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Abstract The rise of functional diversity through gene duplication contributed to the adaption of organisms to various environments. Here we investigate the evolution of putative cellulases of the subfamily 2 of glycoside hydrolase family 5 (GH5_2) in the Cerambycidae (longhorned beetles), a megadiverse assemblage of mostly xylophagous beetles. Cerambycidae originally acquired GH5_2 from a bacterial donor through horizontal gene transfer (HGT), and extant species harbor multiple copies that arose from gene duplication. We ask how these digestive enzymes contributed to the ability of these beetles to feed on wood. We analyzed 113 GH5_2, including the functional characterization of 52 of them, derived from 25 species covering most subfamilies of Cerambycidae. Ancestral gene duplications led to five well-defined groups with distinct substrate specificity, allowing these beetles to break down, in addition to cellulose, polysaccharides that are abundant in plant cell walls (PCWs), namely, xyloglucan, xylan, and mannans. Resurrecting the ancestral enzyme originally acquired by HGT, we show it was a cellulase that was able to break down glucomannan and xylan. Finally, recent gene duplications further expanded the catalytic repertoire of cerambycid GH5_2, giving rise to enzymes that favor transglycosylation over hydrolysis. We suggest that HGT and gene duplication, which shaped the evolution of GH5_2, played a central role in the ability of cerambycid beetles to use a PCW-rich diet and may have contributed to their successful radiation.

The rice weevil, Sitophilus oryzae L. (Coleoptera: Curculionidae), is a formidable global pest of storage grains. This study investigated how the interplay between the physicochemical grain parameters of wheat cultivars and the pest’s physiological response modulates the resistance to the rice weevil. A forced-choice test evaluated the susceptibility of wheat cultivars based on parameters viz., percent weight loss of grains, adult emergence, mean developmental period, and susceptibility index. Among the wheat cultivars, HI 1544 and GW 322 were the least susceptible, while, PBW 343, HD 1563, and HI 8381 were highly susceptible. A significant positive correlation was noticed between percent weight loss and factors such as adult emergence, mean developmental period, total soluble sugars, starch, and insect digestive enzymes. In contrast, grain damage was negatively correlated with grain hardness, protein, amylolytic activity, and phenol content of grains. Principal component Analysis (PCA) accounted for 73.65% of the observed variation, effectively delineating key traits. Agglomerative Hierarchical Clustering (AHC) classified wheat cultivars into two categories based on their susceptibility to rice weevil. We recommend prioritizing less susceptible cultivars (e.g., HI 1544, GW 322) for breeding programs and infestation-prone regions. The findings emphasise that a holistic analysis of physicochemical and biological parameters offers a robust framework for screening resistance of wheat cultivars to rice weevil S. oryzae .

In the carnivorous plant Venus flytrap (Dionaea muscipula), the sequence of events after prey capture resembles the well-known plant defence signalling pathway in response to pathogen or herbivore attack. Here, we used wounding to mimic prey capture to show the similarities and differences between botanical carnivory and plant defence mechanisms. We monitored movement, electrical signalling, jasmonate accumulation and digestive enzyme secretion in local and distal (systemic) traps in response to prey capture, the mechanical stimulation of trigger hairs and wounding. The Venus flytrap cannot discriminate between wounding and mechanical trigger hair stimulation. Both induced the same action potentials, rapid trap closure, hermetic trap sealing, the accumulation of jasmonic acid (JA) and its isoleucine conjugate (JA-Ile), and the secretion of proteases (aspartic and cysteine proteases), phosphatases and type I chitinase. The jasmonate accumulation and enzyme secretion were confined to the local traps, to which the stimulus was applied, which correlates with the propagation of electrical signals and the absence of a systemic response in the Venus flytrap. In contrast to plant defence mechanisms, the absence of a systemic response in carnivorous plant may represent a resource-saving strategy. During prey capture, it could be quite expensive to produce digestive enzymes in the traps on the plant without prey.

