет иммунитет и лезет везде, и размножается везде, на сколько успевает и может. Затем иммунитет "спохватывается", в основном, в воротах инфекции или в крови, и  дальше идет борьба по очищению от вируса, когда щепки летят, особенно в месте, где много вируса- тк там большой напор неспецифического иммунного ответа. Затем, спустя время, подключается и специфический и "дочищает" сам вирус и больные клетки,  где достанет (а может не везде). И порой случается, что вирус или вирусный материал остается кое-где (откуда не удается выкурить) - надолго, обеспечивая "вялотекующую" болезнь с разной симптоматикой- постковидный синдром .

COVID-19 is known to cause multi-organ dysfunction1-68 3 in acute infection, with 69 prolonged symptoms experienced by some patients, termed Post-Acute Sequelae of SARS70 CoV-2 (PASC)4-5. However, the burden of infection outside the respiratory tract and time to viral clearance is not well characterized, particularly in the brain3,6-14. We performed complete autopsies on 44 patients with COVID-19 to map and quantify SARS-CoV-2 distribution, replication, and cell-type specificity across the human body, including brain, from acute infection through over seven months following symptom onset. We show that SARS-CoV-2 is widely distributed, even among patients who died with asymptomatic to mild COVID-19, and that virus replication is present in multiple pulmonary and extrapulmonary tissues early in infection. Further, we detected persistent SARS-CoV-2 RNA in multiple anatomic sites, including regions throughout the brain, for up to 230 days following symptom onset. Despite extensive distribution of SARS-CoV-2 in the body, we observed a paucity of inflammation or direct viral cytopathology outside of the lungs. Our data prove that SARS-CoV-2 causes systemic infection and can persist in the body for months. To inform these pathogen-focused questions and to evaluate for the presence or absence of associated histopathology in matched tissue specimens, we performed extensive autopsies on a diverse population of 44 individuals who died from or with COVID-19 up to 230 days following initial symptom onset. The histopathology findings from our cohort were similar to those reported in other case series . All but five cases were considered to have died from COVID-19 , and, of these, 37 (94.5%) had either acute pneumonia or diffuse alveolar damage at the time of death. Phases of diffuse alveolar damage showed clear temporal associations, with the exudative phase seen mainly within the first three weeks of infection and the fibrosing phase not seen until after a month of infection Pulmonary thromboembolic complications, which were also likely related to SARS-CoV-2 infection, with or without infarction, were noted in 10 (23%) cases. Another finding likely related to SARS-CoV-2 infection included myocardial infiltrates in four cases, including one case of significant myocarditis16 (P3). Some of the cases of microscopic ischemia appeared to be associated with fibrin-platelet microthrombi, and may therefore be related to COVID-19 thrombotic complications. Within the lymph nodes and spleen, we observed lymphodepletion and both follicular and paracortical hyperplasia. Outside the lungs, histological changes were mainly related to complications of therapy or preexisting co-morbidities: mainly obesity, diabetes, and hypertension. Our approach focused on timely, systematic, and comprehensive tissue sampling and preservation of adjacent tissue samples for complementary analyses. We performed droplet digital polymerase chain reaction (ddPCR) for sensitive detection and quantification of SARS-CoV-2 gene targets in all tissue samples collected. To elucidate SARS-CoV-2 cell-type specificity and validate ddPCR findings, we performed in situ hybridization (ISH) broadly across sampled tissues. Immunohistochemistry (IHC) was used to further validate cell-type specificity in the brain where controversy remains on the regional distribution and cellular tropism of SARS-CoV-2 infection. In all samples where SARS-CoV-2 RNA was detected by ddPCR, we performed qRT-PCR to detect subgenomic (sg)RNA, an assay suggestive of recent virus replication15. We confirmed the presence of replication-competent SARS-CoV-2 in extrapulmonary tissues by virus isolation in cell culture. in 6 individuals we measured the diversity 113 and anatomic distribution of intra-individual SARS-CoV- 2 variants using high-throughput, single-genome amplification and sequencing (HT-SGS). We categorized autopsy cases of SARS-CoV-2 infection as “early” (n=17), “mid” (n=13), or “late” (n=14) by illness day (D) at the time of death, being ≤D14, D15-D30, or ≥D31, respectively. We defined persistence as presence of SARS-CoV-2 RNA among late cases. Due to the extensive tissue collection, we analyzed and described the results in terms of grouped tissue categories as the following: respiratory tract; cardiovascular; lymphoid; gastrointestinal; renal and endocrine; reproductive; muscle, skin, adipose, & peripheral nerves; and brain. Between April 26, 2020 and March 2, 2021, we performed autopsies on 44 PCR124 confirmed cases (Extended Data Fig. 1). SARS-CoV-2 seroconversion was detected in 38 of these cases (Supplementary Data 1); three early cases (P27, P36, P37) had not seroconverted and perimortem plasma was unavailable for the other three cases (P3, P4, P15). Extensive sampling of the brain was accomplished in 11 of the 44 cases (Fig. 1). The cohort was 29.5% female with a mean age of 59.2 years and was diverse across race and ethnicity (Extended Data Table 1). 95.5% of patients had at least one comorbidity, with hypertension (54.5%), obesity (52.3%), and chronic respiratory disease (34.1%) being most common. Patients presented to the hospital a mean of 9.4 days following symptom onset and were hospitalized a mean of 26.4 days. Overall, the mean interval from symptom onset to death was 35.2 days and the mean postmortem interval was 26.2 hours. 81.8% of patients required intubation with invasive mechanical ventilation, 22.7% received extracorporeal membrane oxygenation (ECMO) support, and 40.9% required renal replacement therapy. Vasopressors, systemic steroids, systemic anticoagulation, and antibiotics were commonly administered. SARS-CoV-2 RNA was detected in all 44 cases and across 79 of 85 anatomical locations and body fluids sampled (Extended Data Fig. 2, Supplementary Data 1). The highest burden of SARS-CoV-2 RNA (i.e., >100,000 N gene copies/ng RNA input) was detected in the respiratory tract of early cases (Figure 1), but we detected at least 100 N gene copies/ng RNA input from every tissue group besides reproductive tissues from multiple individuals among early cases. The mean SARS-CoV-2 N gene copies/ng RNA detected from tissues in each grouping among early cases are as follows: 9,210.10 across respiratory tissues; 38.75 across cardiovascular tissues; 30.01 across lymphoid tissues; 24.68 across gastrointestinal tissues; 12.76 across renal and endocrine tissues; 0.36 across reproductive tissues; 27.50 across muscle, peripheral nerve, adipose, and skin tissues; 57.40 across ocular tissues; and 32.93 across brain tissues With a few exceptions, the overall burden of SARS-CoV-2 RNA decreased by a log or more across tissue categories among mid cases, and further decreased among late cases. Further, persistence of low-level SARS-CoV-2 RNA (0.0004 to <0.5 N gene copies/ng RNA input) was frequently detected across multiple tissue categories among all late cases, despite being undetectable in plasma. Notably, SARS-CoV-2 RNA was detected in the brains of all six late cases and across most 158 locations evaluated in the brain in five of these six, including P42 who died at D230 Overall, SARS-CoV-2 RNA was detected in respiratory tissue of 43/44 cases (97.7%); cardiovascular tissue of 35/44 cases (79.5%); lymphoid tissue of 38/44 cases (86.4%); gastrointestinal tissue of 32/44 (72.7%); renal and endocrine tissue of 28/44 cases (63.6%); reproductive tissue in 17/40 cases (42.5%); muscle, skin, adipose, and peripheral nervous tissue in 30/44 cases (68.2%); ocular tissue and humors of 22/28 cases (57.9%); and brain tissue in 10/11 cases (90.9%) (Extended Data Table 3). We additionally detected SARS-CoV-2 sgRNA across all tissue categories, predominately among early cases (14/17, 82.4%), as well as in plasma, pleural fluid, and vitreous humor (Fig. 1, Extended Data Fig. 2, Supplementary Data 1). sgRNA was also detected in at least one tissue of 61.5% of mid cases and 42.9% of late cases, including across three tissue categories in a case at D99 (P20). We isolated SARS-CoV-2 in cell culture from multiple pulmonary and extrapulmonary tissues, including lung, bronchus, sinus turbinate, heart, mediastinal lymph node, small intestine, and adrenal gland from early cases up to D7 Spike RNA was detected throughout the respiratory tract in early cases, as well as within the sinus turbinate, trachea, lungs, from late cases (i.e., P33, P20, P42). The heart contained spike RNA within myocytes, endothelium, and smooth muscle of vessels of both early (P18, P19) and late (P3 & P42) cases. The pericardium demonstrated a positive signal for spike RNA within fibroblasts of the stroma. Intimal cells of the aorta were additionally 204 found to contain spike