Role of Flavonoid and Terpenoid Biosynthesis Genes in Atriplex halimus in Suppressing Virulence Gene Expression of Giardia Lamblia

Abstract

Giardia lamblia is a protozoan parasite that causes giardiasis, a widespread intestinal infection affecting millions of people worldwide. Current treatments are limited by drug resistance, adverse effects, and treatment failure, emphasizing the need for alternative therapeutic strategies. Atriplex halimus, a halophytic medicinal plant used in traditional medicine, produces diverse secondary metabolites with potential antiparasitic activity. This study investigated flavonoid and terpenoid biosynthesis genes in Atriplex halimus and evaluated the ability of its extracts to inhibit Giardia lamblia growth and suppress virulence-related gene expression. Plant samples were botanically authenticated, and total RNA was extracted from leaf tissues for complementary DNA synthesis. Key biosynthetic genes, including chalcone synthase, flavonol synthase, and terpene synthase, were amplified by RT-PCR, while their expression levels were assessed using qRT-PCR. Methanolic extracts were analyzed for phytochemical composition, and their antigiardial activity was evaluated against cultured Giardia lamblia trophozoites. In addition, the expression of virulence-associated genes, including alpha-1 giardin, variant surface proteins, and cysteine proteases, was examined by qRT-PCR. Molecular analysis identified chalcone synthase, flavonol synthase, and terpene synthase genes with 89%, 85%, and 82% homology, respectively. Gene expression was highest in mature leaves compared with young leaves and stems. Phytochemical analysis confirmed the presence of flavonoids, including quercetin, kaempferol, and rutin, and terpenoids, including beta-sitosterol and stigmasterol. The extract showed dose-dependent antigiardial activity, with an IC50 value of 78 μg/mL after 48 hours. Treatment also significantly reduced the expression of virulence genes by 68% for alpha-1 giardin, 72% for variant surface proteins, and 64% for cysteine proteases. A positive correlation was observed between flavonoid content and suppression of virulence gene expression. These findings suggest that Atriplex halimus possesses active flavonoid and terpenoid biosynthetic pathways that contribute to its antigiardial activity. The plant extract not only inhibited Giardia lamblia growth but also reduced the expression of genes associated with adherence, immune evasion, and pathogenicity. Therefore, Atriplex halimus may represent a promising source of plant-derived compounds for the development of alternative therapies against giardiasis.