9^th October, 2021; 11am-12noon

Key points of HCFI Expert Round Table

• Platelets are vital from diagnosis, prognosis and monitoring point of view.
• Hematological changes are good navigational tools to diagnose viral infections. 
• Hematology is now restricted to cell counters and flow cytometers. Cell counters are 3-part differential cell counters, 5-part differential cell counters and 7-part differential cell counters. The latest 7-part differential cell counters use laser technology.
• Electronic cell counters are rapid and shorten the time of tests and complete a large number of tests quickly. They also reduce cost of test; maintain and improve the accuracy of tests.
• About 33 parameters of CBC (RBC, WBC and platelet parameters) can be generated with the most sophisticated counters.
• CBC is most frequently done, easily available yet it is the most informative investigation.
• Accurate interpretation of CBC can avoid costly and invasive investigations. 
• Daily monitoring of automated data must be manually validated. A peripheral smear crosscheck is always warranted when reporting.
• Transfusions of all blood products warrants rational use with the help of newer parameters.
• The immature platelet fraction or reticulated platelets are a newer platelet parameter. It contains RNA and can be detected using nucleic acid dyes like new methylene blue. They are the larger strong-staining subset and are typically expressed as a percent of total platelets. 
• IPF differentiates between consumptive vs productive reasons for thrombocytopenia. IPF increases in consumptive reasons but not in the latter. This parameter is increased in the setting of platelet destruction or consumption and decreased with bone marrow failure. IPF decides the status of the marrow, whether it is functioning / responding or not. High IPF means that the marrow is responding and in such a situation, one need not panic even if thrombocytopenia. In dengue, platelet transfusion is not needed even if platelet count is 10,000. If IPF is low, this means that the bone marrow is not working.
• IPF can better differentiate between the causes of thrombocytopenia. It is a reliable parameter even with very low platelet counts and is valuable for effective risk assessment and therapy monitoring of CAD.
• Another new parameter is the platelet larger cell ratio (P-LCR) is indicator of larger (>12fL) circulating platelets (mega platelets, which have very good hemostatic activity). The normal size of platelets is 7.2-11.7 fL. Their normal percentage range is 15-35%. It increases in destructive thrombocytopenia in severe sepsis. P-LCR is inversely related to platelet count and directly related to platelet volume distribution width (PDW) and MPV.
• An old but ignored parameter is mean platelet volume (MPV). The normal MPV ranges from 7.2 to 11.7 fL. When platelet production is increased, young platelets become bigger and more active and MPV levels increase. Increase in MPV during platelet activation is due to change in shape of platelets from biconcave discs to spherical and formation of a pseudopod.
• IPF, P-LCR and MPV must be considered along with platelet count in the clinical condition of the patient to decide if platelet transfusion should be given or not.
• In viral diseases, there will be changes in the WBCs, platelets, RBCs and coagulation and fibrinolytic systems. Covid patients have local or systemic coagulation as was revealed in few autopsies. Changes in the procoagulant and anticoagulant mechanisms caused by the virus.
• RBCs are destroyed by the direct attack of the virus on the heme and similarity of spike protein of the virus and hepcidin which dysregulates the iron metabolism leading to reduction of hemoglobin or hemoglobinopathy.
• WBCs are destroyed due to the direct effect on bone marrow, sepsis, direct effect of the virus on ACE2 of lymphocytes, virus attack on lymph organs, metabolic products such as lactic acid. These mechanisms cause reduction in TLC, apoptosis of lymphocytes.
• Several mechanisms have been postulated for altered coagulation: antiviral antiinflammatory response, injury due to neutrophil extracellular traps (NETs) and activation of different complement pathways. The d-dimer is elevated as is the prothrombin time.
• There is a definite impact of viruses on thrombocytes. 
• Platelets are now regarded as part of the immune system in addition of being capable of forming blood clots.
• Except for viral hemorrhagic fevers and rarely, severe disseminated viral infections, virus induced thrombocytopenia does not lead to significant bleeding and requires judicious platelet transfusions. 
• In viral infections, platelets are reduced because of the direct effect on the bone marrow, immune damage (cytokine storm) and thrombosis (endothelium damage) resulting in decrease in platelet production and increase in platelet consumption (consumptive coagulopathy) resulting in low circulating platelets.
• The early nonspecific immune responses limit multiplication of the virus during the acute phase of the infection. The later specific humoral immune responses help eliminate the virus at the end of the acute phase and subsequently to maintain specific resistance to reinfection.
• Platelets influence the innate immune response through regulation of both the maturation and activation of such innate immune cells as macrophages, neutrophils and dendritic cells.
• Almost every viral disease causes thrombocytopenia. The most frequently associated with thrombocytopenia are dengue, measles, chicken pox, EB virus, mumps and rubella. Herpes simplex, hepatitis B and HTLVIII, Covid-19 may also cause thrombocytopenia.
