Hydrogen peroxide (H2O2) acts as a signaling molecule in response to cold stress. Mitogen-activated protein kinases (MAPKs) and C-repeat/dehydration-responsive factor (CBF) play important roles in cold response regulation. To investigate the roles of MAPKs and CBF in H2O2-induced chilling tolerance, tomato (Solanum lycopersicum cv. Ailsa Craig) plants were treated with 1 mM H2O2 before chilling treatment. The results showed that H2O2 treatment protected subcellular structure, increased concentrations of abscisic acid (ABA), zeatin riboside (ZR), and methyl jasmonate (MeJA), but decreased the concentration of gibberellic acid (GA3). Furthermore, 1 mM H2O2 treatment enhanced the activities of antioxidant enzymes. Meanwhile, relative expressions of SlMAPK1/2/3 and SlCBF1 in H2O2-treated plants were higher than those in the control. Our findings suggest that H2O2 treatment might enhance the chilling tolerance of tomato plants by activating SlMAPK1/2/3 and SlCBF1 gene expression and by regulating phytohormone concentrations and antioxidant enzyme activities.Keywords: antioxidant enzymes; chilling tolerance; hydrogen peroxide; SlCBF1; SlMAPK1/2/3; tomato plants;

Study of endophytic bacteria within plant seeds is very essential and meaningful on account of their heritability and versatility. This study investigated Bacillus bacterial communities within the seeds of four commercial tomato varieties, by 16S rRNA gene PCR-RFLP (restriction fragment length polymorphism). Phylogenetic analysis of 16S rRNA gene sequences indicated that the 22 representative isolates belonged to five species of genus Bacillus and the bacterial compositions showed remarkable differences among tomato varieties. Isolates exhibited multiple plant growth promoting (PGP) traits: 37 % of indole-3-acetic acid production; 37 % of phosphate solubilization; 24 % of siderophores production; 85 % of potential nitrogen fixation and 6 % of 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Isolate HYT-12-1 was shown to have highest ACC deaminase activity (112.02 nmol α-ketobutyrate mg−1 protein h−1) among the five ACC deamiase producing strains. 16S rRNA gene sequencing indicated that the isolate HYT-12-1 shared the highest sequence similarity (100 %) with B. subtilis. PGP experiments under gnotobiotic and greenhouse conditions revealed the ability of strain HYT-12-1 to enhance the growth of tomato seedlings. This is the first study to describe endophytic Bacillus communities within tomato seeds, and the results suggest that B. subtilis strain HYT-12-1 would have a great potential for industrial application as biofertilizer in the future.

A novel gene fragment containing a xylanase was identified from a Holstein cattle rumen metagenomic library. The novel xylanase (Xyln-SH1) belonged to the glycoside hydrolase family 10 (GH10) and exhibited a maximum of 44% identity to the glycoside hydrolase from Clostridium thermocellum ATCC 27405. Xyln-SH1 was heterologously expressed, purified, and characterized. A high level of activity was obtained under the optimum conditions of pH 6.5 and 40 °C. A substrate utilization study indicated that Xyln-SH1 was cellulase-free and strictly specific to xylan from softwood. The synergistic effects of Xyln-SH1 and feruloyl esterase (FAE-SH1) were observed for the release of xylooligosaccharides (XOS) and ferulic acid (FA) from wheat straw. In addition, a high dose of Xyln-SH1 alone was observed to improve the release of FA from wheat straw. These features suggest that this enzyme has substantial potential to improve biomass degradation and industrial applications.

The influence of methyl jasmonate (MeJA) on postharvest quality and enzyme activities, gene expression level, and the functional component content linked to postharvest deterioration in Agaricus bisporus (J.E. Lange) Imbach fruit bodies was investigated. Freshly harvested fruit bodies were treated with 0 (control), 10 and 100 μM MeJA vapor at 20 °C for 12 h and then stored at 10 °C for up to 7 days. The results indicated that treatments with 100 μM MeJA vapor maintained a high level of soluble protein and total sugar, delayed browning, promoted the accumulation of phenolics and flavonoids, and inhibited the increase of respiratory rate and membrane leakage. Furthermore, 100 μM MeJA inhibited the activities of polyphenoloxidase, increased the antioxidant enzymes activities of catalase and superoxide dismutase, and lowered relative expression levels of three genes encoding polyphenol oxidase (AbPPO1, AbPPO2, and AbPPO3) throughout the storage period. Comparatively, 10 μM MeJA also had a clear beneficial effect on postharvest mushroom quality maintenance but was not as effective as 100 μM MeJA treatment. These findings suggest that application of MeJA could have potential in maintaining the quality of harvested A. bisporus fruit bodies.