Background Colibacillosis caused by enterotoxigenic Escherichia coli ( E. coli ) results in economic losses in the poultry industry. Antibiotics are usually used to control colibacillosis, however, E. coli has varying degrees of resistance to different antibiotics. Therefore the use of probiotics is becoming accepted as an alternative to antibiotics. In this study, we evaluated the effects of Clostridium butyricum ( C. butyricum ) on growth performance, immune response, intestinal barrier function, and digestive enzyme activity in broiler chickens challenged with Escherichia coli ( E. coli ) K88. Methods The chickens were randomly divided into four treatment groups for 28 days. Negative control treatment (NC) consisted of birds fed a basal diet without E. coli K88 challenge and positive control treatment (PC) consisted of birds fed a basal diet and challenged with E. coli K88. C. butyricum probiotic treatment (CB) consisted of birds fed a diet containing 2 × 10 7  cfu C. butyricum /kg of diet and challenged with E. coli K88. Colistin sulfate antibiotic treatment (CS) consisted of birds fed a diet containing 20 mg colistin sulfate/kg of diet and challenged with E. coli K88. Results The body weight (BW) and average day gain (ADG) in the broilers of CB group were higher ( P  < 0.05) than the broilers in the PC group overall except the ADG in the 14-21 d post-challenge. The birds in CB treatment had higher ( P  < 0.05) concentration of tumor necrosis factor-α (TNF-α) at 3 and 7 d post-challenge, and higher ( P  < 0.05) concentration of interleukin-4 (IL-4) at 14 d post-challenge than those in the PC treatment group. The concentration of serum endotoxin in CB birds was lower ( P  < 0.05) at 21 d post-challenge, and the concentrations of serum diamine oxidase in CB birds were lower (P < 0.05) at 14 and 21 d post-challenge than in PC birds. Birds in CB treatment group had higher ( P  < 0.05) jejunum villi height than those in PC, NC, or CS treatment at 7, 14, and 21 d post-challenge. In comparison to PC birds, the CB birds had lower ( P  < 0.05) jejunum crypt depth during the whole experiment. The birds in CB or CS treatment group had higher ( P  < 0.05) activities of amylase and protease at 3, 7, and 14 d post-challenge, and higher ( P  < 0.05) activity of lipase at 3, 7 d post-challenge than PC birds. Conclusions In all, these results indicate that dietary supplementation with C. butyricum promotes immune response, improves intestinal barrier function, and digestive enzyme activities in broiler chickens challenged with E. coli K88. There is no significant difference between the C. butyricum probiotic treatment and the colistin sulfate antibiotic treatment. Therefore, the C. butyricum probiotic may be an alternative to antibiotic for broiler chickens.

Plant-derived bioactive compounds with promising nutritional and therapeutic attributes (phytogenics) are among the top priorities in the aquaculture sector. Therefore, the impact of thymol (Thy) and/or thymoquinone (ThQ) on the growth, immune response antioxidant capacity, and Aeromonas sobria (A. sobria) resistance of Nile tilapia was investigated. Four fish groups were fed a control diet and three basal diets supplemented with 200 mg/kg diet of Thy or ThQ and a blend of both Thy and ThQ at a level of 200 mg/kg diet each. At the end of the feeding trial (12 weeks), the tilapias were challenged intraperitoneally with virulent A. sobria (2.5 × 108 CFU/mL) harboring aerolysin (aero) and hemolysin (hly) genes. The results revealed that tilapias fed diets fortified with a combination of Thy and ThQ displayed significantly enhanced growth rate and feed conversion ratio. Notably, the expression of the genes encoding digestive enzymes (pepsinogen, chymotrypsinogen, α-amylase and lipase) and muscle and intestinal antioxidant enzymes (glutathione peroxidase, catalase and superoxide dismutase) was significantly upregulated in Thy/ThQ-fed fish. An excessive inflammatory response was subsided more prominently in the group administrated Thy/ThQ as supported by the downregulation of il-β, il-6 and il-8 genes and in contrast, the upregulation of the anti-inflammatory il-10 gene. Remarkably, dietary inclusion of Thy/ThQ augmented the expression of autophagy-related genes, whilst it downregulated that of mtor gene improving the autophagy process. Furthermore, Thy/ThQ protective effect against A. sobria was evidenced via downregulating the expression of its aero and hly virulence genes with higher fish survival rates. Overall, the current study encouraged the inclusion of Thy/ThQ in fish diets to boost their growth rates, promote digestive and antioxidant genes expression, improve their immune responses and provide defense against A. sorbia infections with great economic benefits.

