|Year : 2021 | Volume
| Issue : 2 | Page : 159-166
Prognostic significance of absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio in COVID-19
KV Padmaprakash1, Kuldeep Kumar Ashta2, Nishant Raman3, Vasu Vardhan4, Sandeep Thareja5, J Muthukrishnan1, KS Rajmohan6, Sudhir Dubey6, Dheeraj Nauhwaar1, Abhinav Kumar2, P Basavaraj2
1 Department of Gastroenterology, Base Hospital, New Delhi, India
2 Department of Internal Medicine, Base Hospital, New Delhi, India
3 Medical Officer, Base Hospital, New Delhi, India
4 Commandant, Base Hospital Delhi Cantt, New Delhi, India
5 Brig IC and Cdr Tps, Base Hospital Delhi Cantt, New Delhi, India
6 Department of Pathology, Base Hospital, New Delhi, India
|Date of Submission||08-Mar-2021|
|Date of Decision||11-May-2021|
|Date of Acceptance||16-May-2021|
|Date of Web Publication||05-Oct-2021|
Medical Officer, Base Hospital, Delhi Cantonment, New Delhi
Source of Support: None, Conflict of Interest: None
Introduction: Effective triage of COVID-19 patients, especially in resource-limited settings, requires cost-effective and readily available markers. The present study looks at the prognostic role of three such laboratory parameters, absolute lymphocyte count (ALC), absolute neutrophil count (ANC), and neutrophil-to-lymphocyte ratio (NLR). Methodology: A retrospective cohort study was done including 328 COVID-19 reverse transcriptase–polymerase chain reaction-confirmed hospitalized patients aged 18 and above in a tertiary center in northern India. Baseline demographic, clinical, and laboratory parameters were collected on the day of admission. Statistical analysis included descriptive statistics, sensitivity–specificity analysis for optimum cutoffs, multiple logistic regression, and Cox proportional hazards regression. Results: The median age of the patients was 45 with 266 (81.1%) males and 62 (18.9%) females. A total of 109 (33.2%) patients were affected with moderate to severe disease. Forty-nine (14.9%) patients had fatal outcomes. Median ALC was lower in patients with moderate to severe disease compared to mild disease (895 vs. 1554.2) and in nonsurvivors compared to survivors (732.0 vs. 1423.9). Median ANC (5182.8 vs. 3057.6) and NLR (5.38 vs. 2.03) were significantly raised in patients with moderate to severe disease as against mild disease and in nonsurvivors(ANC 7040.25 vs. 3448.5, NLR 10.05 vs. 2.35). ALC <1273, ANC >3907, and NLR >2.74 showed considerable sensitivity and specificity for disease severity at admission. ALC and ANC were significantly associated with the odds of moderate to severe disease at admission in the multivariable logistic regression analysis. ALC <1183, ANC >4612, and NLR >3.76 had good sensitivity and specificity as predictors of mortality and emerged as independent risk factors for mortality in the multivariable Cox proportional hazards regression. Conclusion: ALC, ANC, and NLR are relatively cost-effective and readily available routine investigations obtained as a part of complete blood count. These indices show good prognostic significance. Their utility in clinical algorithms can better guide management of patients.
Keywords: Absolute lymphocyte count, absolute neutrophil count, COVID-19, lymphopenia, neutrophilia, neutrophil-to-lymphocyte ratio
|How to cite this article:|
Padmaprakash K V, Ashta KK, Raman N, Vardhan V, Thareja S, Muthukrishnan J, Rajmohan K S, Dubey S, Nauhwaar D, Kumar A, Basavaraj P. Prognostic significance of absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio in COVID-19. J Mar Med Soc 2021;23:159-66
|How to cite this URL:|
Padmaprakash K V, Ashta KK, Raman N, Vardhan V, Thareja S, Muthukrishnan J, Rajmohan K S, Dubey S, Nauhwaar D, Kumar A, Basavaraj P. Prognostic significance of absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio in COVID-19. J Mar Med Soc [serial online] 2021 [cited 2021 Nov 28];23:159-66. Available from: https://www.marinemedicalsociety.in/text.asp?2021/23/2/159/327050
| Introduction|| |
COVID-19 is a rapidly spreading infectious disease caused by SARS-CoV-2, a novel coronavirus. Although most infections are mild and usually self-limiting, severe infection often progresses to severe pneumonia, acute respiratory distress syndrome, sepsis, and Multi organ dysfunction (MODS) ultimately culminating in death.
