**3. Epidemiology of amoebiasis and its occurrence in the era of HIV and AIDS**

The epidemiology of amoebiasis around the world is complicated by the existence of three different forms that are morphological identical but genetically distinct and include *E. histolytica* which is a known pathogen, *E. dispar* and *E. moshkovskii* which are non pathogens (Ali et al., 2008). This is particularly relevant to the African continent as well as many other developing countries in the world, including Latin American and Asian countries, where there is lack of specific diagnostic tools. According to some studies conducted in some African countries (Alonzo et al., 1993; Molback et al., 1994; Njoya et al., 1999; Roche et al., 1999) from 6% to 75% of the population carry the parasite. These studies were conducted using microscopic examination giving a general idea on the distribution of the disease in the population. Such results require confirmation by techniques that clearly differentiate *E. histolytica* from *E. dispar*, which is not pathogenic. Countries in Central and Latin America where the parasite displays endemic behavior include Mexico, Brazil, and Ecuador. In Mexico for example, the incidence rate of intestinal amoebiasis from 1995 to 2000 was reported to be between 1000 and 5000 cases/100,000 inhabitants annually. Incidence values from 2002 to 2006 were 1128.8 to 615.85/100,000 inhabitants per year. As in other developing countries, those under 15 years of age were the most frequently affected group, with a notable increase in children aged 5–9 (Ximenez, 2009). In Aracaju, Brazil, Lawson et al (2004) demonstrated *E. histolytica* in 1% of cases whereas *E. dispar* was found in 13% of the cases. Whilst in Pernambuco state, northeastern Brazil *E. dispar* was found in 74.19% of culture positive samples using the PCR method, no *E. histolytica* was reported (Pinheiro *et al*., 2004). In a remote area of Ecuador, Gatti et al (2002) using isoenzyme analysis reported an 18.9% infection rate with *E. histolytica* while 70.3% were infected with *E. dispar*. In the Indian subcontinent, the prevalence of intestinal amoebiasis among hospitalized patients was found to be around 11.7% using microscopy. However, using molecular biology tools such as PCR, *E. histolytica* was shown to be in 3.5% of those infected (Khairnar et al., 2007). In another endemic country such as Bangladesh and using ELISA antigen detection kits, *E. histolytica* prevalence was found to be 4.2% among children living in the urban slums of Dhaka (Haque et al. 2006). Many studies have been conducted in different parts of the world, (Ghosh et al., 2000) but the region most concerned by this problem (Africa) remains unexplored. Thus, the epidemiology of amoebiasis still remains very uncertain particularly in this part of the world.

Following the HIV/AIDS pandemic, numerous studies demonstrated that intestinal parasites such as *Cryptosporidium* sp, Microsporidia sp, *Isospora belli*, and *Cyclospora cayetenensis* were frequently associated with episodes of severe and often fatal diarrhea in both industrialized and poor countries. There have been controversies around the impact of HIV on the occurrence of amebiasis. However, recent data have shown an increase in the occurrence of *E. histolytica* among HIV patients in countries such as Japan, Mexico, Taiwan, and South Africa (Moran et al., 2005; Hung et al., 2008; Samie et al., 2009; Watanabe et al., 2011). With the hall mark of HIV infection being the depletion of CD4+ T cells count (below 200 cells/µl) and the progressive decline of the mucosal immunologic defense mechanisms, HIV/AIDS patients become prone to life-threatening gastrointestinal manifestations such as diarrhea (Stark et al., 2009). Table 1 provides a summary of the prevalence studies reporting *E. histolytica* and/or *E. histolytica / E. dispar* infections in HIV positive individuals in different countries. The association of *E. histolytica* infections with HIV positive individuals in some studies is not clear-cut. In a Mexican study no clear association between *E. histolytica* and