A metagenomic library of China Holstein cow rumen microbes was constructed and screened for novel gene cluster. A novel feruloyl esterase (FAE) gene was identified with a length of 789 bp and encoded a protein displaying 56% identity to known esterase sequences. The gene was functionally expressed in Escherichia coli BL21 (DE3), and the total molecular weight of the recombined protein was 32.4 kDa. The purified enzyme showed a broad specificity against the four methyl esters of hydroxycinnamic acids and high activity (259.5 U/mg) to methyl ferulate at optimum conditions (pH 8.0, 40 °C). High thermal and pH stability were also observed. Moreover, the enzyme showed broad resistance to proteases. FAE-SH1 can enhance the release of ferulic acid from wheat straw with cellulase, β-1,4-endoxylanase, β-1,3-glucanase, and pectase. These features suggest FAE-SH1 as a good candidate to enhance biomass degradation and improve the health effects of food and forage.

A method for isolating high purity and quantity RNA from Agaricus bisporus which is rich in proteins, carbohydrate, fiber and secondary metabolites, is described. RNA was extracted from mycelium, primordia, sporophores at two development stages and two post-harvest storage stages as well as from pileipellis, inner cap, gill and stipe of the mature sporophore. The A260/A230 and A260/A280 ratios of isolated RNA from fruiting bodies were both ~2 and the yield was about 200 μg/g fresh wt (FW). The yield of RNA from mycelium was approx. 100 μg/g FW. High quality RNA was also extracted from fruiting body tissues of Lentinus edodes, Pleurotus ostreatus, Flammulina velutipes and Pleurotus eryngii with yields from 130 to 225 μg/g FW. RNA extracted from all samples was intact, as demonstrated by gel electrophoresis and was suitable for downstream molecular applications, including RT-PCR and qPCR.

l-Arginine is the precursor of nitric oxide (NO). In order to examine the influence of l-arginine on tomato fruit resistance, preharvest green mature tomato fruits (Solanum lycopersicum cv. No. 4 Zhongshu) were treated with 0.5, 1, and 5 mM l-arginine. The reduced lesion size (in diameter) on fruit caused by Botrytis cinerea, as well as activities of phenylalanine ammonia-lyase (PAL), Chitinase (CHI), β-1,3-glucanase (GLU), and polyphenoloxidase (PPO), was compared between l-arginine treated fruits and untreated fruits. We found that induced resistance increased and reached the highest level at 3–6 days after treatment. Endogenous NO concentrations were positively correlated with PAL, PPO, CHI, and GLU activities after treatment with Pearson coefficients of 0.71, 0.94, 0.97, and 0.87, respectively. These results indicate that arginine induces disease resistance via its effects on NO biosynthesis and defensive enzyme activity.

The effects of methyl salicylate (MeSA) on chilling injury (CI) and gene expression levels, enzyme activities, and metabolites related to arginine catabolism in cherry tomato fruit were investigated. Freshly harvested fruits were treated with 0.05 mM MeSA vapor at 20 °C for 12 h and then stored at 2 °C for up to 28 days. MeSA reduced CI and enhanced the accumulation of putrescine, spermidine, and spermine, which was associated with increased gene expression levels and activities of arginase, arginine decarboxylase, and ornithine decarboxylase at most sampling times. MeSA also increased nitric oxide synthase activity, which at least partly contributed to the increased nitric oxide content. The results indicate that MeSA activates the different pathways of arginine catabolism in cold-stored fruit and that the reduction in CI by MeSA may be due to the coordinated metabolism of arginine and the increase in polyamines and nitric oxide levels.

Metallothioneins (MTs), as a family of low-molecular-weight, cysteine-rich, and metal-binding proteins, show potential for utilization in functional food. Tomato plants were transformed with gene constructs that contained mt-I encoding the mouse MT-I, similar in sense orientation with the constitutively active double 35S promoter from cauliflower mosaic virus. Three independent transformants, which had copies of the gene in their genomes, were obtained. In these transgenic lines, high-level expression of MT-I, high zinc content, and some antioxidant enzyme activities were detected in leaves. The average zinc content in transgenic tomato leaves was 32.7 mg/100 g FW, which about 1.6 times higher than that in wild-type. The superoxide dismutase activity was also higher (68.6, 66.9, and 66.1 U/g FW in the three transformants) than that in wild-type (57.4 U/g FW). In particular, the levels of superoxide free radical scanvenging in the three transformants were 14.2%, 14.6%, and 13.7%, respectively, which about 1.5 times higher than that in control (5.6%). Transgenic MT tomato may potentially be used as an antioxidant and for zinc supplementation.

