Risk Factors Associated With Six-Month Mortality in Hospitalized COVID-19 Patients: A Single-Institution Study

Background Coronavirus disease 2019 (COVID-19) infection can vary from asymptomatic infection to multi-organ dysfunction. The most serious complication of infection with COVID-19 is death. Various comorbid conditions and inflammatory markers have been associated with an increased risk of mortality, specifically within the immediate post-infection period; however, less is known about long-term mortality outcomes. Objectives Our objective is to determine risk factors associated with six-month mortality in hospitalized COVID-19 patients. Methods This is a single-institution, retrospective study. We included patients hospitalized with COVID-19 from the University of Toledo Medical Center in Toledo, Ohio, who were admitted from March 20, 2020, to June 30, 2021. This study was approved by a biomedical institutional review board at the University of Toledo. Patients with available pre-stored blood samples for laboratory testing were included, and hospital charts were assessed up to six months from the date of a positive COVID-19 test result. Two groups were created based on the mortality outcome at six months from COVID-19 positive test results: survivors and non-survivors. The clinical variables or outcomes and laboratory values were compared between the two groups using non-parametric methods due to the small sample size and non-normality of the data. Either the Mann-Whitney U-test for continuous variables or Fisher’s exact test for categorical variables was used for statistical analysis. Results Lactate dehydrogenase (LDH) and D-dimer levels on admission were found to be significantly higher in non-survivors than in survivors. The median high D-dimer level in non-survivors was 5.96 micrograms/milliliter (μg/mL) (interquartile range (IQR): 3.95-11.29 μg/mL) vs 1.82 μg/mL (IQR 1.13-5.55 μg/mL) in survivors (p = 0.019). Median LDH levels were also higher in non-survivors vs survivors, i.e., 621.00 international units per liter (IU/L) (IQR 440.00-849.00 IU/L) vs 328.00 IU/L (IQR 274.00-529.00 IU/L), respectively (p = 0.032). The demographic profile, comorbidity profile, and laboratory data (typically associated with short-term mortality, inflammation, and organ dysfunction) were similar between survivors and non-survivors, except for LDH and D-dimer. Conclusion Higher LDH and D-dimer levels on admission were found to be associated with an increased six-month mortality rate in hospitalized COVID-19 patients. These hematologic data can serve as risk stratification tools to prevent long-term mortality outcomes and provide proactive clinical care in hospitalized COVID-19 patients.

The coronavirus disease 2019 (COVID- 19) pandemic has undoubtedly altered the course of world history since the first cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were reported in adult patients in December 2019. The virus is a single-stranded, enveloped RNA virus that primarily infects mammalian hosts [1]. SARS-CoV-2 is highly transmissible in humans and is spread commonly via respiratory droplets. The most common symptoms include fever, cough, shortness of breath, fatigue, and nausea. Common pulmonary manifestations include acute viral pneumonia; however, COVID-19 has a wide spectrum of extra-pulmonary symptom manifestations as well, ranging from asymptomatic infection to multi-organ dysfunction [2]. The most serious complication of infection with COVID-19 is death; there have been 542 million reported cases, and COVID-19 has claimed the lives of over 6.33 million people worldwide since 2019 [3].
Patients hospitalized with COVID-19 infection have unique characteristics that can impact their short-term mortality rates and disease severity. Between 60% and 90% of hospitalized patients have other comorbidities (most commonly hypertension, diabetes mellitus, cardiovascular disease (CVD), and chronic obstructive pulmonary disease), and approximately 35% of hospitalized patients experience severe disease necessitating intensive care unit level of care [1]. Symptoms, organ dysfunction, and/or psychological damage may persist for weeks to months after infection with COVID-19 in a minor subset of patients; long-term COVID-19 is a distressing complication for some after acute infection.
The pathophysiology, disease course, and validated treatment modalities have been extensively examined since the advent of the worldwide COVID-19 pandemic three years ago; however, post-acute infection data continues to surface, which dynamically shapes our knowledge of the impact COVID-19 has on patients and communities. Laboratory data obtained on hospitalized patients during acute COVID-19 infection may be a useful tool to predict post-acute mortality risk. Much of the study data relating to post-acute COVID-19 infection morbidity and mortality risk in the United States (US) is 90-day data; however, some European studies have followed patients for six months post-infection [4]. The purpose of this study is to retrospectively examine single-center data from hospitalized adult patients suffering from COVID-19 infection in Toledo, Ohio, USA, between March 2020 and June 2021 to determine the demographic, social, and laboratory factors that may be associated with a higher six-month mortality rate.