An aspect of COVID-19, other than its potentially fatal nature, is the huge impact on health-care burden it has. While for milder forms of the disease, general isolation with supportive management is at most adequate, patients with severe disease often require intensive care. Early identification of risk factors for critical illness is hence a prerequisite for provision of an effective triage and for forecasting intensive care unit requirement.
Numerous markers of disease severity and mortality have been identified previously. The pathological background behind many of these markers involves an unregulated immune response leading to a multisystemic involvement of the disease.,, Hyperinflammation and cytokine storm have been shown to be a cardinal feature of disease severity. It is yet not well understood what drives and propagates this overexuberant immune response. Recent literature points toward neutrophil infiltration found in pathological specimens from autopsied COVID-19 patients.,, Neutrophil infiltration in pulmonary capillaries, acute capillaries with fibrin deposition, extravasation of neutrophils into the alveolar space, and neutrophilic mucositis were observed.,, Simultaneously, increased neutrophil counts were observed in the peripheral blood of COVID-19 patients who died., Of note, a high incidence of lymphopenia in COVID-19 patients has also been reported.,, This sparked interest in neutrophilia and lymphopenia as markers of COVID-19 severity and mortality. In this background, neutrophil-to-lymphocyte ratio (NLR), a dividend of the absolute neutrophil (ANC) and lymphocyte (ALC) counts, has also been studied with respect to disease severity and mortality.
NLR is an inflammatory biomarker and has been utilized previously in the prognostication of other diseases including cancer, community-acquired pneumonia, and sepsis.,,, The NLR is a simple test, ANC and ALC being routinely performed as a part of the complete blood count and hence do not add to the cost of diagnosis.
In the above premise, the present study was planned to determine ALC, ANC, and NLR as predictors of disease severity as well as their prognostic value.
| Methodology|| |
A retrospective study was done in a tertiary hospital in northern India and involving 328 COVID-19 patients. Cases included in the study were patients with COVID-19 aged 18 and above and confirmed by a positive result from real-time reverse transcriptase–polymerase chain reaction (RTPCR) assay for nasal, nasopharyngeal, and throat swab specimens. Demographic and clinical data were collected at presentation and were inclusive of age, gender, and comorbidities. Laboratory investigations were performed on day 1 of admission and were inclusive of total leukocyte count (TLC), differential leukocyte count, ALC, ANC platelet count (PLT), blood chemistry, and C-reactive protein (CRP).
Patients were categorized at presentation based on national guidelines into two categories, i.e., mild disease and moderate to severe disease. COVID-19 patients in the mild category included those who maintained an oxygen saturation of >94% in room air, with a respiratory rate of <24/min and with no signs and symptoms of pneumonia. The second category of moderate to severe comprised patients with an oxygen saturation of <94% at room air with a respiratory rate of >24/min and with evidence of pneumonia. The primary endpoint was taken as in-hospital all-cause mortality.
Categorical variables included gender, comorbidities, disease severity, and outcome and were described as frequencies and percentages. Continuous variables including age, TLC, ALC, ANC, NLR, PLT, and CRP were described as medians and interquartile ranges (IQR). Chi-square tests were utilized for comparing categorical variables. Continuous variables were compared by means of t-test or one-way analysis of variances if normally distributed. Mann–Whitney U-test was employed for comparison of nonnormally distributed data. Continuous variables were further dichotomized using appropriate sensitivity and specificity from ROC curves. Survival analysis was done using Cox proportional hazards regression. Statistical significance was considered at alpha < 0.05 for 95% confidence interval and power of 80%. MS Excel 2016 and Excel for Microsoft office 365 (Microsoft Corporation), SPSS version 23 for Windows (IBM SPSS Statistics for Windows, Version 23.0. Armonk NY: IBM Corp) were used respectively for data handling and analysis.