•Preharvest sodium nitroprusside (SNP treatment increased the NO content in ‘Golden Delicious’ apples.•Preharvest SNP treatment inhibited ethylene metabolism.•Preharvest SNP treatment promoted the accumulation of sucrose.•Preharvest SNP treatment might be a potential measure for ripening control.In order to examine the influence of preharvest nitric oxide (NO) treatment on ethylene biosynthesis and soluble sugar metabolism in ‘Golden Delicious’ apples, apple trees were sprayed with 50 μM sodium nitroprusside (SNP) (a donor of NO) 14 days before harvest. The results indicated that preharvest SNP treatment can increase the NO content and the NOS activity in apple fruit, therefore, delay the accumulation of ethylene due to its inhibition on the activities of 1-aminocyclopropane-1-carboxylate synthase (ACS) and 1-aminocyclopropane-1-carboxylate oxydase (ACO). Fructose is the main sugar in ‘Golden Delicious’ apple. The synthesis of sucrose was stimulated and the decomposition of sucrose was inhibited by this treatment, thus causing the accumulation of sucrose. We can draw a conclusion that pre-harvest SNP (50 μM) treatment can increase the NO content of fruit during storage, while higher NO content can further regulate fruit ripening through its effect on ethylene and sugar metabolism in ‘Golden Delicious’ apple fruit during storage at 18 °C.

Treatment of tomato (Solanum lycopersicum L. cv. Messina) fruit with hot air (HA) at 38 °C enhanced the transcript levels of LeARG1 and LeARG2, the two genes encoding arginase, and arginase activity. The strongest induction of LeARG1 and LeARG2 transcripts was observed after fruit treated with 38 °C HA for 12 h, which also effectively alleviated chilling injury (CI) of tomato fruit, manifested as decreased CI index, electrolyte leakage and malondialdehyde content during cold storage. To investigate the potential role of arginase in HA-induced chilling tolerance, fruit were treated with HA, or arginase inhibitor Nω-hydroxy-nor-l-arginine (nor-NOHA) combined with HA and then stored at 2 °C for up to 28 d. The results showed that HA-induced arginase activity was strongly inhibited by pretreatment with nor-NOHA and the reduction of CI by HA was nearly abolished by the arginase inhibitor. In addition, HA treatment increased activities of superoxide dismutase, catalase and ascorbate peroxidase, inhibited peroxidase activities, and promoted the accumulation of arginine, proline and putrescine. These effects were partially counteracted by nor-NOHA except that arginine and putrescine accumulation was unaffected. Our results indicate that arginase induction may be partly involved in HA-induced chilling tolerance in tomato fruit, possibly by a mechanism involving activation of antioxidant enzymes and an increase in proline levels.Highlights► Hot air (HA) treatment induced gene expression and activity of arginase in tomato fruit. ► HA-induced chilling tolerance was almost abolished by an arginase inhibitor nor-NOHA. ► The changes in activities of antioxidant enzymes in HA-treated fruit was diminished by nor-NOHA. ► Inhibition of arginase activity by nor-NOHA reduced HA-induced proline accumulation. ► Arginase induction is involved in HA-induced chilling tolerance in tomato fruit.

Freshly harvested tomato fruit were pretreated with 0.2 mM arginine at −35 kPa for 0.5 min and then stored at 2 °C for 28 d to investigate the effect of exogenous arginine treatment on endogenous arginine catabolism in relation to chilling injury (CI). Arginine treatment reduced the CI index of fruit and enhanced accumulation of polyamines, especially putrescine, and proline, which resulted from the increased activities of the catabolic enzymes arginase, arginine decarboxylase, ornithine decarboxylase and ornithine δ-aminotransferase at most sampling times. Nitric oxide synthase activity was also increased by arginine treatment, which at least partly contributed to the increased nitric oxide concentration. These results revealed that the reduction in CI by exogenous arginine may be due to the accumulation of putrescine, proline and nitric oxide induced by activating the different pathways of endogenous arginine catabolism.Highlights► Arginine treatment alleviated postharvest chilling injury in tomato fruit. ► Exogenous arginine regulated enzymes related to endogenous arginine catabolism. ► The levels of putrescine, proline and NO in fruit were enhanced by arginine treatment. ► Arginine alleviated chilling in fruit due in part to activated arginine catabolism.

