Malaria, B lymphocytes and Epstein-Barr virus : emerging concepts on Burkitt's lymphoma pathogenesis

Plasmodium falciparum and Epstein-Barr virus (EBV) infections are recognized co-factors in the genesis of endemic Burkitt's Lymphoma (eBL), the most common paediatric cancer in equatorial Africa. This thesis work examines and discusses interactions between Pfalciparum and EBV that could promote the emergence of eBL. Special focus is given to the effect of malarial antigens on the B cell compartment and of P. falciparum infection on EBV persistence. Plasmodium falciparum infected erythrocytes (IE) express P. falciparum membrane protein 1 (PfEMP1) on their membrane, conferring them with multi-adhesive properties. The demonstration that one of the domains of PfEMP1, the cysteine-rich interdomain region 101 (CIDR1alpha), binds to nonimmune immunoglobulins (Igs), led us to investigate the interaction between IE and human B cells. IE directly adhere to and activate purified B cells from malaria-naive individuals, an interaction involving CIDR1alpha. We have identified and characterized CIDR1alpha as a T cell-independent polyclonal B cell activator that can: (i) induce proliferation and release of lgM, IL-6 and TNF-alpha; (ii) rescue tonsillar B cells from apoptosis; (iii) favour the activation and expansion of the B cell memory compartment, where EBV persists after primary infection; and (iv) bind human IgM and IgG at low affinity but sufficient to lead to B cell receptor signalling. The B cell activation is partially mediated by binding of CIDR1alpha to surface Igs. However, comparison of the gene expression profiles induced by CIDR1alpha and anti-Ig activation, using a cDNA microarray, demonstrated a low degree of homology in the signatures imposed by both stimuli suggesting that CIDR1alpha-activation arises from multiple receptor signalling. Acute malaria infection impairs EBV-specific immune responses and increases the number of EBV-carrying B cells in circulation. Whether this reflects proliferation of previously infected B cells, and/or enhanced virus production and bystander infection of B cells, is not understood. Thus we analysed the effect of malaria on the EBV load in children living in malaria endemic areas. To this end, we studied the occurrence and quantified cell-free EBV-DNA in plasma from Ghanaian children with and without acute malaria infection. Viral DNA was detected in 40% of samples (47% in the malaria-infected and 34% in the non-malaria group, respectively) but was absent in plasma from Ghanaian adults and healthy Italian EBV sero-positive children. The impact of malaria on the control of EBV persistence seems to be evident only before immunity to malaria is fully acquired, in fact adults living in the same area did not have detectable EBV-DNA in the plasma. These findings indicate that viral reactivation is common among children living in malaria-endemic areas, and may contribute to the increased risk for eBL. They also suggest different mechanisms of EBV persistence in these children as compared to adults living in the same region or to children never exposed to malaria. To evaluate the direct relation between EBV viral load and the course of malaria infection, we quantified EBV DNA in plasma and in saliva from Ugandan children with acute malaria (M+) before (day 0) and 14 days after receiving anti-malaria treatment. Controls included children without malaria (M) and children with eBL. EBV DNA was detected in 31% of the plasma samples and in 79% of the saliva samples collected from M+ on day 0, anti-malaria treatment cleared the viral load in plasma without affecting saliva levels. The plasma load varied significantly between the groups; the lowest levels were detected in the M-group, increased in the M+, and reached the highest values in eBL patients. The same trend was evident in the frequency and levels of and-BZLF1 antibodies, indicative of viral reactivation. In the M+ group, the positive plasma samples clustered in a children age group of 7-9 years, the peak-age incidence of eBL. The clearance of circulating EBV after malaria treatment indicates a direct relation between active malaria infection and viral reactivation. In conclusion, this thesis provides unique insight into the mechanisms by which two pathogens, P. falciparum and EBV, exploit the immune system by subverting homeostatic control of B cell proliferation, apoptosis and differentiation thus favouring EBV reactivation