| Results|| |
A total of 328 patients were included in the study with 266 (81.1%) males and 62 (18.9%) females. The median age of patients was 48 (IQR: 25) years. Mild disease at presentation was observed in 219 (66.8%), whereas 109 (33.2%) patients were affected with moderate to severe disease. Forty-nine (14.9%) patients had fatal outcomes. The median duration from admission to death was 7 days (IQR: 5 days). For patients who survived, the median duration of admission was 13 days (IQR: 7 days). A total of 123 (37.5%) patients had at least one underlying comorbidity with 62 (18.9) patients having multiple (two or more) comorbidities. The median ALC value was 1402.15 (IQR: 1052.27), median ANC value was 3856.10 (IQR: (2865.52), and the median NLR value was 2.49 (IQR: 3.78). Lymphopenia and neutrophilia were observed in 128 (39%) and 60 (18.3%) patients. The demographic, clinical, and laboratory characteristics are presented in [Table 1].
|Table 1: Demographic, clinical, and laboratory indices recorded at the time of admission|
Click here to view
Statistically significant differences existed between the values of ALC, ANC, and NLR in patients with mild disease as against those with moderate to severe disease. ALC was lower in patients with moderate to severe disease (median:895.5, IQR: 686.73) compared to mild disease (median: 1554.2, IQR: 1025.8). ANC in patients with moderate to severe disease was significantly higher (median: 5182.8, IQR: 5222.85) compared to that in patients with mild disease (median: 3057.6, IQR: 2068.5). Similarly, NLR in patients with moderate to severe disease (median: 5.38, IQR: 8.93) was significantly higher than in patients with mild disease (median: 2.03, IQR: 1.64) [Table 2], [Figure 1]. ROC curves showed ALC <1273 (Sn: 71.6%, Sp: 70.8%, area under the curve [AUC]: 0.238), ANC >3907 (Sn: 73.4%, Sp: 73.5%, AUC: 0.787), and NLR >2.74 (Sn: 78%, Sp: 74.4%, AUC: 0.840) to be good predictors of disease severity at presentation [Figure 2]. Multiple logistic regression adjusted to age, gender, and comorbidities showed a significant association of ANC >3907 with disease severity (adjusted odds ratio [aOR] 5.417, 95% confidence interval [CI] 3.051–9.617). ALC and NLR were not significantly associated with disease severity [Table 3].
|Table 2: Demographic, clinical, and laboratory indices in patients with mild disease versus moderate-to-severe disease|
Click here to view
|Figure 1: Boxplot representing distribution of absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio among patients with moderate-to-severe disease as compared to those with mild disease'|
Click here to view
|Figure 2: Sensitivity-specificity analysis for absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio as predictors of mortality|
Click here to view
|Table 3: Univariable and multivariable logistic regression model for predictors of disease severity|
Click here to view
The values of ALC, ANC, and NLR varied considerably between survivors and nonsurvivors. ALC was significantly lower in nonsurvivors (median: 732.0, IQR: 716.35) compared to survivors (median: 1423.9, IQR: (50.0). ANC was significantly higher in patients who died (median: 7040.25, IQR: (5533.87) compared to those who survived (median: 3448.5, IQR: 295.8). NLR in patients who died (median: 10.05, IQR: 16.14) was also significantly higher than in survivors (median: 2.35, IQR: 2.21) [Table 4]], [Figure 3]. ALC <1183 (Sn: 75.5%, Sp: 68.1%, AUC: 0.199), ANC >4612 (Sn: 81.6%, Sp: 76.7%, AUC: 0.841), and NLR >3.76 (Sn: 81.6%, Sp: 78.9%, AUC: 0.884) performed well as predictors of mortality [Figure 4]. Multivariable Cox proportional hazards regression revealed ALC <1183 (adjusted hazard ratio [aHR]: 2.703, 95% CI: 1.312–5.567), ANC >4612 (aHR: 3.537, 95% CI: 1.558–8.030), and NLR >3.76 (aHR: 3.019, 95% CI: 1.357–6.716) to be significant predictors of the risk of mortality [Table 5].