Methyl jasmonate (MeJA), a major derivative of the plant hormone jasmonic acid, plays a critical role in inducing resistance to fungal pathogen. To study the endurance of MeJA-induced resistance and its cause, green mature tomatoes (Solanum esculentum cv. Lichun) were treated with 100 μM MeJA and nordihydroguaiaretic acid (NDGA, LOX inhibitor) at −35 kPa for 0.5 min and incubated at 25 ± 1 °C, 85–90% RH. Treatment with MeJA reduced disease symptoms in tomato fruit soon after being inoculated with Botrytis cinerea. Lesion size in MeJA-treated fruit was inhibited by 42.5%, 27.9% and 13.9% respectively (P < 0.05) in fruit inoculated 1, 3 and 6 d after treatments. At advanced stages (inoculation carried out 9 and 12 d after treatments), no inhibitory effect of MeJA were found. Ethylene biosynthesis was activated in the response of green mature tomatoes to methyl jasmonate with a rapid (1 d) and enhanced ethylene peak (0.9 ng kg−1 FW s−1). However the ethylene level was below that of the control from 6 d to 12 d. This rise was closely related with conversion of ACC to ethylene, especially a rise in ACO activity (6 h), which preceded an increase in ACS (12 h) after MeJA treatment. The development of ethylene biosynthesis was accompanied by a significant increase in LOX activity. Two significant O2− peaks (P < 0.05) were detected in MeJA-treated fruit during storage (6.18 μmol g−1 FW min−1 at 6 h and 5.68 μmol g−1 FW min−1 at 3 d). The correlations between LOX, and O2− and ACO activities were 0.75, 0.73 respectively (P < 0.05). These results indicate that MeJA-induced resistance against B. cinerea is durable, MeJA induces LOX and the superoxide radicals formed by LOX may activate ACO and ethylene biosynthesis.

Nitric oxide (NO), an important signaling molecule in plants, is known to play crucial roles in the regulation of both defense responses and inducing resistance to fungal pathogens. To investigate NO synthesis during induced resistance and its mechanism, green mature tomatoes (Lycopersicon esculentum cv. No.4 Zhongshu) were treated with the elicitor from Botrytis cinerea, considered an effective inducer of disease resistance, nitric oxide synthase (NOS) inhibitor N-nitro-l-arginine (l-NNA) and nitric reductase (NR) inhibitor sodium azide (NaN3). Our data indicated that reduced disease symptoms, and lesion size, in tomato fruit with B. cinerea were inhibited by 93.3 and 80.9% respectively three and four days after inoculation. NO biosynthesis was rapidly initiated one day after green mature tomatoes were treated with the elicitor and the peak NO content reached 3.43 μmol/g FW. The activities of NOS and defense enzymes, including phenylalanine ammonia-lyase (PAL), chitinase (CHI), β-1,3-glucanase (GLU), and polyphenoloxidase (PPO), were also increased by the elicitor treatment. Gene expression of pathogenesis-related protein 1(PR-1) increased within 2 h, and total phenols contents increased significantly within four days. A treatment of NOS inhibitor + elicitor increased disease incidence and lesion area and reduced the activities of defense enzymes compared with elicitor-treated fruit, indicating that elicitor-induced NO generation and disease resistance could be blocked by a NOS inhibitor. The NR inhibitor did not obviously influence the content of NO and disease resistance after incubation with the elicitor, and elicitor treatment did not induce NR activity. These results imply that NOS is a key enzyme regulating NO production in induced resistance.Research highlights▶ The elicitor from Botrytis cinerea was considered as an effective inducer of disease resistance. ▶ Increased disease resistance and NO biosynthesis in tomato fruit with the elicitor treatment were found. ▶ The elicitor-induced NO generation and disease resistance could be blocked by a NOS inhibitor, which implies that NOS is a key enzyme regulating NO production in induced resistance.

•We tested PGPB on tomato seedlings growth under low night temperature.•Plants were adversely affected by night chilling stress.•PGPB enhanced plants growth and nutrient uptake.•Antioxidant system was induced by PGPB under night chilling stress.This study investigated the effects of inoculation with selected 1-aminocyclopropane-1-carboxylate (ACC) deaminase-producing endophytic bacterium TPs-04 on growth promoting, nutrient uptake and antioxidant system of tomato (Solanum lycopersicum L.) seedlings under low night temperature (LNT, i.e., 6, 9 and 15 °C). Growth and nutrient uptake of tomato seedlings was reduced under different LNT stress level. Inoculation with strain TPs-04 significantly increased the length and dry weight of root and shoot, and chlorophyll content of leaves. Further analysis showed that it could enhance the uptake of N, P and K. The improvement of seedling growth might due to the plant growth promoting traits such as siderophore production, phosphorous solubilization, nitrogen fixation and ACC deaminase production indicated by TPs-04. Chilling stress resulted in rapid accumulation of hydrogen peroxide (H2O2) and malondialdehyde (MDA). Moreover, severe LNT (6 °C) raised the antioxidant enzyme activities compared to 9 and 15 °C treatments. The selected strain TPs-04 induced a higher increase in these antioxidant enzyme activities, and substantially decreased the accumulation of H2O2 and MDA in response to night chilling stress. Strain TPs-04 might be used as a potential bioinoculant of tomato plants protection under night chilling stress.