Evaluation of Laboratory Findings of Patients with Coronavirus Disease 2019 in Kerman, Iran

Background & Objective: Since December 2019 in Wuhan, China there is a new form of pneumonia and after expansion in other countries, World Health Organization (WHO) called it Coronavirus Disease 2019 (COVID-19). Since the clinical laboratory findings have played an important role in the progression of the disease, this study aimed to evaluate the laboratory findings in COVID-19 patients (before vaccination). Methods: In this case-control study that was conducted from February to August 2020; the laboratory test status in 101 positive COVID-19 patients was evaluated and compared with 101 healthy individuals. Results: The results of our study showed that 21% of patients had low WBC, 24.75% low RBC, 37.62%, low Hb, 18.81% with low HCT, 29.7%, low Plt, 41.58% had High PT, 71.29% high CRP, 17.82% high urea, 11.88% high CR, 15.84% high LDH, 10.89% low sodium, 14.75% low potassium (K). The quantitative examination of blood factors showed that lymph%, mixed%, PLT, HCT, Hb, and RBC were higher in the control group than in the case group. While Neu%, WBC, PTT, CRP, UREA, LDH, K in the patient group were higher than in the control group. Conclusion: According to the results of the study, it can be concluded that in the clinical treatment of COVID-19 patients, much attention should be paid to the laboratory indicators to identify and intervene early in critically ill patients.


Introduction
Since December 2019 epidemic of Coronavirus Disease 19  worldwide, studies are started on this virus.This kind of Coronavirus was isolated from human airway epithelial cells, and it can cause severe acute respiratory syndrome COV2 (1).In the past two decades after severe acute respiratory syndrome coronavirus (SARS-COV) and Middle East respiratory syndrome coronavirus (MERS-COV), SARS-COV2 is the third virus from this group that treatment public health around the world (1)(2)(3)(4)(5).
The mean incubation period was 5.2 days.Studies show human-to-human transmission has occurred among close contacts (13,14).Common clinical symptoms include fever, cough, tiredness, dyspnea, and loss of taste and smell.Less common symptoms are sore throat, headache, diarrhea, and chest pain (5).
The gold standard for diagnosis of COVID-19 is the identification of viral genome with PCR in respiratory tract secretion during the first week of symptoms (6).Mortality rate increased in older age, with high Sequential Organ Failure Assessment (SOFA) score and when d-dimer greater than 1 μg/mL (on admission) (15)(16)(17)(18).
COVID-19 infection leads to many abnormal laboratory parameters.Hematological, biochemical, and coagulation-fibrinolysis tests are adequate indicators.Therefore, the evaluation of clinical laboratory parameters can help to stop disease progression, decrease the side effects of the virus, and can help to diagnose patients in an early stage of Vol.18 No. 3 Summer, 2023 IRANIAN JOURNAL OF PATHOLOGY COVID-19 infection before showing positive PCR tests.This study aimed to determine and compare the significant differences in laboratory tests (CBC and biochemistry) between the COVID-19 patients with healthy control groups (normal populations before the COVID-19 pandemic) in Kerman, Iran.

Material and Methods
This case-control study's population consisted of 101 patients with COVID-19 infection, which were confirmed by polymerase chain reaction (PCR) test (according to WHO (World Health Organization) guidelines) and admitted to Afzalipour Hospital, Kerman, south-eastern Iran, between February and November 2020, in addition to 102 healthy controls that had no disease with the same laboratory exam before COVID-19 pandemic.Included criteria in the study were patients between 20-80 years of age, without a history of using anti-coagulant drugs (such as Warfarin) recently, and no liver or kidney disease.Patients diagnosed with DIC (Disseminated Intravascular Coagulation) or increased INR (International Normalized Ratio), PTT (Partial Thromboplastin time), and PT (Prothrombin Test) at the same time were excluded.

Data Collection
All patients with COVID-19 infection were confirmed by PCR tests which were performed according to WHO guidelines.Laboratory information was collected from admission files.
Using the records data about demographic, laboratory tests including CBC, PT, PTT, INR, BUN, Cr, Na, K, BUN, ALK-p, LDH, and CRP on the first day of hospitalization and also for demographic data, habits, lifestyle and past medical history of both groups, questioners were filled.
Coagulation function tests, which included Partial Thromboplastin Time (P.T.T), Prothrombin Time (P.T), and International Normalized Ratio (I.N.R), were performed using manual tests and duplicate analysis.
All participants signed a written informed consent, which explicitly provided permission for gathering the relevant clinical data.Those who were unable or unwilling to participate were excluded.