|Table 4: Demographic, clinical, and laboratory indices in survivors versus nonsurvivors|
Click here to view
|Figure 3: Boxplot representing distribution of absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio among survivors and nonsurvivors|
Click here to view
|Table 5: Univariable and multivariable Cox proportional hazards regression model for predictors of mortality|
Click here to view
|Figure 4: Sensitivity-specificity analysis for absolute lymphocyte count, absolute neutrophil count, and neutrophil-to-lymphocyte ratio as predictors of mortality|
Click here to view
Data were analyzed to determine whether ALC, ANC, and NLR determined the time period of seroconversion in survivors or the time from admission to death in nonsurvivors. Neither of the three variables showed a significant association (data not shown).
| Discussion|| |
In the present study, the data of 328 COVID-19 RTPCR-confirmed hospitalized patients were analyzed for baseline demographic and clinical characteristics and basic laboratory investigations. The independent association of ALC, ANC, and NLR with criticality of the disease and with mortality was screened.
Lymphocytes and their subsets play an important role in maintaining the immune system function and constitute a part of the adaptive immune response against viral infections. Recent studies have reported a higher incidence of lymphopenia in patients with severe disease and have drawn an association with adverse outcomes.,,, The findings from the present study are corroboratory. It was observed in this study that the median ALC was lower in patients with severe disease and among nonsurvivors. In the present study, it was also observed that ALC <1273 predicted disease severity with good sensitivity and specificity and was also independently associated with the odds of severe disease after adjusting for age, gender, and underlying comorbidities. ALC <1183 showed good sensitivity and specificity as a prognostic marker for mortality and emerged as an independent risk factor in the adjusted logistic regression model.
Different mechanisms have explaining lymphopenia have been proposed, including the virus's ability to infect T-cells through the angiotensin-converting enzyme 2 receptors and cluster of differentiation 147-spike proteins, the inflammatory cytokine storm, and exhaustion of T-cells.
On the other hand, neutrophils are a major component of the leukocyte population that activates and migrates from the venous system to the immune organ or system. Neutrophils are a vital component of the innate immune response and are one of the first cells to be recruited and are primarily associated with clearance of pathogens and debris through phagocytosis. Excessive neutrophil counts in peripheral blood of COVID-19 patients have drawn significant attention. Excessive neutrophil counts, exerting their influence on tissue injury through oxidative stress, phagocytosis, and formation of neutrophil NETS, have been shown to correspond with disease severity. Wang et al., in a recent study, have further demonstrated that the dynamics of neutrophil counts in COVID-19 patients during hospitalization exhibited the same trend as the corresponding lung injury. Their study also showed a median ANC of 3400 in patients with severe disease and 2900 in mild disease.
The association of neutrophil counts with severe and fatal disease has been described previously.,, Similar results were observed in the present study with ANC in patients with moderate to severe disease being significantly higher compared to that in patients with mild disease and in nonsurvivors as against survivors. A sensitivity–specificity analysis further showed that ANC >3907 and ANC >4612, respectively, predicted disease severity and mortality with considerable sensitivity and specificity. An independent association was observed in the multivariable logistic regression analysis with disease severity and with mortality.
A composite marker encompassing the effects of both neutrophilia and lymphopenia in the form of NLR was also studied. NLR >2.74 predicted moderate to severe disease with good sensitivity and specificity but was not independently associated with disease severity. NLR > 3.76, however, was observed to be an independent risk factor for mortality with a hazard ratio of 3.019 in the multivariable Cox proportional hazards regression analysis. The prognostic role of NLR has been studied previously. Yang et al. in a study involving 93 patients reported NLR > 3.3 as an independent biomarker for indicating poor clinical outcomes. Liu et al., in a prospective study, observed NLR ≥ 3.13 to be a predictor of critical illness. Wang et al. suggested a much lower cutoff for NLR by observing that the risk of severity and mortality for patients with NLR ≥ 2.14 was significantly higher than those with NLR ≤ 0.48.