• Platelets play a main role in fighting against pathogen including viruses in addition to their hemostatic function. The interaction between the virus and platelets through their receptors activates the platelets.
• Viruses cause a decrease in platelet production by infection of megakaryocytes leading to their apoptosis, decreased maturation and ploidy of megakaryocytes or decreased expression of thrombopoietin receptor. The systemic inflammatory response due to viral infection leads to platelet activation. Also, platelet bind to neutrophils forming platelet-neutrophil aggregates, which in turn trigger the phagocytosis of platelets.
• Various mechanisms that contribute to thrombocytopenia in viral infections include aggregation, impaired hemostasis, sequestration and intravascular destruction, platelet expression of pattern recognition receptors (PRR), platelets can induce inflammation and secrete antimicrobial proteins and act as antigen presenting cells.
• Sequestration and intravascular destruction are the primary mechanisms, which leads to correction of thrombocytopenia with use of low dose steroids in post viral cases.
• Platelet expression of pattern recognition receptors (PRR) is the mechanism which justifies the use of steroids in low platelet counts following viral infection as autoimmunity is suppressed.
• It is important to know the technical points as a proper management can only be defined when one knows the why of it. Knowing the how and why of thrombocytopenia helps to draft a proper management plan.
• Isolated platelet transfusions do help in acute crisis management but correcting the underlying cause helps in the long-term management of the situation.
• Hematological analysis is the most easily accessible and helpful tool in assisting the diagnosis of Covid-19. CT scan and molecular diagnosis are expensive.
• In Covid-19, CBC is the primary method to screen suspected Covid-19. TLC can be normal or decreased in the early stages of infection; lymphocytopenia and eosinopenia are often seen; increased CRP and ESR, d-dimer is increased in severely ill patients; alteration of T cell subpopulation can also be observed with CD4+ reduction.
• CRP increases within 4-6 hours of inflammation; the level doubles every 8 hours and peaks at 36-50 hours, which is 100-1000 % higher than the normal value.
• CRP level and duration is proportional to the severity of infection. Most studies have shown that CRP was markedly raised in all patients, especially in the severe and critically ill patients.
• There are morphological changes in neutrophils; apoptotic and immature granulocytes are seen in PS. Cytokine storm and hyperinflammation are implicated as the possible factors causing these changes.
• NLR is a new prognostic indicator. It is an easy-to-use parameter. The cut-off value of NLR is 3.13 (sensitivity 0.875 and specificity 0.717). if NLR is >3.13 and age is >50 years, the patient should be transferred to ICU.
• NLR is a useful parameter for prognostic evaluation and risk stratification of Covid-19 patients. It predicts severe illness in Covid-19 patients in the early stage.
• An elevated platelet-lymphocyte ratio (PLR) may be a prognostic marker in Covid-19.  The cut-off value is 180. It is elevated in severe compared to non-severe Covid-19 patients.
• Presence of widespread microvascular thrombi in both pulmonary and extrapulmonary vessels indicate a systemic prothrombotic state.
• A low to low-normal platelet count is present during peak symptomatic illness, with increased MPV and PDW.
• Desensitization of bone marrow is also a mechanism for thrombocytopenia in Covid-19.
• Platelet-monocyte complexes are formed in severe Covid-19.
• Hematology analysis is helpful in guiding Covid-19 treatment. Along with NLR and CRP, it is of significant clinical value in evaluating disease outcome.
• The requirement of transfusions is low in Covid-19 patients, including in severe cases.
• The hematological parameter of utmost importance in dengue is platelet count. Decrease in platelet count and rise in hematocrit are predictive and recovery parameters of DHF/DSS. 
• CBC in dengue shows high hemoglobin and hematocrit from D3-D10 (highest on D7) due to plasma leakage, lower WBC on D2-D10 (lowest on D4) and lower platelet count D3-D10 (lowest on D6). High monocytes on D1-4 (highest on D2), which can be used to predict severity of dengue infection.
• High atypical lymphocytes between D5 and D9 (highest on D7); predict severity of dengue infection – high in DHF than in dengue fever.
• High eosinophils on D9-10 (highest on D9).
• NLR  is >1 during the first 5 days of the infection and then is reversed on D6 to D9.
• Thrombocytopenia and platelet dysfunction go hand in hand during dengue infection.
• Leukopenia in dengue may be due to virus induced destruction or inhibition of myeloid progenitor cells.
• Thrombocytopenia results from destruction of peripheral platelets and BM megakaryocytes by viruses, which consequently reduce the platelet production.
• Influenza infection is associated with thrombocytopenia which depends on severity of infection. In adults, severe influenza is accompanied by an increased risk of pulmonary thromboembolism and cardiovascular events suggesting that platelet activation occurs during infection.

Excerpts from a presentation “Hematological Changes in prevalent viral fevers” by Prof Dr DP Lokwani, Founder Vice Chancellor MP Medical Sciences University, Consultant Pathologist Jabalpur hospital & Research Center, Ex Prof and Head, NSCB Medical College Jabalpur, MP

Participants

Dr Ashok Gupta

Dr DP Lokwani

Dr KK Kalra

Dr Suneela Garg

Dr Arun Jamkar

Dr B Kapoor

Ms Ira Gupta

Mr Saurabh Aggarwal

Dr S Sharma