Statistical Analysis
The STATA (version 14) was used for data analysis.

Results
This study was a case-control study with 101 hospitalized patients (M: 60%, F: 40%) with PCRconfirmed SARS-CoV-2 infection and 102 control group (M:61%, F:39%).In Table 1 we compared demographic data (such as age, gender, marital status, etc.), habits (like cigarette, opium, alcohol, etc.) and, being at high-risk places such as hospitals, care clinics dorms, hotels, and nursing homes in both control and the patient group.No significant difference in sex or age was observed between the patient (49.46±17.75years) and the control group (51.03±14.69years) (P= 0.955).Most of the patient and the control group population were married (95% vs. 93%).
Table 2 shows the past medical history of the study population; inpatient group asthma was reported more than in the control group.About other medical histories, there was no significant difference (P> 0.005).
Different cancer types in study groups were shown in Table 3.Two patients mentioned a history of lung cancer and one mentioned cervical cancer in the patient group.In the control group pancreas cancer (2 persons), bladder carcinoma (2 persons), laryngeal carcinoma (1 person), and 1 person mentioned cervical cancer were reported.Our data about laboratory parameters was arranged and the main laboratory findings were compared in both groups (Tables 4 and 5).Main laboratory tests including WBC, RBC, HB, HCT, PLT, PT, CRP, Urea, Cr, LDH, Na, and K were in the normal range in both patient and control groups, but about 22% of patients show leukopenia (WBC < 4x10 9 /µL), 25% decrease RBC count and 38% of patients show anemia (Less than 12 gr/dl in Female or less than 14 gr/dL in Male) in Hb concentration.
WBC differentiation also showed a significant difference in the patient and control groups (Neutrophils % 71.34±1.16

Table 4. Laboratory parameters in patient and control groups
The quantitative examination of blood factors showed that lymph%, PLT, HCT, HB, RBC, and mixed% were significantly higher in the control group than in the patient group.While Neu%, WBC, PTT, CRP, UREA, LDH, and K in the patient group were significantly higher than the control group.In Tables 7 and 8 past medical history and opium intake were evaluated respectively that shows asthma can play a role in predicting underlying disease to COVID-19 infection (P=0.036).Also, in COVID-19 patients, last month opium users increased (P=0.028).