A 95-day feeding trial was conducted to evaluate the outcomes of feeding Bacillus subtilis fermented Azolla (BSFA) on nonspecific immunity, antioxidative status, intestinal digestive enzymes and histomorphometry, and disease resistance in the Nile tilapia. We formulated five isonitrogenous and isocaloric experimental diets to incorporate BSFA at level of (0%, 15%, 30%, 45%, 60%). The growth performance parameters (FBW, BWG, SGR, PER, and FCR) revealed a significant increase in the BSFA30 tilapia group compared to the control group followed by BSFA45 (P < 0.05). The BSFA30 group exhibited the highest nonspecific immunity parameters including (lysozyme activity, phagocytic index, and phagocytic activity) compared to other groups (P < 0.05). SOD and GPx reported the highest values in the BSFA60 group. Nile tilapia carcass composition was not influenced by BSFA inclusion level (P > 0.05). Interestingly, Nile tilapia fed with BSFA15 diet exhibited the highest protease activity level (P < 0.05), while those fed on BSFA30 documented the highest amylase activity. Intestinal histomorphology was significantly enhanced with the gradual increase of administrated BSFA. Regarding the tilapia disease resistance against Aeromonas septicemia, BSFA significantly diminished the cumulative mortality compared to the control group. To sum up, BSFA was more effective in improving the growth performance and immunity of Nile tilapia.

Host plant species will influence the population and physiological performance of insects. Frankliniella occidentalis is a well-known invasive pest commonly found on flowering plants. Herein, the population development of F. occidentalis was investigated on the flowers of different Rosa chinensis cultivars (Ruby, Love, Parade, Pink Peace, and Mohana), and the digestive enzyme activities in thrips were measured after feeding on these flowers.The developmental times of F. occidentalis from egg to adult were 10.07, 10.37, 11.64, 10.66, and 10.90 d on Ruby, Love, Parade, Pink Peace, and Mohana, respectively. Significant differences in fecundity were also observed, with the greatest fecundity levels of F. occidentalis on Ruby (82.96) and the lowest on Mohana (63.40). F. occidentalis showed the greatest R0 on Ruby (43.57), followed by Love (36.46), Parade (33.00), Pink Peace (27.97), and Mohana (23.21). The rm showed a similar trend, with values of 0.156, 0.145, 0.141, 0.134, and 0.130, respectively.There were significant differences in digestive enzyme activities in F. occidentalis on different flowers, and different digestive enzymes showed different performance among these plants.The highest amylase and lipase activities in F. occidentalis were on Ruby, on which F. occidentalis had the fastest development rate and the highest R0, whereas the highest trypsin activity was on Pink Peace. All three digestive enzymes in thrips showed the lowest activities on Mohana.The varied population development of F. occidentalis associated with R. chinensis cultivars may be related to their digestive enzyme performance, which plays important roles in nutrient metabolism and insect growth. Graphical Abstract

The effects of heating duration on digestibility of the muscle powder of three fish (pollock [WP], yellowtail [YT], mackerel [CM]) and the whole-body powder from mackerel (WM) were assessed in vitro using a digestive enzyme complex (EC) extracted from yellowtail pyloric caeca. This research will contribute to establishing a theoretical basis for feed processing conditions for yellowtail. Transcriptomic analysis showed that the most dominantly expressed genes in the pyloric caeca examined were those for trypsin and that their activity was well maintained in the EC. To assess heating effects, the powders were heated at 99 °C for 0, 3, 20, 72 and 240 min, followed by hydrolysis with the EC. After 3 min of heating the WP, YT and CM, the average production of essential amino acids (EAAs) decreased to 81.3%, 72.0% and 66.9%, respectively, compared to that detected in controls without heating. The production further decreased with increased heating duration from 3 to 240 min, but much of the decrease was within the first 3 min in WP, YT and CM. The decreasing rate of EAA production was higher in WM than CM, and increased with heating time. These results suggest that WP is the most suitable feed material for yellowtail of the four samples tested and that the shortening of heating time and removal of viscera are also important factors.

This study aimed to evaluate the effect of adding molasses in different times on water quality, growth performance, body biochemical composition, digestive and hepatic enzymes of Nile tilapia in the biofloc system. Tilapia fingerlings (1.53 ± 0.14 g) were distributed in five treatments including control, BFT24 (adding molasses to the tanks every 24 h), BFT48 (48 h), BFT72 (72 h), and BFT96 (96 h) and reared for 37 days in fiberglass tanks (130 L), with a stocking density of one fish per litre. The results showed that highest increases in biomass (740.12 g) and survival (98.97%) were obtained in BFT24 treatment (P<0.05). The body composition was affected by the experimental treatments so that the highest protein content was obtained in the BFT72 (P<0.05). Digestive enzymes activities were significantly (P<0.05) higher in BFT treatments than the control group. The current study showed higher biomass and survival ratio for Nile tilapia were observed in BFT24 treatment. The liver and digestive enzymes of Nile tilapia were affected by the different addition times of molasses to the rearing tanks.