Asymptomatic malaria (presence of malaria parasite without symptoms) is prevalent in highly endemic areas in Africa [24
]. It is suggested that asymptomatic parasitemia confers partial immunity and protection against symptomatic malaria [5
]. On the other hand, asymptomatic individuals constitute a remarkably large reservoir for malaria transmission or could be a precursor in the progression of symptomatic form of malaria. This study was conducted after reviewing data generated in a previous study [19
] and finding a remarkably high number (36%) of asymptomatic patients recorded as healthy controls. We acknowledge that the cross sectional measurement of parasitemia and biological factor levels in host blood and the lack of follow-ups to determine the true impact of asymptomatic parasitemia is a limitation of this study. Nevertheless, the importance of our observation is predicated on recent studies that indicate that individuals with asymptomatic parasitemia may have a higher risk of developing symptomatic malaria than those without evidence of parasitemia [26
]. In our study, the AC group had lower hemoglobin level compared to the HC group (). This lower level of hemoglobin could be due to the presence of chronically low parasitemia causing extensive destruction of red blood cells. Many asymptomatic infections are undetected by microscopy and thus most asymptomatic individual do not receive required treatment thereby rendering them susceptible to the risks associated with both symptomatic and asymptomatic malaria. This persistent problem has resulted in the proposed presumptive intermittent treatment strategy for asymptomatic individuals, regardless of their malaria infection status, to reduce malaria morbidity and mortality [16
]. This therapeutic method may adversely result in accelerating the development of resistance against anti-malarial drugs suggesting an urgent need to determine the true infection status before treatment. This approach is useful in decreasing the prevalence of asymptomatic malaria and the negative consequence of drug resistance to anti-malarial drugs.
The present study examined a broad range of immune mediators, including cytokines, chemokines, and markers of apoptosis and angiogenesis in healthy and asymptomatic malaria carrier pregnant women. The immunological relevance of these biological factors associated with asymptomatic malaria have been previously established [27
]. Due to the lack of concurrent studies of inflammatory, apoptotic, and angiogenic factors in appropriate time-matched symptomatic patients, the results are discussed in context. Proinflammatory Th1-type cytokines are thought to be critical to the control of exoerythrocytic and erythrocytic P. falciparum
], but their exaggerated production may also contribute to organ damage, particularly in the brain in cerebral malaria cases. It is widely accepted that anti-inflammatory Th2-type cytokines downregulate Th1-derived cytokines. Th2-type cytokines, such as IL-10, regulate Th1-cytokines and prevent severe forms of malaria in some animal models [31
]. The regulation of TNF levels by IL-10 appears to contribute to the prevention of severe malarial anemia in humans [32
]. However, the role that IL-10 plays may depend on its levels, since very high levels of IL-10 have been associated with severe malaria in humans [35
] and some animal models [36
]. In our analysis, higher levels of IL-10 were observed in the AC group than the HC group. IL-10 is a key cytokine involved in both protection and immunopathology during malaria. High levels of IL-10 may be beneficial by reducing the inflammatory response, but may be detrimental by decreasing antiparasitic cellular immune responses. IL-10 is an anti-inflammatory cytokine that acts in part by blocking monocyte and macrophage production of inflammatory cytokines such as IL-6, TNF, and IL-l [37
]. Animal studies have suggested that IL-10 may play a regulatory role that modulates susceptibility to parasite infection. In particular, IL-10 inhibits microbicidal activity of IFN-γ
-treated macrophages against intracellular parasites such as Toxoplasma gondii
], Trypanosoma cruzi
] and Leishmania major
], and the killing of extracellular Schistosoma mansoni schistosomulas
]. IL-10 has been previously observed to be elevated during malarial episodes in nonpregnant [42
] and pregnant women [44
]. Both increased and decreased levels of IL-10 have been associated with poor malaria outcomes. Low levels of IL-10 or low IL-10 to TNF ratios were associated with severe malarial anemia in African children [32
], while high IL-10 levels were associated with reduced ability to eliminate malaria parasitemia in Tanzanian children [45
]. Thus, the high levels of IL-10 observed in the AC group may reduce their ability to eliminate or clear malaria parasitemia as seen in the HC.