These observations from the present study suggest increased neutrophilia and lymphopenia as a marker of disease severity and a forecast of unfavorable outcomes. An increased neutrophil count indicates the degree of the inflammatory response, and the decreased lymphocytes indicate the degree of immune imbalance. Virus-triggered human immune response primarily relies on lymphocytic response. In severe COVID-19 disease, lymphopenia caused by systemic inflammation and other virus-associated factors results in rapid replication and inefficient clearance of the virus. The condition is further complicated by an upregulation of neutrophils and monocyte/macrophage recruitment to the site of infection resulting in excessive infiltration of the lungs. These associations are better represented by the concept of NLR. COVID-19-triggered inflammation hence increased NLR. An elevated NLR was not significantly associated with disease severity at admission but emerged as an independent risk factor for mortality. Elevated NLR hence promotes COVID-19 progression.
The limitations of the present study include relatively smaller size and retrospective design which limited the number of variables that could be considered in the multivariable analysis. Further, outcomes were evaluated at the end of the follow-up period and not during fixed intervals throughout the course of the disease.
| Conclusion|| |
Neutrophilia and lymphopenia predict disease severity at admission as well as are independent risk factors for mortality. NLR emerged as an independent risk factor for mortality, though was not significantly associated with the odds of severe disease at admission. These routine laboratory parameters hence have a potential role in the triage of patients, prognostication, and guiding treatment.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Hui DS, I Azhar E, Madani TA, Ntoumi F, Kock R, Dar O, et al
. The continuing 2019-nCoV epidemic threat of novel coronaviruses to global health – The latest 2019 novel coronavirus outbreak in Wuhan, China. Int J Infect Dis 2020;91:264-6.
Liu J, Liu Y, Xiang P, Pu L, Xiong H, Li C, et al
. Neutrophil-to-lymphocyte ratio predicts critical illness patients with 2019 coronavirus disease in the early stage. Transl Med 2020;18:1-12.
Zhu Z, Cai T, Fan L, Lou K, Hua X, Huang Z, et al
. Clinical value of immune-inflammatory parameters to assess the severity of coronavirus disease 2019. Int J Infect Dis 2020;95:332-9.
Ragab D, Salah Eldin H, Taeimah M, Khattab R, Salem R. The COVID-19 cytokine storm; what we know so far. Front Immunol 2020;11:1446.
Yao XH, Li TY, He ZC, Ping YF, Liu HW, Yu SC, et al
. A pathological report of three COVID-19 cases by minimally invasive autopsies. Zhonghua Bing Li Xue Za Zhi 2020;49:E009.
Barnes BJ, Adrover JM, Baxter-Stoltzfus A, Borczuk A, Cools-Lartigue J, Crawford JM, et al
. Targeting potential drivers of COVID-19: Neutrophil extracellular traps. J Exp Med 2020;217:1-7. Available from: https://pubmed.ncbi.nlm.nih.gov/32302401/
. [Last accessed on 2020 Dec 22].
Fox SE, Akmatbekov A, Harbert JL, Li G, Quincy Brown J, Vander Heide RS. Pulmonary and cardiac pathology in African American patients with COVID-19: An autopsy series from New Orleans. Lancet Respir Med 2020;8:681-6.
Wang J, Li Q, Yin Y, Zhang Y, Cao Y, Lin X, et al
. Excessive neutrophils and neutrophil extracellular traps in COVID-19. Front Immunol 2020;11:2063.
Borges L, Pithon-Curi TC, Curi R, Hatanaka E. COVID-19 and neutrophils: The relationship between hyperinflammation and neutrophil extracellular traps. Mediators Inflamm 2020;2020:1-7.
Nathan C. Neutrophils and COVID-19: Nots, NETs, and knots. J Exp Med 2020;217:1-3
Henry BM, Cheruiyot I, Vikse J, Mutua V, Kipkorir V, Benoit J, et al.
