Selenocysteine methyltransferase SMT catalyzed the synthesis of Se-methylselenocysteine to regulate the accumulation of glucosinolates and sulforaphane in broccoli

Selenocysteine methyltransferase(SMT)is a key enzyme involved in the Se metabolism pathway,and it is responsible for the catalysis of Se-methylselenocysteine(SeMSC)compound formation.Previous studies showed that selenium treatment activated SMT expression and promoted the accumulation of glucosinolates(GSLs)and sulforaphane,but the roles and functional mechanisms of SMT in mediating GSLs and sulforaphane synthesis remain unclear.In this study,we identified the BoSMT gene in broccoli and uncovered its roles in mediating GSLs biosynthesis.Transgenic assays revealed that BoSMT is involved in SeMSC biosynthesis in broccoli.More importantly,the contents of GSLs and sulforaphane were significantly increased in the BoSMT-overexpressing broccoli lines but decreased in the knockdown lines,suggesting that BoSMT played a positive role in regulating GSLs and sulforaphane synthesis.Further evidence indicated that BoSMT-mediated overaccumulation of GSLs and sulforaphane might be due to the increase in the endogenous SeMSC content.Compared with the mock(water)treatment,selenite-induced significantly increases of the SeMSC content in the BoSMT-knockdown plants partially compensated the phenotype of GSLs and sulforaphane loss.Compared with the mock treatment,exogenous SeMSC treatment significantly increased the contents of GSL and sulforaphane and activated GSL synthesis-related gene expression,suggesting that SeMSC acted as a positive regulator for GSL and sulforaphane production.Our findings provided novel insights into selenium-mediated GSLs and sulforaphane accumulation.The genetic manipulation of BoSMT might be a useful strategy for improving the dietary nutritional values of broccoli.

UV-B irradiation enhances the accumulation of beneficial glucosinolates induced by melatonin in Chinese kale sprout

Cruciferous sprout is a new form of vegetable product rich in bioactive compounds,especially glucosinolates.Previous studies have focused on increasing the accumulation of glucosinolates in cruciferous sprouts by applying different chemical regulators,with a particular focus on their contribution to nutritional quality and health benefits.Nevertheless,the effects of melatonin and UV-B irradiation on glucosinolate biosynthesis remain unclear.In this study,it was found that changes in melatonin concentrations significantly affected the contents of individual as well as total aliphatic and indolic glucosinolates.The 5μmol·L^(-1)melatonin was decided as the optimum concentration that could increase the content of beneficial glucosinolates including glucoraphanin and 4-methoxy glucobrassicin in Chinese kale sprouts.Notably,the enhancement of glucosinolate accumulation by melatonin treatment could be further amplified by UV-B irradiation.Furthermore,our results showed that R2R3-MYB transcription factor BoaMYB28 and BoaMYB51,which are central regulators of aliphatic and indolic glucosinolate biosynthesis respectively,were both involved in the regulation of glucosinolate biosynthesis by melatonin and UV-B irradiation.Additionally,the expression of glucosinolate biosynthetic genes,including BoaCYP79F1,BoaCYP83A1,BoaSUR1,BoaUGT74B1,BoaCYP79B2,BoaCYP79B3,and BoaCYP83B1 participated in the formation of core structures and BoaFMOGS-OX5,BoaAOP2,BoaCYP81F2,and BoaIGMT1 involved in the sidechain modification of aliphatic and indolic glucosinolate,was regulated by melatonin or UV-B irradiation.Taken together,these findings provide a potential strategy for improving the nutritional quality and resistance of Chinese kale sprouts.

The effect of glutathione on glucosinolate biosynthesis through the sulfur assimilation pathway in pakchoi associated with the growth conditions

Glucosinolates(GSLs) are a group of nitrogen-and sulfur-containing secondary metabolites, synthesized primarily in members of the Brassicaceae family, that play an important role in food flavor, plant antimicrobial activity, resistance to insect attack, stress tolerance, and human anti-cancer effects. As a sulfur-containing compound, glutathione has a strong connection with GSLs biosynthesis as a sulfur donor or redox system, and exists in reduced(glutathione;GSH) and oxidized(glutathione disulfide;GSSG) forms. However, the mechanism of GSH regulating GSLs biosynthesis remainds unclear. Hence, the exogenous therapy to pakchoi under normal growth condition and sulfur deficiency condition were conducted in this work to explore the relevant mechanism. The results showed that exogenous application of buthionine sulfoximine, an inhibitor of GSH synthesis, decreased the transcript levels of GSLs synthesis-related genes and transcription factors, as well as sulfur assimilation-related genes under the normal growth condition. Application of exogenous GSH inhibited the expression of GSLs synthesis-and sulfur assimilation-related genes under the normal condition, while the GSLs biosynthesis and the sulfur assimilation pathway were activated by exogenous application of GSH when the content of GSH in vivo of plants decreased owing to sulfur deficiency. Moreover,exogenous application of GSSG increased the transcript levels of GSLs synthesis-and sulfur assimilation-related genes under the normal growth condition and under sulfur deficiency. The present work provides new insights into the molecular mechanisms of GSLs biosynthesis underlying glutathione regulation.

