The regulation of digestive enzyme release in the two-spotted field cricket Gryllus bimaculatus (de Geer): effects of endogenous and environmental factors

Insects are the most abundant animal species on earth with a huge economical and ecological impact. In spite of intensive research in the field of integrated pest management there are still a lot of questions concerning the adaptation mechanism of insects to their environment. As the digestive tract displays a putative target for effective pest management, this study worked on the effects of endogenous and environmental factors on digestive enzyme release in the omnivorous cricket, Gryllus bimaculatus. The age-dependent enzyme release of carbohydrases, proteases and lipase correlates with the daily feeding rate of the crickets and peaked between days 2 to 4 in last instar larvae as well as in adult crickets. In contrast, the secretion of chitinase was affected by the moulting cycle of the insects reaching maximum activity at the day of moult. Therefore, chitinase plays only a minor role in food digestion. The cellulase activity in the midgut of G. bimaculatus resulted from an endogenous cellulase and was not caused by bacteria or eukaryotic endosymbionts in the digestive tract. The endoprotease trypsin was stored in the caecal tissue as an inactive precursor, and is secreted to the lumen by exocytosis. Following activation Gryllus-trypsin (~24 kDa) is protected from proteolytic degradation, but there is no endogenous protease inhibitor in the midgut. Gene knockdown by RNA interference was used to analyse the endogenous regulation of digestive enzyme release by the neuropeptides allatostatin A and sulfakinin, which had already been shown to affect feeding in G. bimaculatus. Functional analysis of the AST-A gene was investigated for last instar larvae and adult crickets, whereby female crickets seemed to be more sensitive to this method. The gene suppression of AST-A resulted in a decreased synthesis of amylase, trypsin, aminopeptidase and lipase in the caecal tissue, but enzyme release varied between sexes and developmental stages. The knockdown of SK expression led to an increase of amylase and cellulase secretion in female crickets, and to a reduction of protease and lipase release in males. As food plays a fundamental role in digestive enzyme release, both quality and quantity of nutrition are ample factors. There was always a higher digestive enzyme activity in fed crickets compared to starved ones. Furthermore, starvation resulted in a decrease of enzyme synthesis in the caecal tissue. In general, nutrients in the incubation medium led to a stimulation of digestive enzyme secretion, but in the case of cellulase the presence of both cellubiose in the incubation medium and cellulose in the diet caused a strong decline in cellulase release. Addition of the plant protease inhibitor SBTI to the diet caused a dose-dependent inhibition of protease activity in the caeca, whereby minor concentrations of SBTI were compensated by enzyme hyperproduction. In addition to the food uptake, the daily light-dark cycle seems to affect digestive enzyme release. Crickets started to feed at the beginning of the scotophase, which led to an increase of protease and lipase secretion in larvae and adults. The secretion of carbohydrases was highest during the photophase. This means that enzyme release is not solely affected by the time of food uptake. Temperature is one of the most important environmental factors, but seems to play only a minor role in the release of digestive enzymes. All tested enzymes showed a broad optimal temperature range (30°C-40°C), but there was no difference in the release of amylase or lipase after tissue incubation at 25°C or 35°C. In contrast, trypsin and aminopeptidase showed a higher secretion after incubation at 35°C compared to 25°C. Furthermore, insect rearing at 22°C and 32°C during various developmental stages resulted in a positive acclimation of trypsin secretion to rearing temperature.

Level thesis:degree_level

thesis.doctoral

Grantor dc:publisher

Universität Bayreuth

Year

2013

Author dc:creator

• Weidlich, Sandy

Contributors dc:contributor

• Hoffmann, Klaus H.

Repository record source_url

https://epub.uni-bayreuth.de/id/eprint/102/

OAI identifier oai:identifier

oai:epub.uni-bayreuth.de:102