Lymphopenia and neutrophilia at admission predicts severity and mortality in patients with COVID-19: A meta-analysis. Acta Biomed 2020;91:1–16.
Huang C, Wang Y, Li X, Ren L, Zhao J, Hu Y, et al. Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China. Lancet 395(10223):497-06.
Howard R, Kanetsky PA, Egan KM. Exploring the prognostic value of the neutrophil-to-lymphocyte ratio in cancer. Sci Rep 2019;9:1-10.
Yilmaz H, Cakmak M, Inan O, Darcin T, Akcay A. Can neutrophil-lymphocyte ratio be independent risk factor for predicting acute kidney injury in patients with severe sepsis? Ren Fail 2015;37:225-9.
Kaushik R, Gupta M, Sharma M, Jash D, Jain N, Sinha N, et al.
Diagnostic and prognostic role of neutrophil-to-lymphocyte ratio in early and late phase of sepsis. Indian J Crit Care Med 2018;22:660-3.
] [Full text]
de JagerCP, WeverPC, GemenEF, KustersR, van Gageldonk-LafeberAB, van der Poll T, et al. The neutrophil-lymphocyte count ratio in patients with community-acquired pneumonia. PLoS One 2012;7(10):e46561
He R, Lu Z, Zhang L, Fan T, Xiong R, Shen X, et al.
The clinical course and its correlated immune status in COVID-19 pneumonia. J Clin Virol 2020;127:104361.
Wagner J, DuPont A, Larson S, Cash B, Farooq A. Absolute lymphocyte count is a prognostic marker in Covid-19: A retrospective cohort review. Int J Lab Hematol 2020;42:761-5.
Zhao Q, Meng M, Kumar R, Wu Y, Huang J, Deng Y, et al.
Lymphopenia is associated with severe coronavirus disease 2019 (COVID-19) infections: A systemic review and meta-analysis. Int J Infect Dis 2020;96:131-5.
Zhao Y, Nie HX, Hu K, Wu XJ, Zhang YT, Wang MM, et al
. Abnormal immunity of non-survivors with COVID-19: Predictors for mortality. Infect Dis Poverty 2020;9:108.
Kong M, Zhang H, Cao X, Mao X, Lu Z. Higher level of neutrophil-to-lymphocyte is associated with severe COVID-19. Epidemiol Infect 2020;148:e139.
Tavakolpour S, Rakhshandehroo T, Wei EX, Rashidian M. Lymphopenia during the COVID-19 infection: What it shows and what can be learned. Immunol Letters 2020;225:31-2.
Yang AP, Liu JP, Tao WQ, Li HM. The diagnostic and predictive role of NLR, d-NLR and PLR in COVID-19 patients. Int Immunopharmacol 2020;84:106504.
Sun S, Cai X, Wang H, He G, Lin Y, Lu B, et al
. Abnormalities of peripheral blood system in patients with COVID-19 in Wenzhou, China. Clin Chim Acta 2020;507:174-80.
Liu J, Liu Y, Xiang P, Pu L, Xiong H, Li C, et al
. Neutrophil-to-lymphocyte ratio predicts critical illness patients with 2019 coronavirus disease in the early stage. Transl Med 2020;18:206.
Wang S, Fu L, Huang K, Han J, Zhang R, Fu Z. Neutrophil-to-lymphocyte ratio on admission is an independent risk factor for the severity and mortality in patients with coronavirus disease 2019. J Infect 2021;82:e16-8.
Liu J, Li S, Liu J, Liang B, Wang X, Wang H, et al
. Longitudinal characteristics of lymphocyte responses and cytokine profiles in the peripheral blood of SARS-CoV-2 infected patients. EBioMedicine 2020;55:102763.
Channappanavar R, Zhao J, Perlman S. T cell-mediated immune response to respiratory coronaviruses. Immunol Res 2014;59:118-28.
[Figure 1], [Figure 2], [Figure 3], [Figure 4]
[Table 1], [Table 2], [Table 3], [Table 4], [Table 5]