Exploring Anemia and Hemolysis in COVID-19 Patients


As of February 17, 2023, the global impact of the COVID-19 pandemic remains staggering, with over 673 million infections and 6.8 million deaths recorded worldwide. Belgium, in particular, has been severely affected, reporting 4.7 million cases and 33,000 deaths by the same date.

Amidst this health crisis, healthcare professionals have encountered a myriad of complications associated with COVID-19, including an intriguing and relatively rare phenomenon – cold agglutinin hemolytic anemia.

This article delves into the fascinating world of this autoimmune disorder, shedding light on its prevalence, clinical manifestations, and its intricate connection with SARS-CoV-2 infection.

Understanding Cold Agglutinin Hemolytic Anemia

Cold agglutinin hemolytic anemia is an autoimmune disorder characterized by the presence of autoantibodies, predominantly of the IgM isotype (though rare IgG cases exist), which become active at low temperatures (0°C–4°C), causing the agglutination of red blood cells. This initial agglutination sets in motion the classical complement pathway, leading to extravascular hemolysis, primarily in the liver, as red blood cells opsonized by C3d are phagocytosed.

A key diagnostic indicator for cold agglutinin hemolytic anemia is a positive direct Coombs test (DCT) for C3d, along with a cold agglutinin titer ≥1/64 at 4°C. Notably, the thermal amplitude, rather than IgM antibody titers alone, plays a crucial role in determining the clinical significance and severity of hemolysis.

This condition can be divided into ‘cold agglutinin disease,’ a clonal lymphoproliferative pathology, and ‘cold agglutinin syndrome,’ which is typically polyclonal and secondary to infections, with mycoplasma infections being common in adults and Epstein–Barr virus or cytomegalovirus in children.

Cold Agglutinins in the Context of COVID-19

The relationship between cold agglutinin hemolytic anemia and COVID-19 has piqued the curiosity of medical practitioners. During the early stages of the pandemic’s first wave, two cases of cold agglutinin hemolytic anemia in patients with SARS-CoV-2 infection were identified within a hospital setting.

Both patients tested positive for C3d and IgM, and their cold agglutinin titers were notably elevated (1/64 and 1/2048 at 20°C, respectively). Remarkably, as the COVID-19 infection resolved, hemolysis spontaneously improved, accompanied by IgM-to-IgG seroconversion and the disappearance of cold agglutinins.

This intriguing correlation between COVID-19 and cold agglutinin hemolytic anemia is not isolated. The literature contains reports of various autoimmune cytopenias and skin disorders, such as acrosyndrome and frostbite, which have been attributed to cold agglutinins in COVID-19 patients. This convergence of two distinct medical phenomena raises questions about the underlying mechanisms and their significance in the context of the SARS-CoV-2 virus.

Complement Activation and Inflammation in COVID-19

The complement system, an integral part of the immune response, has garnered significant attention in the pathophysiology of SARS-CoV-2 infection. Several studies have underscored the role of complement activation in COVID-19. These interactions between the complement system, inflammation, and coagulation may provide insights into previously unexplained aspects of the disease’s progression and complications.

A study published in 2020 revealed that 46% of COVID-19 patients exhibited a positive DCT, which correlated with increased anemia and greater transfusion requirements. This finding points to the interplay between complement activation, anemia, and the severity of COVID-19.

Inflammation, Iron Metabolism, and Anemia

Another well-established cause of anemia is inflammation, which impacts iron metabolism. Interleukin-6, a proinflammatory cytokine, plays a pivotal role by increasing the synthesis of hepcidin in the liver. Hepcidin is a key regulator of iron homeostasis. Its production is influenced by factors such as iron stores, anemia, erythropoiesis, and hypoxemia.

Hepcidin binds to ferroportin on enterocytes, macrophages, and hepatocytes, leading to its degradation, thereby blocking the release of iron into the bloodstream. Within cells, iron is bound to ferritin, reflecting the body’s iron stores. Erythroferrone, a recently identified protein produced by erythroblasts via the JAK2-STAT5 pathway, counteracts hepcidin synthesis, making more iron available for erythropoiesis.

The Study’s Objectives

Against this backdrop of complex interactions between COVID-19, cold agglutinin hemolytic anemia, complement activation, and inflammation, a study conducted at the University Hospital of Liège aimed to unravel the mysteries surrounding anemia in SARS-CoV-2 infected patients. The study had several specific objectives:

  • To evaluate the prevalence of positive DCT and autoimmune hemolytic anemia, particularly cold agglutinin anemia, in COVID-19 patients hospitalized between September and December 2020.
  • To investigate potential links between a positive DCT, anemia, and the development of complications and mortality in COVID-19 patients.
  • To explore the origins of anemia and assess the impact of SARS-CoV-2 infection on iron metabolism. Additionally, the study aimed to identify correlations between iron parameters and the occurrence of complications.