Statistical analysis
The primary objective of this study is to identify risk factors that may be associated with six-month mortality. The clinical variables or outcomes and laboratory values were compared using non-parametric methods due to the small sample size and non-normality of the data. Either the Mann-Whitney U-test for continuous variables or Fisher's exact test for categorical variables was used for statistical analysis. All analyses were performed using R 4.0.3 (R Foundation for Statistical Computing, Vienna, Austria) and an alpha level of 0.05 was considered statistically significant.

Ethical approval
The approval for our retrospective study conducted at the University of Toledo Medical Center was granted by the biomedical Institutional Review Board (IRB) prior to beginning the collection of data, IRB number 300722-UT.

Results
Our final analysis included a total of 28 patients. Eleven of the 28 were non-survivors and seventeen of the 28 were survivors six months after hospitalization for COVID-19 infection. No difference was found in the demographic profile of patients in the two groups. The median age, sex, and ethnicity were statistically similar between the two groups. The median age was 64 years (interquartile range (IQR): 59.00-67.00 years) for non-survivors and 61 years (IQR: 53.00-68.00 years) for survivors (p = 0.524). There was no statistically significant difference in sex distribution or classification by the racial and ethnic group either. About 45.5% of non-survivors were male, while 76.5% of survivors were male (p = 0.125). Among non-survivors, 36.4% were White, 54.5% were Black, and 9.1% were Hispanic; among survivors, 47.1% were White, 41.2% were Black, and 0% were Hispanic (p = 0.486). No statistically significant difference in six-month mortality outcome was found based on the selected comorbidity profile: coronary artery disease, atrial fibrillation, cerebrovascular accidents, peripheral artery disease, cancer, and smoking.