Discussion
Since December 2019, novel coronavirus disease (COVID-19; previously known as 2019-nCoV) has been reported in Wuhan, China, and subsequently affected worldwide.In general, COVID-19 is an acute resolved disease, but it can also be deadly, with a 2% case fatality rate (diffuse alveolar damage) (19).The most common symptoms were fever, shortness of breath, expectoration, fatigue, dry cough, and myalgia (38).Most COVID-19 patients presented with dyspnea with comorbidities such as hypertension and diabetes (39).Our study also showed diabetes, cardiovascular disturbances, respiratory disease including COPD and asthma, and increased blood pressure (hypertension) as comorbidities in COVID-19 patients.Also, asthma increased the hospitalization rate in patients (5).In a study by Haung et al., most COVID-19 patients were men and underlying diseases include diabetes, hypertension, and cardiovascular disorders.Also, in most of the patients, lymphopenia was detected in the CBC test (20,21).No significant changes were seen between the patient and the control group (P>0.005).Different studies on laboratory parameters in COVID-19 patients showed significant changes in WBC count; Qin et al. (2020), revealed that severe patients of COVID-19 tend to have lower lymphocyte counts, higher leukocyte counts, and neutrophil-lymphocyte ratio (NLR), as well as lower percentages of monocytes, eosinophils, and basophils (mixed).Also, they claimed that the number of T cells significantly decreased.Both helper T (Th) cells and suppressor T cells in patients with COVID-19 were less than normal levels (22).The present study showed that in 21% of patients with low WBC, Neu% in the patient group was significantly higher than in the control group.
In another study, Yuan et al. showed lower lymphocyte count, decreased red blood cell and hemoglobin, low levels of immunoglobulin G, and significantly higher D-dimer, fibrinogen, white blood cell count, neutrophil count, interleukin-6, C-reactive protein, procalcitonin, erythrocyte sedimentation rate, ferritin, and lactate dehydrogenase in severe and critically ill patients (23,24).Elevated TnT, CRP, and D-dimer, and declined PaO2/FiO2 suggest that fatality due to COVID-19 was associated with multiple organ dysfunction (23) studies show other coagulation parameters such as PT, PTT, and fibrinogen in non-ICU-patients were less significant in identifying patients needing ICU care (25).
In another study by Badel on 1810 pediatric patient population with positive PCR test for COVID-19 leukopenia, lymphopenia, elevated ferritin, procalcitonin, and CRP was detected in a laboratory study (25-29).In the transmission of COVID-19, cold and dry weather and low levels of ultraviolet radiation are moderately associated with increased SARS-CoV-2 transmissibility, with humidity playing an important role.Quarantine, physical distancing, and social isolation are effective ways to reduce coronavirus transmission (30)(31)(32)(33)(34).There was no significant difference between being in high-risk places such as hotels, care clinics, dorms, and nursing homes between the control and the patient groups.In multiple studies that analyzed the ACE2 RNA expression in the respiratory tract, ACE2 expression appears in nasal epithelial cells and the size of this population of ACE2-expressing nasal epithelial cells is comparable with the size of the population of ACE2expression in type II alveolar cells (AT2) (35,36).In the inflammatory process and cytokines storms IL-19 may play important roles in inflammatory responses because it up-regulates IL-6 and TNF-alpha and induces apoptosis (36).Aggrawal evaluated the role of TNF-induced apoptosis in T cell deficiency in lymphocytes from aged humans, expression of TNF receptors (TNFRI and TNFRII), and the adapter molecules, including TNFR-associated death domain protein (TRADD), TNFR-associated factor 2 (TRAF-2), and receptor-interacting protein (RIP).An increased constitutive expression of TNFRI and TRADD and decreased expression of TNFRII and TRAF-2 was observed in lymphocytes from aged patients, also increased activation of caspases (caspase-8 and caspase-3) involved in TNFR/TNF signaling pathway (37).Maybe in our study increased CRP levels and lymphopenia can complain with these hypotheses.Maybe viral infection could be a potential risk factor for idiopathic pulmonary fibrosis because of cytokine storms (38).Probable reasons for the SARS-associated lymphopenia may be direct infection of lymphocytes by SARS-CoV, lymphocyte sequestration in the lung, or cytokine-mediated lymphocyte trafficking.Also, it could be due to immune-mediated lymphocyte destruction, bone marrow or thymus suppression, or apoptosis (24,39).Henry studied 1189 children with COVID-19 infection and showed that monitoring lymphocyte count, CRP, PCT (Procalcitonin), and IL-6 in severe patients would help to manage the treatment of the disease (40).Liu mentioned NLR as an independent risk factor of the inhospital mortality for COVID-19 patients (more than 6.5) 8% higher risk of in-hospital mortality for each unit increase in NLR and may help identify high-risk individuals with COVID-19 (41).
Hematological factors such as platelet count, prothrombin time, D-dimer, and neutrophil-tolymphocyte ratio can help clinicians to assess the severity and prognosis of patients with COVID-19.The sepsis-induced coagulopathy scoring system can be used for the early assessment and management of patients with critical disease (42,43).The result of the present study that used a significant Coagulation test including PT, PTT, and I.N.R showed a significant difference in the patient and the control group (P <0.001).It may be because of coagulopathy during sepsis.

Conclusion
According to the results of the study, it can be concluded that in the clinical treatment of COVID-19 patients, much attention should be paid to laboratory indicators to identify and intervene early in critically ill patients.

Table 1 .
Demographic data & habits of patient and control groups M means male and F shows female, tobacco means cigarette, hookah, or any type of nicotine.

Table 2 .
Underlying disease in patients and control groups

Table 3 .
Malignancy typing in patient and control groups Cr in 11.88% LDH in 15.84% and blood electrolytes Na in 5.94%, and K in 4.95% of the patient group increased significantly (P<0.001).C.R.P as an inflammatory factor in blood tests increased in 71% of patients.

Table 5 .
The laboratory tests range in both patient and control group

Table 6 .
Relationship between demographic data, habits, and COVID-19 infection in patient and control group *Tobacco include cigarette, hookah or other type of nicotine

Table 7 .
The relationship between underlying disease and COVID-19 infection in patient group

Table 8 .
Relationship between times of opium use and infection in the patient group