Cytokines may stimulate beneficial immunological responses in falciparum malaria by inducing acute phase responses, inhibition of parasitic growth, and clearance of vascular parasites and debris. Neutrophils are stimulated by cytokines such as TNF or INF-γ
in order to increase parasite destruction [46
]. In the present study, it seems that G-CSF may be involved in controlling P. falciparum
parasitemia, which ultimately results in the asymptomatic condition observed in our AC group. G-CSF has both anti-infective and anti-inflammatory effects [47
]. G-CSF enhances phagocytosis and increases chemotaxis and antibody-dependent cytotoxicity suggesting its involvement in anti-malarial defense by activating neutrophils and acting as an immunomodulatory factor [47
]. Surprisingly, G-CSF levels were elevated in the AC compared to HC suggesting that this factor may be involved in maintaining low parasitemia levels observed in the asymptomatic individual. Functional studies on the association of G-CSF and IL-10 with asymptomatic malaria are much needed to reveal their contribution to the spread and severity of malaria.
Maternal leukocytes produce predominately Th2-type cytokines shifting the cytokine balance towards Th2-type response during pregnancy to prevent initiation of inflammatory that might damage the integrity of the placental barrier [17
]. In response to invading pathogens,Th1-type cytokines may be produced to reverse the Th2-type bias [18
] to aid in elimination of parasites by enhancing phagocytic activity of macrophages, generating reactive oxygen intermediates, and stimulating the proliferation of T cells [48
]. However, our median ratio result indicates that there was significance bias towards a Th2-type profile in the AC group with low pro- to anti-inflammatory ratios compared to the HC group (). This deviation in immune response towards a Th2 bias may explain the asymptomatic observation in the AC group due to the fact that this group has a less robust Th1-type response to eliminate malaria parasites compared to the HC group. In addition, the Th2 bias may leave pregnant AC more open to Th1-dependent infections that may result in increased morbidity in pregnancy.
We acknowledge that this study has limitations that can be investigated in future studies. The current study did not include cytokine profiles of nonpregnant women and pregnant women with malaria as well as asymptomatic nonpregnant women to determine whether our observations were unique to pregnant women. However, IL-10 has been observed to be elevated during malarial episodes in nonpregnant [42
] and pregnant women [44
]. Although our study lacked malaria positive pregnant women, comparing the results to other studies in which malaria positive pregnant women were included suggests that the levels of peripheral IL-10 in the AC group were lower than pregnant women with placental malaria [49
]. In addition, studies in murine models of malaria have suggested that IL-10 response during infection may be associated with the disease exacerbation [36
To the best of our knowledge, there are no comprehensive prospective studies that describe cytokine profile during pregnancy, therefore, we do not know whether the observation made in this study may have been influenced by pregnancy-specific conditions. However, the increase in asymptomatic cases [19
] highlights the alarming potential of malaria as a serious public health issue underscoring the need for tighter surveillance [50
]. Asymptomatic individuals not only serve as a source of maintenance of intense malaria transmission during the peak season but could also perpetuate resistance through the inappropriate use of drugs [51
In summary, cytokine, chemokine, and growth factor evaluation in asymptomatic healthy pregnant women reveal an association between increased plasma concentrations of IL-10 and G-CSF. Thus, susceptibility to symptomatic malaria in pregnancy may be associated with high levels of IL-10 and G-CSF and dysregulation of pro and anti-inflammatory factors in pregnant women. In the absence of studies explaining the functional implication of this finding for the asymptomatic condition observed in these pregnant women, we propose that assessment of these biological factors in conjunction with rapid detection of P. falciparum HRP-II and PCR-based methods could be a useful tool for predicting risk for asymptomatic malaria carriage.