The investigation into the relationship between COVID-19 infection and cold agglutinin hemolysis yielded intriguing results, which prompt further exploration. Although some case reports have suggested that anti-SARS-CoV-2 IgM antibodies may exhibit cold agglutinin activity, our study did not establish a clear correlation between COVID-19 infection and cold agglutinin hemolysis. The isolated nature of these cases suggests that this phenomenon may be uncommon in the broader context of COVID-19.

However, our study did align with the findings of Berzuini and colleagues, which indicated that patients with positive direct Coombs test (DCT) results exhibited lower hemoglobin levels upon admission, an increased need for transfusion, and a higher number of erythrocyte units transfused per patient.

Furthermore, these DCT-positive patients experienced longer hospital stays and mechanical ventilation periods. It is essential to note that the study by Berzuini and our study differ in terms of the prevalence of positive DCT cases, with their study reporting a higher rate (46%) compared to our findings (20.3%). This discrepancy may stem from differences in patient populations, as their study focused exclusively on severe COVID-19 cases that required pre-transfusion assessments.

Algassim et al. reported a prevalence of positive DCT of 14.7% for anemic patients in intensive care units (ICU) and 9% for those in general hospitalization. In their study, DCT-positive patients exhibited statistically lower hemoglobin levels and longer hospital stays. Notably, LDH levels were higher in DCT-positive patients, and the presence of spherocytes in these patients led to the conclusion of autoimmune hemolytic anemia. However, our study did not find evidence of hemolysis, distinguishing it from Algassim et al.’s findings.

A recent publication by Hafez et al. also reported results similar to ours, with a prevalence of 20% positive DCT in a cohort of 135 COVID-19 patients. These DCT-positive patients exhibited more pronounced anemia and presented with more severe forms of infection, but there were no biological signs of hemolysis.

It is essential to acknowledge that our study did not collect data on comorbidities, which could potentially introduce an imbalance between DCT-positive and DCT-negative patient groups, contributing to variations in anemia rates.

The etiologies of positive DCT in COVID-19 patients have been discussed, including potential erythrocyte membrane modifications, the influence of the complement system, and the role of medications.

A French study demonstrated a significant decrease in CD35 (complement type 1 receptor) and an increase in C4d expression on erythrocyte surfaces in over 80% of severe COVID-19 patients, suggesting complement system activation. Drug-induced hemolysis was considered unlikely in our cohort because Coombs determinations were conducted upon admission, unaffected by subsequent treatments.

While our study found that DCT-positive patients were more anemic, no evidence of hemolysis was observed, casting doubt on the autoimmune nature of anemia in these patients. Other potential etiologies of anemia in COVID-19 patients include inflammation, iron, vitamin B12, or folic acid deficiency. Iron metabolism investigations in our study revealed a strong association between anemia and inflammation, indicated by elevated ferritin and hepcidin levels, which were highly correlated.

Inflammation plays a pivotal role in anemia development during COVID-19 infection. Interleukins and tumor necrosis factor (TNF) interfere with erythropoiesis, reducing erythropoietin (EPO) production and inducing direct cellular toxicity on renal cells. Our study found that EPO and erythroferrone levels were significantly elevated in DCT-positive patients, suggesting an attempt to compensate for anemia. However, this compensation may be hindered by the prevalence of inflammation, which hampers the release of iron needed for effective erythrocyte regeneration.

The correlations established between iron parameters, hemoglobin, and C-reactive protein (CRP) in our study support the hypothesis that inflammation contributes significantly to anemia in COVID-19 patients. Additionally, studies have identified associations between iron parameters and COVID-19 complications, such as disease severity and mortality.

Meta-analyses have shown that ferritin levels are higher in deceased COVID-19 patients compared to survivors, indicating the prognostic value of iron-related markers. Moreover, hepcidin levels have been linked to disease severity and mortality, with increased mortality observed in patients with elevated hepcidin levels.

Frost et al.’s study on iron metabolism in COVID-19 patients revealed a significant correlation between serum iron levels and disease severity, emphasizing the importance of monitoring iron parameters in COVID-19 management. The study found that hemoglobin levels did not correlate with disease severity but noted low hemoglobin values in non-survivors. Similarly, our study identified high ferritin and low hemoglobin and erythroferrone levels as significantly associated with complications in COVID-19 patients.

Despite these findings, our study has limitations, including a relatively small sample size and variability in the timing of iron parameter measurements. Comorbidities, not accounted for in our study, could also contribute to differences in anemia rates among DCT-positive and DCT-negative patients. Further research with larger cohorts and standardized measurements is needed to elucidate the intricate relationship between COVID-19, anemia, and iron metabolism fully.


As the world continues to grapple with the profound impact of the COVID-19 pandemic, healthcare professionals are continually uncovering new facets of this complex disease. The interplay between SARS-CoV-2 infection, cold agglutinin hemolytic anemia, complement activation, inflammation, and disruptions in iron metabolism highlights the multifaceted nature of COVID-19’s effects on the human body.

Studies like the one conducted at the University Hospital of Liège are crucial for deepening our understanding of these phenomena and improving patient care in the face of this ongoing global health crisis.

reference link : https://journals.sagepub.com/doi/full/10.1177/20406207231199837



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