ORIGINAL ABSTRACT

UNLABELLED: Intensive use of glyphosate-based herbicides in genetically modified (GM) and non-GM agriculture has resulted in widespread glyphosate-resistant weeds. In response, the agricultural biotechnology industry has launched GM crops tolerant to glyphosate plus 2,4-D and glyphosate plus dicamba. Consequently, people are increasingly exposed to mixtures of these herbicides, yet no studies have been conducted to assess health risks. Intestinal structure and integrity, as well as gut microbiome composition and function, are recognised contributors to disease. Therefore, we investigated the effects of glyphosate alone and in combination with 2,4-D and dicamba on gut structure and function. Pregnant Wistar rats were administered via drinking water from gestation day 6 with glyphosate at the European Union (EU) no observed adverse effect level (NOAEL: 50 mg/kg bw/day) and acceptable daily intake (ADI: 0.5 mg/kg bw/day), and with an ADI mixture of glyphosate (0.5 mg/kg bw/day), 2,4-D (0.02 mg/kg bw/day) and dicamba (0.3 mg/kg bw/day). Offspring continued this regimen for 13 weeks post-weaning. Large and small intestinal tissues and gut content were isolated and analysed for inflammation, gut epithelial integrity, oxidative stress, microbiota composition and histopathology. The glyphosate NOAEL and, to a greater degree, the glyphosate, dicamba, 2,4-D mixture resulted in increased gut inflammation and permeability, associated with oxidative stress and altered microbial composition. Histological analysis confirmed structural alterations and inflammation in large and small intestine. Effects were more pronounced in large intestine and females. Our results identify exposure to glyphosate alone and a mixture of glyphosate, 2,4-D and dicamba as risk factors for gut structure and function dysbiosis. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s00204-026-04409-9.