**3. Epidemiology of amoebiasis and its occurrence in the era of HIV and AIDS**  The epidemiology of amoebiasis around the world is complicated by the existence of three different forms that are morphological identical but genetically distinct and include *E. histolytica* which is a known pathogen, *E. dispar* and *E. moshkovskii* which are non pathogens (Ali et al., 2008). This is particularly relevant to the African continent as well as many other developing countries in the world, including Latin American and Asian countries, where there is lack of specific diagnostic tools. According to some studies conducted in some African countries (Alonzo et al., 1993; Molback et al., 1994; Njoya et al., 1999; Roche et al., 1999) from 6% to 75% of the population carry the parasite. These studies were conducted using microscopic examination giving a general idea on the distribution of the disease in the population. Such results require confirmation by techniques that clearly differentiate *E. histolytica* from *E. dispar*, which is not pathogenic. Countries in Central and Latin America where the parasite displays endemic behavior include Mexico, Brazil, and Ecuador. In Mexico for example, the incidence rate of intestinal amoebiasis from 1995 to 2000 was reported to be between 1000 and 5000 cases/100,000 inhabitants annually. Incidence values from 2002 to 2006 were 1128.8 to 615.85/100,000 inhabitants per year. As in other developing countries, those under 15 years of age were the most frequently affected group, with a notable increase in children aged 5–9 (Ximenez, 2009). In Aracaju, Brazil, Lawson et al (2004) demonstrated *E. histolytica* in 1% of cases whereas *E. dispar* was found in 13% of the cases. Whilst in Pernambuco state, northeastern Brazil *E. dispar* was found in 74.19% of culture positive samples using the PCR method, no *E. histolytica* was reported (Pinheiro *et al*., 2004). In a remote area of Ecuador, Gatti et al (2002) using isoenzyme analysis reported an 18.9% infection rate with *E. histolytica* while 70.3% were infected with *E. dispar*. In the Indian subcontinent, the prevalence of intestinal amoebiasis among hospitalized patients was found to be around 11.7% using microscopy. However, using molecular biology tools such as PCR, *E. histolytica* was shown to be in 3.5% of those infected (Khairnar et al., 2007). In another endemic country such as Bangladesh and using ELISA antigen detection kits, *E. histolytica* prevalence was found to be 4.2% among children living in the urban slums of Dhaka (Haque et al. 2006). Many studies have been conducted in different parts of the world, (Ghosh et al., 2000) but the region most concerned by this problem (Africa) remains unexplored. Thus, the epidemiology of amoebiasis still remains very uncertain particularly

Following the HIV/AIDS pandemic, numerous studies demonstrated that intestinal parasites such as *Cryptosporidium* sp, Microsporidia sp, *Isospora belli*, and *Cyclospora cayetenensis* were frequently associated with episodes of severe and often fatal diarrhea in both industrialized and poor countries. There have been controversies around the impact of HIV on the occurrence of amebiasis. However, recent data have shown an increase in the occurrence of *E. histolytica* among HIV patients in countries such as Japan, Mexico, Taiwan, and South Africa (Moran et al., 2005; Hung et al., 2008; Samie et al., 2009; Watanabe et al., 2011). With the hall mark of HIV infection being the depletion of CD4+ T cells count (below 200 cells/µl) and the progressive decline of the mucosal immunologic defense mechanisms, HIV/AIDS patients become prone to life-threatening gastrointestinal manifestations such as diarrhea (Stark et al., 2009). Table 1 provides a summary of the prevalence studies reporting *E. histolytica* and/or *E. histolytica / E. dispar* infections in HIV positive individuals in different countries. The association of *E. histolytica* infections with HIV positive individuals in some studies is not clear-cut. In a Mexican study no clear association between *E. histolytica* and

in this part of the world.

HIV has been noted. In this study, the prevalence of *E. histolytica* in HIV/AIDS patients was 25.3% compared to 18.4% in a control HIV-group (Moran et al., 2005). Other studies in South American countries have shown no obvious association. However, a significant association between high levels of serum anti-*E. histolytica* antibodies and the presence of *E. histolytica* in the stool has been noted in studies from both Vietnam (Blessman et al., 2006) and Africa (Stauffer et al., 2006). In a South African study in the Vhembe district in the northern part of the country, a positive association between *E. histolytica* infection and HIV-positive individuals has been indicated. Among the HIV-positive individuals, those with CD4+ count less than 200 cells/µl, were relatively more likely to be seropositive for *E. histolytica* (Samie et al., 2010). In a Chinese study, a higher seroprevalence of *E. histolytica* infections was also found in HIV-infected patients (Chen et al, 2007). Furthermore, two studies conducted in Taiwan revealed a positive association as well (Hung et al., 2005; Tsai et al., 2006).


Table 1. Global prevalence of *E. histolytica* in HIV-infected and non-infected persons.

