ABSTRACT
Vibrio parahaemolyticus strains recovered from human diarrheal stools (one in 1975 and two in 2001) and environmental sources (four, between 2008 and 2010) were investigated for the presence of virulence genes (trh, tdh, and vpadF), pandemic markers (orf8, toxRS new), and with respect to their pathogenic potential in two systemic infection models. Based only on the presence or absence of these genetic markers, they were classified as follows: the environmental strains were non-pathogenic, whereas among the clinical strains, the one isolated in 1975 was pathogenic (non-pandemic), and the other two were pathogenic (pandemic). The pathogenic potential of the strains was evaluated in mice and Galleria mellonella larvae infection models, and except for the clinical (pathogenic, non-pandemic) isolate, the others produced lethal infection in both organisms, regardless of their source, serotype, and genotype (tdh, orf8, toxRS new, and vpadF). Based on mice and larval mortality rates, the strains were then grouped according to virulence (high, intermediate, and avirulent), and remarkably similar results were obtained by using these models: The clinical strain (pathogenic and non-pandemic) was classified as avirulent, and other strains (four non-pathogenic and two pandemic) were considered of high or intermediate virulence. In summary, these findings demonstrate that G. mellonella larvae can indeed be used as an alternative model to study the pathogenicity of V. parahaemolyticus. Moreover, they raise doubts about the use of traditional virulence markers to predict pathogenesis of the species and show that reliable models are indispensable to determine the pathogenic potential of environmental isolates considered non-pathogenic, based on the absence of the long-standing virulence indicators.
ABSTRACT
Changes in protein levels in different components of the apical junctional complex occur in colorectal cancer (CRC). Claudin3 is one of the main constituents of tight junctions, and its overexpression can increase the paracellular flux of macromolecules, as well as the malignant potential of CRC cells. The aim of this study was to investigate the molecular mechanisms involved in the regulation of claudin3 and its prognostic value in CRC. In silico evaluation in each of the CRC consensus molecular subtypes (CMSs) revealed that high expression levels of CLDN3 (gene encoding claudin3) in CMS2 and CMS3 worsened the patients' longterm survival, whereas a decrease in claudin3 levels concomitant with a reduction in phosphorylation levels of epidermal growth factor receptor (EGFR) and insulinlike growth factor 1 receptor (IGF1R) could be achieved by inhibiting Nglycan biosynthesis in CRC cells. We also observed that specific inactivation of these receptor tyrosine kinases (RTKs) led to a decrease in claudin3 levels, and this regulation seems to be mediated by phospholipase C (PLC) and signal transducer and activator of transcription 3 (STAT3) in CRC cells. RTKs are modulated by their Nlinked glycans, and inhibition of Nglycan biosynthesis decreased the claudin3 levels; therefore, we evaluated the correlation between Nglycogenes and CLDN3 expression levels in each of the CRC molecular subtypes. The CMS1 (MSI immune) subtype concomitantly exhibited low expression levels of CLDN3 and Nglycogenes (MGAT5, ST6GAL1, and B3GNT8), whereas CMS2 (canonical) exhibited high gene expression levels of CLDN3 and Nglycogenes (ST6GAL1 and B3GNT8). A robust positive correlation was also observed between CLDN3 and B3GNT8 expression levels in all CMSs. These results support the hypothesis of a mechanism integrating RTK signaling and Nglycosylation for the regulation of claudin3 levels in CRC, and they suggest that CLDN3 expression can be used to predict the prognosis of patients identified as CMS2 or CMS3.