Molecular mechanisms underlying the vulnerability of Pacific abalone (Haliotis discus hannai) to Vibrio harveyi infection at higher water temperature

Abstract

Climate change is one of the most important threats to farmed abalone worldwide. Although abalone is more susceptible to vibriosis at higher water temperatures, the molecular mode of action underlying this has not been fully elucidated. Therefore, this study aimed to address the high susceptibility of Halitotis discus hannai to V. harveyi infection using abalone hemocytes exposed to low and high temperatures. Abalone hemocytes were divided into four groups, 20C, 20 V, 25C, and 25 V, depending on co-culture with (V)/without (C) V. harveyi (MOI = 12.8) and incubation temperature (20 °C or 25 °C). After 3 h of incubation, hemocyte viability and phagocytic activity were measured, and RNA sequencing was performed using Illumina Novaseq. The expression of several virulence-related genes in V. harveyi was analyzed using real-time PCR. The viability of hemocytes was significantly decreased in the 25 V group compared to cells in the other groups, whereas phagocytic activity at 25 °C was significantly higher than at 20 °C. Although a number of immune-associated genes were commonly upregulated in abalone hemocyte exposed to V. harveyi, regardless of temperature, pathways and genes regarding pro-inflammatory responses (interleukin-17 and tumor necrosis factor) and apoptosis were significantly overexpressed in the 25 V group compared to the 25C group. Notably, in the apoptosis pathway, genes encoding executor caspases (casp3 and casp7) and pro-apoptotic factor, bax were significantly up-regulated only in the 25 V group, while the apoptosis inhibitor, bcl2L1 was significantly up-regulated only in the 20 V group compared to the control group at the respective temperatures. The co-culture of V. harveyi with abalone hemocytes at 25 °C up-regulated several virulence-related genes involved in quorum sensing (luxS), antioxidant activity (katA, katB, and sodC), motility (flgI), and adherence/invasion (ompU) compared to those at 20 °C. Therefore, our results showed that H. discus hannai hemocytes exposed to V. harveyi at 25 °C were highly stressed by vigorously activated inflammatory responses and that the bacterial pathogen overexpressed several virulence-related genes at the high temperature tested. The transcriptomic profile of both abalone hemocytes and V. harveyi in the present study provide insight into differential host-pathogen interactions depending on the temperature conditions and the molecular backgrounds related to increased abalone vulnerability upon global warming.

Keywords: Cell death; Haliotis discus hannai; Hemocyte; Innate immunity; Phagocytosis; Quorum sensing; Reactive oxygen species; Transcriptome; Vibrio harveyi; Virulence; Water temperature.