Discussion
To our knowledge, this is the first study in the United States analyzing risk factors associated with six-month mortality in hospitalized COVID-19 patients though some literature from European countries exists in this context [4,5]. Studies on the predictors of acute-mortality outcomes in patients with COVID-19 have been published in the US, but not in the context of six-month mortality. Our study found higher median lactate dehydrogenase (LDH) and D-dimer levels on initial hospitalization were associated with six-month mortality in hospitalized patients with COVID-19 infection. Our results found that the median D-dimer in the nonsurvivor group was 3.3 times higher than that of the survivor group (alive at six months). The median LDH in the non-survivor group was approximately 1.9 times higher than that of the survivor group.
LDH is an enzyme found in many tissues in our bodies; consequently, tissue damage leads to increased LDH released into the blood. Other studies involving COVID-19 patients have found an association between higher LDH levels and poor prognostic outcomes [6][7][8]. Fan et al. found elevated LDH at admission in ICU patients compared to their non-ICU group (p-value <0.001) [6]. Yan et al. analyzed blood samples from 485 infected patients from Wuhan, China, to identify biomarkers of COVID-19 mortality via machine learning tools, and they found LDH, lymphocytes, and high-sensitivity CRP predicts mortality more than 10 days in advance with more than 90% accuracy [7]. Specifically, levels of LDH alone in the blood were highly predictive of COVID-19 mortality. Higher levels of LDH are used as markers of tissue damage in liver disease and lung tissue death. Furthermore, published reports suggest LDH is a prognostic indicator of lung fibrosis in severe interstitial lung disease [8]. In a study conducted in Yichang, China, Lv et al. reported higher LDH levels in non-survivors of COVID-19 than survivors in the post-COVID lung fibrosis stage. The median survival time was 27.5 days in patients with high LDH, quantified as greater than 263 IU/L, compared to those with low LDH, quantified as less than 263 IU/L, who had a median survival time of 40 days [9].
D-dimer is a degradation product of cross-linked fibrin and is often used as a marker for hemostatic abnormalities, specifically blood clots. In many studies [7], increasing D-dimer levels have been associated with COVID-19 severity as well as mortality. In a study on persistent D-dimer levels in patients with convalescent COVID-19, the observed rate of increased D-dimer levels was higher in hospitalized patients and predicted thrombotic complications [10]. In a case-control study by Yao et al., D-dimer elevation upon admission was associated with increased disease severity and in-hospital mortality [11]. Similar to our study, Li et al. concluded normal D-dimer on presentation, day one, is predictive of 28-day survival [12]. Furthermore, a normal D-dimer on day three is even more predictive of 28 days of survival. In contrast to Li et al., our study analyzed a longer time frame of six months vs 28-day survival. Previous research has shown that D-dimer values are frequently elevated in COVID-19 patients; however, they are even higher in patients with severe COVID-19 compared to those with milder forms. In a retrospective cohort study in Wuhan, China, conducted by Zhou et al., it was reported that D-dimer levels, specifically those greater than 1 ug/mL, were approximately ninefold higher in non-survivors than survivors in a 22-day mortality period [13].
We did not find significant differences between the two groups in either peak or nadir values of hematologic parameters, i.e., hemoglobin (Hgb), white blood cell (WBC), platelet count (plt), absolute neutrophil count (ANC), and absolute lymphocyte count (ALC). Anemia and thrombocytopenia have previously been found to be associated with disease severity. Thrombocytopenia has also been linked to an increased risk of COVID-19 mortality [14,15]. One of the first studies to report an association between thrombocytopenia as a risk factor for mortality in COVID-19 patients was conducted in China. Thrombocytopenia was associated with an approximately three times higher risk of mortality compared to those without thrombocytopenia. This data is in contrast with our findings and may be attributed to the difference in sample size [16]. A metaanalysis of the clinical characteristics of COVID-19 patients reported normal leukocyte counts in 69.7% of patients, lymphopenia in 59.5% of patients, and leukocytosis in 12.6% [17]. We did not report significant differences in leukocyte counts between survivors and non-survivors. The peak and nadir of median ferritin, c-reactive protein (CRP) levels, and erythrocyte sedimentation rate (ESR) were similar between the two groups. In a systematic review by Iwamura et al., ESR and CRP levels were elevated regardless of the degree of severity of the infection [18]. Our data is consistent with these findings.
No significant association between six-month mortality and demographic characteristics such as age, sex, race, and ethnicity was found. In contrast to this data, a cohort study in Germany found the odds of sixmonth all-cause mortality increased by 2.21 for every 10 years of age [4]. They also report that the female sex is protective as women had improved long-term outcomes than males. The limited sample size of our study may explain the difference within our findings. Larger studies to elucidate associations between demographic characteristics and six-month mortality may be warranted. Comorbid conditions such as coronary artery disease (CAD), atrial fibrillation (Afib), cerebrovascular accidents (CVAs), and peripheral artery disease (PAD) did not predict mortality. A systemic review of predictors of mortality by Tian et al. reports cardiovascular disease and cerebrovascular disease were associated with a higher risk of mortality, even though mechanisms are not yet understood [19]. In a French study, coronary artery calcifications (CACs) were found to be an independent predictor of six-month mortality in COVID-19 patients, even those without known atherosclerotic cardiovascular disease [20]. Published data is unclear in the association of various comorbid conditions and the prediction of mortality and severity of COVID-19.

Limitations
This study has limitations. The study was conducted in one specific institution and had a limited sample size. The external validity of our results would increase with studies in various regions and institutions and with larger sample sizes. Due to the retrospective nature of the study, previously collected data was utilized, which limited the variables that were analyzed. Lastly, laboratory data were analyzed from a one time frame, from initial hospitalization; therefore, further studies are needed to elucidate possible time-dependent changes in prognostic variables.