Integrative Study Supports the Role of Trehalose in Carbon Transfer From Fungi to Mycotrophic Orchid

Orchids rely on mycorrhizal symbiosis, especially in the stage of mycoheterotrophic protocorms, which depend on carbon and energy supply from fungi. The transfer of carbon from fungi to orchids is well-documented, but the identity of compounds ensuring this transfer remains elusive. Some evidence has been obtained for the role of amino acids, but there is also vague and neglected evidence for the role of soluble carbohydrates, probably trehalose, which is an abundant fungal carbohydrate. We therefore focused on the possible role of trehalose in carbon and energy transfer. We investigated the common marsh orchid (Dactylorhiza majalis) and its symbiotic fungus Ceratobasidium sp. using a combination of cultivation approaches, high-performance liquid chromatography, application of a specific inhibitor of the enzyme trehalase, and histochemical localization of trehalase activity. We found that axenically grown orchid protocorms possess an efficient, trehalase-dependent, metabolic pathway for utilizing exogenous trehalose, which can be as good a source of carbon and energy as their major endogenous soluble carbohydrates. This is in contrast to non-orchid plants that cannot utilize trehalose to such an extent. In symbiotically grown protocorms and roots of adult orchids, trehalase activity was tightly colocalized with mycorrhizal structures indicating its pronounced role in the mycorrhizal interface. Inhibition of trehalase activity arrested the growth of both symbiotically grown protocorms and trehalose-supported axenic protocorms. Since trehalose constitutes only an inconsiderable part of the endogenous saccharide spectrum of orchids, degradation of fungal trehalose likely takes place in orchid mycorrhiza. Our results strongly support the neglected view of the fungal trehalose, or the glucose produced by its cleavage as compounds transported from fungi to orchids to ensure carbon and energy flow. Therefore, we suggest that not only amino acids, but also soluble carbohydrates are transported. We may propose that the soluble carbohydrates would be a better source of energy for plant metabolism than amino acids, which is partially supported by our finding of the essential role of trehalase.

Effects of Diallyl Trisulfide, an Active Substance from Garlic Essential Oil, on Energy Metabolism in Male Moth Sitotroga cerealella (Olivier)

This study investigated the effects of diallyl trisulfide (DAT), an active substance from garlic essential oil, on the metabolism of the main energy substances of pre- and postmating males of Sitotroga cerealella. Males at 12 h postemergence were fumigated with DAT at a concentration (LC10 = 0.010 µL/L) in a glass jar for 7 h. The main energy metabolites from pre- and postmating males were determined, including protein, triglyceride, glycogen, total soluble sugar, trehalose, and trehalase. The contents of total protein and total soluble sugar and the trehalase activity of premating males were significantly increased following DAT treatment, whereas the contents of protein from the accessory gland, triglyceride, glycogen, and trehalose were significantly decreased after treatment. Additionally, after mating, the total protein and soluble sugar contents were significantly increased and the glycogen content was significantly decreased in the treatment group relative to the levels in controls, but there was no significant difference observed in triglyceride, accessory gland proteins, trehalose content, or trehalase activity between the treatment and control groups. Furthermore, the changes in the main energy substances between pre- and postmating in males after the DAT treatment (∆DAT) were smaller than those in the control group (∆CK). This result indicated that DAT can accelerate the rate of metabolism in males at LC10, leading to the accumulation of greater levels of total soluble sugar to support life activities and to the increased synthesis of proteins to resist an adverse environment.

Biochemical changes of the cereal cyst nematode, Heterodera avenae, at low temperatures

Cereal cyst nematode (Heterodera avenae) diapause is induced by high temperatures and is broken by low temperatures. In this study, metabolic responses were monitored in diapause and non-diapause H. avenae during exposure to 4°C for 10 weeks. The results showed that there was no difference in total carbohydrate content. The content of glycogen and glycerol at 0 week was relatively high but decreased with increased storage time at 4°C. The content of trehalose of the nematode at 10 weeks was significant lower than that at 5 weeks at 4°C. Protein content increased significantly after incubation for 5 and 10 weeks. Esterase and trehalase activity increased with the increasing period at 4°C and showed a significant difference between treatments for esterase activity but there was no significant difference between 5 and 10 weeks for trehalase activity. The SDS-PAGE pattern indicated that a 15.5 kDa protein was absent at 10 weeks and present at 0 and 5 weeks. Esterase isoenzyme patterns of H. avenae showed that at 10 weeks there were four bands: EST 0.21, EST 0.24, EST 0.30 and EST 0.34 (Rf values). EST 0.24 was the common band in the three treatments. Biochemical tests were conducted to correlate with hatching experiments using the same treatments. 2-DE patterns of H. avenae showed that diapause and non-diapause nematodes had 409 and 412 protein spots, respectively, and 19 protein spots were unique: 11 distinct proteins in non-diapause and eight distinct proteins in diapause. This information could be helpful in understanding the diapause mechanism of the cereal cyst nematode.

Trehalase as a possible marker of intestinal ischemia--reperfusion injury.

Different pathological affections of the small intestine cause corresponding morphological and functional changes. The present study was aimed to assess intestinal trehalase activities during ischemia and following reperfusion, correlate them with the pathological changes and determine whether trehalase could be used as a biochemical marker of the intestinal ischemia, ischemia - reperfusion injury. Wistar rats, randomly divided into 5 experimental groups (IR) (each n=15), were subjected to one hour mesenteric ischemia followed by 0, 1, 4, 12 and 24 hours of reperfusion. As a control group sham operated animals were used (n=15). The activity of trehalase was determined using an adapted Dahlqwist method. The range of intestinal injury was determined using histological (histopathological injury index and goblet cell quantification) and immunohistochemical (Ki67, InSitu TUNEL) methods. The highest activities of trehalase were recorded in the control group (C=4.42 ± 0.373 μmol/mg/h). The most altered intestinal histology detected in group IR1 was accompanied by the lowest trehalase activity (IR1=0.97 ± 0.209 μmol/mg/h; p < 0.001 C vs. IR1). Improved histological structure in the remaining reperfusion periods correlated with increase in trehalase activity. Almost normal mucosal histological architecture and 72% of the enzymatic activity were restored after 24 hours of reperfusion (IR24=3.20 ± 0.266 μmol/mg/h; p < 0.01 IR1 vs. IR24). The correlation between intestinal histology and trehalase activities during intestinal injury has been shown. Trehalase activity is closely associated with the status of the histological architecture of the small intestine.