Contrast-enhanced ultrasound imaging for assessment of intestinal inflammation in rainbow trout.

The intestine is essential for nutritional uptake and as a barrier to pathogens. Inflammation of the intestine can result from chemical contaminants, dietary irritants, or disease and may lead to serious health consequences, including reduced growth rates or increased pathogen susceptibility. Traditionally, intestinal inflammation in fish has been detected through histology completed post-mortem through excision and processing of the affected tissue. However, in human clinical settings, tools have been developed to assess intestinal inflammation non-invasively. Contrast-enhanced ultrasound (CEUS) imaging is an important tool for measuring inflammation in patients because it is cost-effective and minimally invasive. Specifically, CEUS allows real-time visualization and quantification of vascular perfusion. Changes in blood flow are typical in areas of inflamed or diseased tissue, and by measuring these changes, it is possible to assess the degree of inflammation. We demonstrate that standard CEUS protocols used for small mammals can be used to quantify vascular perfusion in the intestines of rainbow trout. Our resolution was sufficient to measure a significant difference in perfusion between control and TNBS-inflamed trout intestines, with inflamed intestines showing decreased perfusion. The presence of inflammation in the TNBS-treated intestines was verified ex vivo histologically and was characterized by the thickening of intestinal folds. The minimally invasive nature of CEUS imaging presents the opportunity to evaluate intestinal health in novel ways by allowing for longitudinal observations and avoiding mortality in valuable or at-risk specimens. Recent developments of highly portable, cost-effective CEUS systems will allow broad application of this tool, from industry to research.

Chemically-induced trout model of acute intestinal inflammation using TNBS.

Chemically-induced models of intestinal inflammation are a useful tool for the study of immune responses and inflammation. Although well established in mammals, application of these models is currently limited in teleosts. Based on a variety of factors, including genetic diversity, known toxicological sensitivity, and economic importance, we propose salmonids as a model family of fishes for studying intestinal inflammation. We present a rainbow trout model of chemically-induced intestinal inflammation using 2,4,6-trinitrobenzene sulfonic acid (TNBS), assessed through histological analysis of primary and secondary intestinal folding, enterocyte morphology, goblet cell size and frequency, tissue layer thickness, and immune cell infiltration. Twenty-four hours after treatment with one of three concentrations of TNBS, trout developed classic signs of intestinal inflammation, including notably increased thickness of primary and secondary folds, and increased immune cell infiltration as compared to controls. This study provides a simple, reproducible model of rapid TNBS-induction of moderate intestinal inflammation.

Embryonic exposure to predation risk and hatch time variation in fathead minnows.

Organisms are exposed to a wealth of chemical information during their development. Some of these chemical cues indicate present or future dangers, such as the presence of predators that feed on either the developing embryos or their nearby parents. Organisms may use this information to modify their morphology or life-history, including hatching timing, or may retain information about risk until it gains relevance. Previous research has shown predation-induced alterations in hatching among embryonic minnows that were exposed to mechanical-injury-released alarm cues from conspecific embryos. Here, we test whether minnows likewise hatch early in response to alarm cues from injured adult conspecifics. We know that embryonic minnows can detect adult alarm cues and use them to facilitate learned recognition of predators; however, it is unknown whether these adult alarm cues will also induce a change in hatching time. Early hatching may allow animals to rapidly disperse away from potential predators, but late hatching may allow animals to grow and develop structures that allow them to effectively escape when they do hatch. Here, we found here that unlike embryonic fathead minnows (Pimephales promelas) exposed to embryonic cues, embryonic minnows exposed to adult alarm cues do not exhibit early hatching. The ability of embryos to recognize adult alarm cues as a future threat, but not a current one, demonstrates sophisticated ontogenetic specificity in the hatching response of embryonic minnows.