RNA. Mononuclear leukocytes within the lymph node, spleen, and appendix of an early case (P19) contained spike RNA, as did colonic epithelium . Epithelial cells along the intestinal tract in early cases (P16, P18, P19) contained viral RNA, as well as stratified squamous epithelium of the esophagus. Mononuclear leukocytes were again visualized with SARS-CoV-2 RNA in lymphoid aggregates and the interstitium of the small and large intestine, with infected cells still present in the colon of late cases (P33, P42). Kupffer cells, hepatocytes, and bile duct epithelium within the liver were additionally found to contain spike RNA. Within the kidney, spike RNA could be visualized within parietal epithelium of Bowman’s capsule, collecting duct cells, distal tubule cells, and glomerular endothelium. The adrenal glands contained spike RNA within endocrine cells. Endocrine follicular cells of the thyroid and glandular cells of the pancreas were also positive for spike RNA (Fig. 2). Among reproductive organs, spike RNA was visualized within Leydig and Sertoli cells of the testis, germ cells within the testicular tubules, endometrial gland epithelium, endometrial stromal cells, uterine smooth muscle cells, and stromal cells of the post-menopause ovary (Fig. 2). Myocytes within skeletal muscle contained spike RNA in both early (P18) and late (P20) cases. In addition to the organ-specific cell type infection of SARS-CoV-2, endothelium, muscularis of atrial vessels, and Schwann cells were identified as infected throughout the body, and were similarly positive across early and late cases. Spike RNA was found in neurons, glia and ependyma, as well as endothelium of vessels 5 across all lobes of the brain of early, mid, and late cases. Within the cerebellum specifically,neurons, Purkinje cells, and endothelium of vasculature 226 also contained spike protein via IHC In the examination of the 11 brains, we found few histopathologic changes, despite the evidence of substantial viral burden. Vascular congestion was an unusual finding that had an unclear etiology and could be related to the hemodynamic changes incurred with infection. Global hypoxic/ischemic change was seen in two cases... We show SARS-CoV-2 disseminates across the human body and brain early in infection at high levels, and provide evidence of virus replication at multiple extrapulmonary sites during the first week following symptom onset. We detected sgRNA in at least one tissue in over half of cases (14/27) beyond D14, suggesting that prolonged viral replication may occur in extra271 pulmonary tissues as late as D99. While others have questioned if extrapulmonary viral presence is due to either residual blood within the 272 tissue8,17 or cross-contamination from the lungs during tissue procurement8, our data rule out both theories. Only 12 cases had detectable SARS-CoV-2 RNA in a perimortem plasma sample, and of these only two early cases also had SARS-CoV-2 sgRNA in the plasma, which occurred at Ct levels higher than nearly all of their tissues with Furthermore, blood contamination would not account for the SARS-CoV-2 sgRNA or virus isolated from tissues. Contamination of additional tissues during procurement, is likewise ruled out by ISH demonstrating widespread SARS-CoV-2 cellular tropism across the sampled organs, by IHC detecting viral protein in the brain, and by several cases of virus genetic compartmentalization in which spike variant sequences that were abundant in extrapulmonary tissues were rare or undetected in lung samples. Using both ddPCR and sgRNA analysis to inform our selection of tissue for virus isolation and ISH staining allow us to describe a number of novel findings. Others6,8-12,17 have previously reported SARS-CoV-2 RNA within the heart, lymph node, small intestine, and adrenal gland. We demonstrate conclusively that SARS-CoV-2 is capable of infecting and replicating within these tissues. Current literature has also reported absent or controversial expression of ACE2 and/or TMPRSS2 in several extrapulmonary tissues, such as the colon, lymphoid tissues, and ocular tissues, calling into question if these tissues can become infected by SARS-CoV-21-3. However, we observed high levels of SARS-CoV-2 RNA and evidence of replication within these organs, as well as SARS-CoV-2 RNA via ISH in colonic mucosal epithelium and mononuclear leukocytes within the spleen, thoracic cavity lymph nodes, and GI lymphoid aggregates. We believe these ISH positive cells represent either infection or phagocytized virus in resident mac 294 rophages. Further, we isolated virus from a mediastinal lymph node and ocular tissue from two early cases (P19, P32). Our use of a single-copy sequencing approach for the SARS-CoV-2 spike allowed us to demonstrate homogeneous virus populations in many tissues, while also revealing informative virus variants in others. Low intra-individual diversity of SARS-CoV-2 sequences has been observed frequently in previous studies18-20, and likely relates to the intrinsic mutation rate of the virus as well as lack of early immune pressure to drive virus evolution in new infections. It is important to note that our HT-SGS approach has both a high accuracy and a high sensitivity for minor variants within each sample, making findings of low virus diversity highly reliable21. The virus genetic compartmentalization that we observed between pulmonary and extrapulmonary sites in several individuals supports independent replication of the virus at these sites, rather than spillover from one site to another. Importantly, lack of compartmentalization between these sites in other individuals does not rule out independent virus replication, as independently replicating populations may share identical sequences if overall diversity is very low. It was also interesting to note several cases where brain-derived virus spike sequences showed non-synonymous differences relative to sequences from other tissues. These differences may indicate differential selective pressure on spike by antiviral antibodies in brain versus other sites, though further studies will be needed to confirm this speculation. Our results collectively show while that the highest burden of SARS-CoV-2 is in the airways and lung, the virus can disseminate early during infection and infect cells throughout the entire body, including widely throughout the brain. While others have posited this viral dissemination occurs through cell trafficking11 due to a reported failure to culture virus from blood3,22, our data support an early viremic phase, which seeds the virus throughout the body following pulmonary infection. R 317 ecent work by Jacobs et al.22 in which SARS-CoV-2 virions were pelleted and imaged from COVID-19 patient plasma, supports this mechanism of viral dissemination. Although our cohort is primarily made up of severe cases of COVID-19, two early cases had mild respiratory symptoms (P28; fatal pulmonary embolism occurred at home) or no symptoms (P36; diagnosed upon hospitalization for ultimately fatal complications of a comorbidity), yet still had SARS-CoV-2 RNA widely detected across the body, including brain, with detection of sgRNA in multiple compartments. Our findings, therefore, suggest viremia leading to body-wide dissemination, including across the blood-brain barrier, and viral replication can occur early in COVID-19, even in asymptomatic or mild cases. Further, P36 was a juvenile with no evidence of multisystem inflammatory syndrome in children, suggesting infected children without severe COVID-19 can also experience systemic infection with SARS328 CoV-2. Finally, a major contribution of our work is a greater understanding of the duration and locations at which SARS-CoV-2 can persist. While the respiratory tract was the most common location in which SARS-CoV-2 RNA tends to linger, ≥50% of late cases also had persistence in the myocardium, thoracic cavity lymph nodes, tongue, peripheral nerves, ocular tissue, and in all sampled areas of the brain, except the dura mater. Interestingly, despite having much lower levels of SARS-CoV-2 in early cases compared to respiratory tissues, we found similar levels between pulmonary and the extrapulmonary tissue categories in late cases. This less efficient viral clearance in extrapulmonary tissues is perhaps related to a less robust innate and adaptive immune response outside the respiratory tract. We detected sgRNA in tissue of over 60% of the cohort. These data coupled with ISH suggest that SARS-CoV-2 can replicate within tissue for over 3 months after infection in some individuals, with RNA failing to clear from multiple compartments for up to D230. This persistence of viral RNA and sgRNA may represent infection with defective virus, which has been described in persistent infection with measles virus – another single-strand enveloped RNA virus—in cases of subacute sclerosing panencephalitis25. The mechanisms contributing to PASC are still being investigated; however, ongoing systemic and local inflammatory responses have been proposed to play a role5. Our data provide evidence for delayed viral clearance, but do not support significant inflammation outside of the respiratory tract even among patients who died months after symptom onset.

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