AIG1 affects in vitro and in vivo virulence in clinical isolates of <i>Entamoeba histolytica</i>

<div><p>The disease state of amebiasis, caused by <i>Entamoeba histolytica</i>, varies from asymptomatic to severe manifestations that include dysentery and extraintestinal abscesses. The virulence factors of the pathogen, and host defense mechanisms, contribute to the outcomes of infection; however, the underlying genetic factors, which affect clinical outcomes, remain to be fully elucidated. To identify these genetic factors in <i>E</i>. <i>histolytica</i>, we used Illumina next-generation sequencing to conduct a comparative genomic analysis of two clinical isolates obtained from diarrheal and asymptomatic patients (strains KU50 and KU27, respectively). By mapping KU50 and KU27 reads to the genome of a reference HM-1:IMSS strain, we identified two genes (EHI_089440 and EHI_176590) that were absent in strain KU27. In KU27, a single <i>AIG1</i> (<i>a</i><i>vrRpt2</i>-<u>i</u>nduced <u>g</u>ene 1) family gene (EHI_176590) was found to be deleted, from a tandem array of three <i>AIG1</i> genes, by homologous recombination between the two flanking genes. Overexpression of the EHI_176590 gene, in strain HM-1:IMSS cl6, resulted in increased formation of cell-surface protrusions and enhanced adhesion to human erythrocytes. The EHI_176590 gene was detected by PCR in 56% of stool samples from symptomatic patients infected with <i>E</i>. <i>histolytica</i>, but only in 15% of stool samples from asymptomatic individuals. This suggests that the presence of the EHI_176590 gene is correlated with the outcomes of infection. Taken together, these data strongly indicate that the AIG1 family protein plays a pivotal role in <i>E</i>. <i>histolytica</i> virulence via regulation of host cell adhesion. Our <i>in-vivo</i> experiments, using a hamster liver abscess model, showed that overexpression or gene silencing of EHI_176590 reduced and increased liver abscess formation, respectively. This suggests that the <i>AIG1</i> genes may have contrasting roles in virulence depending on the genetic background of the parasite and host environment.</p>