Over the past decade, there has been an increasingly reported risk of amebiasis in East Asian countries like Japan, Taiwan and South Korea particularly among men who have sex with men (MSM) probably due to oral-anal sexual contact (Hung et al., 2008; Watanabe et al., 2011). In Japan, *E. histolytica* often occur in institutions of mentally retarded individuals where outbreacks of amebiasis have been described with the prevalence rate and positive serology rate as high as 38.2% and 67.1%, respectively (Nishise et al., 2010) and has been

Amoebiasis in the Tropics: Epidemiology and Pathogenesis 209

takes between 1-4 weeks to perform and requires sophisticated laboratory equipment making it not feasible as a routine procedure especially in the developing world where *E. histolytica* is rampant. The rate of success of *E. histolytica* culture in reference laboratories has been reported to be between 50 and 70%. Moreover, isoenzyme (zymodeme) analysis is labor intensive, costly and often produces false-negative results for many microscopy

Serological methods may be useful diagnostically to detect infections with *E. histolytica* in developed countries where infections are not as common as in endemic developing nations (Ohnishi et al., 1997). In developing countries individuals are constantly exposed to *E. histolytica* making serological tests unable to definitively distinguish past from current infections (Caballero et al., 1994). Amoebic serology is highly sensitive and specific for the diagnosis of ALA (Zengzhu et al., 1999). Conversely, a study of asymptomatic individuals living in an *E. histolytica* endemic area of Vietnam revealed that about 83% of those infected had detectable anti-amoebic antibodies (Blessmann et al., 2002). Several assays for the detection of antibodies to *E. histolytica* infections have been developed (Table 2). These include: indirect hemagglutination (IHA), latex agglutination, immunoelectrophoresis, counterimmunoelectrophoresis (CIE), the amebic gel diffusion test, immunodiffusion, complement fixation, indirect immunofluorescence assay (IFA), and enzyme-linked immunosorbent assay (ELISA). With the exception of ELISA, all the other tests have been either costly to perform (Complement fixation), less sensitive and nonspecific (IHA and Latex agglutination test), time consuming (immunodiffusion) or requires skills in culture

**Serological Assay Sensitivity (%) Specificity (%) Reference(s)** 

IHA 100*a*, 99 90.9-100*a* , 99.8 Pillai et al., 1999;

Novagnost *Entamoeba* IgG >95 >95 Manufacturer's

I.H.A. Amoebiasis 93 97.5 Robert et al., 1990

Table 2. List of some of the commercially available antibody assays used for the diagnosis of

ELISA is a reliable, easy to perform and rapid method for the diagnosis of *E. histolytica*  infections especially in developing countries. It has been used widely for the study of the epidemiology and diagnosis of symptomatic amoebiasis (intestinal and/or extraintestinal). An ELISA to detect antibodies to *E. histolytica* has been shown to be 97.9% sensitive and 94.8% specific for detection of *E. histolytica* antibodies in ALA patients in a non endemic country (Hira et al., 2001). Unlike IgG, immunoglobulin M (IgM) is short lived and does not

95 97 Manufacturer's

100, 97.7-100 (100) 95.6, 97.4 (100) Manufacturer's

Hira et al., 2001

recommendation

recommendation

recommendation; Knappik et al., 2005

positive stool specimens (Strachan et al., 1988).

and antigen preparation (IFA) (Fotedar et al., 2007).

Amebiasis Serology microplate

RIDASCREEN *Entamoeba* (IgG

ELISA

detection)

amoebiasis.

occurring more often in HIV positive patients (Watanabe et al., 2011). In addition to HIV/AIDS, the increasing use of organ transplants and other immunosuppressed conditions such as neutropenia have been considered important risk factor for invasive amoebiasis in many countries. In Colombia for example, a study of organ transplant patients revealed that about 24.7% had detectable antiamoebic antibodies (Reyes et al., 2006) whereas in another study 14.3% neutropenic patients were found to have antiamoebic antibodies (Cardona et al., 2004).

Certain risk behaviors, such as homosexual relations and practicing oro-anal sex, can exacerbate the possibility of acquiring *E. histolytica* infections as well as other intestinal parasites such as *Cryptosporidium* spp., where the symptomatic pictures are more severe than those of immunocompetent individuals (Tatiana et al., 2008; Hung 2008). A recent study in Vietnam had indicated that socio-economic and personal hygiene factors determined infection with *E. histolytica*, rather than exposure to human and animal excreta in agricultural activities (Pham duc et al., 2011). In a study in Bangladesh, it was shown that wet environment is not the only factor that affects the detection curve of *E. histolytica*, but anti-Carbohydrate Recognition Domain IgA level in the gut is another determining factor for its occurrence in a closed population (Haque et al., 2006). Although, numerous seroprevalence studies suggest that HIV/AIDS individuals are at a higher risk of *E. histolytica* infections and are therefore more likely to develop symptomatic infections or severe forms of the disease, modest data exist to support these findings and further research is needed to confirm this hypothesis.
