Cell-Free DNA: Potential Application in COVID-19 Diagnostics and Management.

WHO has declared COVID-19 as a worldwide, public health emergency. The elderly, pregnant women, and people with associated co-morbidities, including pulmonary disease, heart failure, diabetes, and cancer are the most predisposed population groups to infection. Cell-free DNA is a very commonly applied marker, which is elevated in various pathological conditions. However, it has a much higher sensitivity than standard biochemical markers. cfDNA appears to be an effective marker of COVID-19 complications, and also serves as a marker of certain underlying health conditions and risk factors of severe illness during COVID-19 infection. We aimed to present the possible mechanisms and sources of cfDNA released during moderate and severe infections. Moreover, we attempt to verify how efficiently cfDNA increase could be applied in COVID-19 risk assessment and how it corresponds with epidemiological data.

Cell-free DNA maps COVID-19 tissue injury and risk of death and can cause tissue injury.

INTRODUCTIONThe clinical course of coronavirus 2019 (COVID-19) is heterogeneous, ranging from mild to severe multiorgan failure and death. In this study, we analyzed cell-free DNA (cfDNA) as a biomarker of injury to define the sources of tissue injury that contribute to such different trajectories.METHODSWe conducted a multicenter prospective cohort study to enroll patients with COVID-19 and collect plasma samples. Plasma cfDNA was subject to bisulfite sequencing. A library of tissue-specific DNA methylation signatures was used to analyze sequence reads to quantitate cfDNA from different tissue types. We then determined the correlation of tissue-specific cfDNA measures to COVID-19 outcomes. Similar analyses were performed for healthy controls and a comparator group of patients with respiratory syncytial virus and influenza.RESULTSWe found markedly elevated levels and divergent tissue sources of cfDNA in COVID-19 patients compared with patients who had influenza and/or respiratory syncytial virus and with healthy controls. The major sources of cfDNA in COVID-19 were hematopoietic cells, vascular endothelium, hepatocytes, adipocytes, kidney, heart, and lung. cfDNA levels positively correlated with COVID-19 disease severity, C-reactive protein, and D-dimer. cfDNA profile at admission identified patients who subsequently required intensive care or died during hospitalization. Furthermore, the increased cfDNA in COVID-19 patients generated excessive mitochondrial ROS (mtROS) in renal tubular cells in a concentration-dependent manner. This mtROS production was inhibited by a TLR9-specific antagonist.CONCLUSIONcfDNA maps tissue injury that predicts COVID-19 outcomes and may mechanistically propagate COVID-19-induced tissue injury.FUNDINGIntramural Targeted Anti-COVID-19 grant, NIH.

Microbial cell-free DNA in plasma of patients with sepsis: a potential diagnostic methodology.

Sepsis is a life-threatening clinical condition demanding accurate and rapid diagnosis of the culprit pathogen, thereby to improve prognosis. Pathogen determination through blood culture is the gold standard for diagnosis but has limitations due to low sensitivity. Recently, circulating DNAs derived from pathogenic organisms were found in the plasma of patients with sepsis and were further proved to be more sensitive biomarkers for the diagnosis of the pathogen origin in sepsis. However, the fundamental molecular characteristics of circulating DNA in patients with sepsis remain unclear. Here, we used specific PCR and Sanger sequencing to verify the microbiology culture results via the corresponding plasma circulating DNA. We analyzed the composition and molecular characteristics of circulating DNA in septic patients using next-generation sequencing technology. We showed the presence of pathogen-derived circulating DNA in the plasma of patients with sepsis. The sizes of circulating DNA fragments derived from pathogenic bacteria showed a skewed unimodal distribution, while those derived from host cells showed a normal unimodal distribution. Lengths of fragments at peak concentration for both origins ranged from 150 bp to 200 bp, and reads mapping to pathogenic bacteria genome distributed uniformly on the reference. Our findings have improved our understanding of microbial circulating DNA in patients with sepsis as a potential methodology for the accurate diagnosis of sepsis, especially in light of an urgent need for such a diagnosis associated with the COVID-19 infection.

Use of Donor-Derived Cell-Free DNA for Assessment of Allograft Injury in Kidney Transplant Recipients During the Time of the Coronavirus Disease 2019 Pandemic.

BACKGROUND: Kidney allograft biopsy is the gold standard for diagnosis of rejection. Under the current extraordinary circumstances of the coronavirus disease 2019 (COVID-19), in which social distancing is key to limiting the spread of the virus, the model used to provide care to transplant recipients has undergone a very rapid transformation. In the spirit of medical distancing, we have been using the donor-derived cell-free DNA (dd-cfDNA) test for screening for rejection. METHODS: This article describes our experience with this approach between March 15th and May 20th, 2020. RESULTS: This test was obtained for-cause in 23 patients and for monitoring in 9 patients. Normal results aided in forgoing biopsy in 63% of the patients for whom the test was obtained in the outpatient setting. The test is neither 100% sensitive nor specific for rejection; however, when used in combination with the available clinical information, it can be used for determining whether bringing in a transplant recipient into a medical facility is necessary. CONCLUSIONS: In the event COVID-19 becomes a long-term challenge for our community, noninvasive biomarkers such as the dd-cfDNA may become more relevant than ever in enhancing our ability to care for our transplant patients while maximizing the distancing measures.

Making the most of the first prenatal visit: The challenge of expanding prenatal genetic testing options and limited clinical encounter time.

OBJECTIVE: Advances in prenatal genetics place additional challenges as patients must receive information about a growing array of screening and testing options. This raises concerns about how to achieve a shared decision-making process that prepares patients to make an informed decision about their choices about prenatal genetic screening and testing options, calling for a reconsideration of how healthcare providers approach the first prenatal visit. METHODS: We conducted interviews with 40 pregnant women to identify components of decision-making regarding prenatal genetic screens and tests at this visit. Analysis was approached using grounded theory. RESULTS: Participants brought distinct notions of risk to the visit, including skewed perceptions of baseline risk for a fetal genetic condition and the implications of screening and testing. Participants were very concerned about financial considerations associated with these options, ranking out-of-pocket costs on par with medical considerations. Participants noted diverging priorities at the first visit from those of their healthcare provider, leading to barriers to shared decision-making regarding screening and testing during this visit. CONCLUSION: Research is needed to determine how to restructure the initiation of prenatal care in a way that best positions patients to make informed decisions about prenatal genetic screens and tests.