Development of a gut perfusion model as an alternative to the use of live fish

Abstract

An alternative fish model with the principal aim of studying the interaction between fish pathogens and the intestinal tissue was developed. The preparation consisted of an excised gut tractus from rainbow trout (Oncorhynchus mykiss), perfused through cannulation of the aorta intestinalis ventralis with filtered and heparinized Cortland+dextran 1% as the perfusion fluid. This perfusion fluid was delivered by means of a drip. The perfused gut tractus was suspended in a circular bath filled with Ringer solution, which was aerated and kept at a constant temperature of 12 degrees C. Unperfused gut placed in Ringer solution at the same temperature served as the negative control. Perfusion was effective in maintaining the gut in a healthy condition for at least 60 min with only slight oedema and sloughing of the epithelium. Conversely, the unperfused gut revealed excessive tissue degeneration and severe necrosis.

Interactions of virulent and avirulent Yersinia ruckeri strains with isolated gill arches and intestinal explants of rainbow trout Oncorhynchus mykiss

“…The gut perfusion model was used as described by Nematollahi et al (2005). Briefly, the fish were euthanized, the aorta intestinalis ventralis was cannulated after the pyloric caeca and the cranial and caudal ends were severed.…”

“…Subsequently, the gut was flushed with 10 ml PBS and a ligature was tied around both the cranial and caudal end. The gut was suspended in an aerated organ bath surrounded by a 20°C water jacket and perfused with Cortland solution + dextran 1% (Nematollahi et al 2005). For inoculation, ~10 8 CFU in 0.4 ml PBS were injected inside the lumen of the ligated gut.…”

Intestinal Explant Cultures from Gilthead Seabream (Sparus aurata, L.) Allowed the Determination of Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet

“…In this regard, systems have been developed, based on the ex vivo maintenance of intestine fragments, to evaluate successfully the effect of different bacterial strains on intestinal health and provide very reliable information on the interactions between the bacteria and the host. The ex vivo intestinal sack method [ 4 ] has been used to assess the histological and microbial changes in fish in response to bacteria exposure [ 3 , 5 , 6 , 7 , 8 , 9 ]; however, this method is highly restricted by the tissue’s viability under experimental conditions [ 3 , 4 ]. The development of new experimental models based on tissue explants culture has allowed the maintenance of tissue lifespan, as well as immune and histological features [ 10 , 11 , 12 ].…”

Intestinal Explant Cultures from Gilthead Seabream (Sparus Aurata, L.) Allowed Determining the Mucosal Sensitivity to Bacterial Pathogens and the Impact of a Plant Protein Diet

“…In this regard, systems have been developed, based on the ex vivo maintenance of intestine fragments, to evaluate successfully the effect of different bacterial strains on the intestinal health and providing very reliable information on the interactions between the bacteria and the host. The ex vivo intestinal sack method [4] has been used to assess the histological and microbial changes in sh in response to bacteria exposure [3,[5][6][7][8][9]; however, this method is highly restricted by the tissue viability under experimental conditions [3,4]. The development of new experimental models based on tissue explants culture has allowed maintaining the tissue lifespan, as well as immune and histological features [10][11][12].…”