Isolation of a strain-specific Entamoeba histolytica cDNA clone

Abstract

Entamoeba histolytica is an intestinal parasite causing significant morbidity and mortality worldwide. More tools are needed to understand the epidemiology and molecular pathogenesis of amebiasis. A cDNA library was constructed by using poly(A)+ RNA isolated from an axenic strain of E. histolytica, HM1:IMSS, which expresses a pathogenic isoenzyme pattern (zymodeme). Differential screening of the library yielded a strain-specific 3' polyadenylated cDNA clone, C2, possessing nine 26-nucleotide tandem repeats. RNA and DNA transfer blot analysis of four axenic strains of E. histolytica possessing the same pathogenic zymodeme revealed that the gene is present and expressed in pathogenic E. histolytica HM1:IMSS and 200:NIH but is not present in pathogenic strains HK-9 and Rahman. In addition, Southern blot analysis using the C2 clone showed heterogeneity of genomic organization between HM1:IMSS and 200:NIH. DNA dot blot hybridization analysis demonstrated that cDNA clone C2 was also able to distinguish xenically cultured E. histolytica strains possessing pathogenic zymodemes from those possessing nonpathogenic zymodemes and could detect as few as 100 amebic trophozoites. We conclude that C2 is a strain-specific E. histolytica cDNA clone that, in conjunction with other E. histolytica-specific probes, could serve as a useful epidemiologic tool.

Infection by Human Immunodeficiency Virus-1 Is Not a Risk Factor for Amebiasis

“…Recently, the polymorphic structure of E. histolytica has been described. [32][33][34][35] Polymorphic DNA loci have been characterized both in proteincoding 36,37 and noncoding 33,35,38 sequences. These polymorphic loci are potentially useful in the molecular epidemiology of amebiasis.…”

DNA probes and PCR for diagnosis of parasitic infections

“…Single-copy genomic DNA sequences encoding a 125-kDa surface antigen that differed between pathogenic and nonpathogenic isolates but were 90% similar in nucleic acid sequences made up the basis for differentiation of isolates by DNA probes and restriction enzyme fragment length polymorphism analysis of PCR products (33,121). A DNA clone isolated from a cDNA library containing 26-nucleotide tandem repeats was found to distinguish between xenically cultured pathogenic and nonpathogenic E. histolytica strains; however, this sequence was not present in all axenically cultured pathogenic strains (17). Two pairs of oligonucleotide primers targeted to sequences coding for a 30-kDa surface antigen were the basis of a PCR assay discriminating between pathogenic and nonpathogenic laboratory isolates (120), while another 30-kDa assay featured multiple sets of primers and restriction enzyme analysis to differentiate pathogenic from nonpathogenic laboratory isolates (119).…”

Entamoeba histolytica and amoebic liver abscess in northern Sri Lanka: a public health problem

“…Entamoeba infection can be divided into symptomatic and asymptomatic. Asymptomatic individuals tend to be infected/colonized by mostly non-pathogenic and commensal Entamoeba species, primarily Entamoeba dispar (E. dispar), rather than the pathogenic E. histolytica [15][16][17]. In fact, Herbinger et al (2011) demonstrated that, in a cohort of 5378 travelers returning back to Germany from elsewhere with symptoms of intestinal infections, PCR had detected E. histolytica and E. dispar in 9.7% and 88.3% of cases, respectively; more importantly, however, was the fact that almost all of those with PCR evidence of E. histolytica had symptoms typical of amebiasis, compared with only half with evidence of E. dispar [18].…”

“…The rest of those with E. dispar were determined to be coinfected with other organisms such as Campylobacter spp., Giardia lamblia, and Salmonella typhi. Moreover, some studies have even showed E. dispar colonization in completely asymptomatic HIV-positive individuals, with no evidence of mucosal invasion [15,19].…”