Transcriptomics integrated with metabolomics reveals the mechanism of CaCl_(2)-HCl electrolyzed water-induced glucosinolate biosynthesis in broccoli sprouts

Glucosinolates are important phytochemicals in Brassicaceae.We investigated the effect of CaCl_(2)-HCl electrolyzed water(CHEW)on glucosinolates biosynthesis in broccoli sprouts.The results showed that CHEW treatment significantly decreased reactive oxygen species(ROS)and malondialdeh yde(MDA)contents in broccoli sprouts.On the the 8^(th)day,compared to tap water treatment,the the total glucosinolate content of broccoli sprouts with CHEW treatment increased by 10.6%and calcium content was dramatically enhanced from 14.4 mg/g DW to 22.7 mg/g DW.Comparative transcriptome and metabolome analyses revealed that CHEW treatment activated ROS and calcium signaling transduction pathways in broccoli sprouts and they interacted through MAPK cascades.Besides,CHEW treatment not only promoted the biosynthesis of amino acids,but also enhanced the expression of structural genes in glucosinolate synthesis through transcription factors(MYBs,bHLHs,WRKYs,etc.).The results of this study provided new insights into the regulatory network of glucosinolates biosynthesis in broccoli sprouts under CHEW treatment.

Screening of glucosinolates degrading lactic acid bacteria and their utilization in rapeseed meal fermentation

Rapeseed meal is a promising food ingredient, but its utilization is limited by the presence of some potentially harmful ingredients, such as glucosinolates. Fermentation is a cost-effective method of detoxication but a food-grade starter culture with glucosinolates degradation capacity is required. In this study, 46 strains of lactic acid bacteria from traditional paocai brines were screened for their ability to glucosinolate degradation. The results showed that more than 50% of the strains significantly degraded glucosinolates. Two strains of Lactiplantibacillus(p7 and s7) with high capacity of glucosinolates degradation through producing enzymes were identified. Then,an optimized condition for rapeseed meal fermentation by p7 was established to degrade glucosinolates, which can achieve about 80% degradation. UPLC/Q-TOF-MS analysis showed that the degradation rate of individual glucosinolates was different and the degradation rate of gluconapin and progoitrin in rapeseed meal can reach more than 90%. Meanwhile, fermentation with p7 can improve safety of rapeseed meal by inhibiting the growth of Enterobacteriaceae and improve its nutritional properties by degrading phytic acid. The in vitro digestion experiments showed that the content of glucosinolates in rapeseed meal decreased significantly during gastric digestion. Meanwhile, fermentation with p7 can greatly improve the release of soluble protein and increase the contents of free essential amino acids, such as lysine(increased by 12 folds) and methionine(increased by 10 folds).

Effects of fresh-cut and storage on glucosinolates profile using broccoli as a case study

Glucosinolates(GLS) contribute to the unique flavour, nutrition, and plant defence of the Cruciferous vegetables. Understanding the GLS changes through postharvest processing is essential for defined preservation. In this study, four different fresh-cut types, whole flower(W),floret(F), quarterly cut floret(QF) and shredded floret(FS) of broccoli, were stored for 0, 1, 2 and 3 day(s) to explore GLS responses to postharvest treatments. As a result, seven GLS were identified, mainly including glucoraphanin(RAA), neoglucobrassicin(NEO), and glucobrassicin(GBC)and accounting for 52.69%, 20.12% and 14.99% of the total GLS(21.92 ± 0.48) μmol · g ^(-1 )DW, respectively. FS had the sharpest decrease in GLS after three days of storage(6.55 ± 0.37) μmol · g-1DW, while QF had the least(10.16 ± 0.33) μmol · g ^(-1 )DW. All GLS components decreased over storage, except for 4-methoxyglucobrassicin(4 ME) in FS and QF, suggesting its key role in serious wound defence. The results suggested certain postharvest approaches influenced the flavour and nutrition of broccoli.

Diversity of glucosinolates among common Brassicaceae vegetables in China

Brassicaceae vegetables are an important traditional daily food in China and around the world, which provide nutrients and phytochemicals that are beneficial for human health. Among them, Brassica and Raphanus are widely cultivated and eaten, have been evolved and/or bred for special characteristics during the long history of cultivation. Epidemiological studies suggest that the health benefits of Brassicaceae vegetables are mainly associated with glucosinolates(GSLs) and their hydrolytic products. In this review, we discuss the diversity of common consumed Brassicaceae vegetables and their GSL composition in edible parts. We also discuss the diversity factors affecting GSL content, and the diversity roles and functions of GSL. The information in this review provides guidance for consumers to select vegetables with a high GSL content,optimum edible stages, suitable edible methods, and provides a theoretical basis for crop molecular breeding and market development of GSL products.

Dissecting the genetic architecture of glucosinolate compounds for quality improvement in flowering stalk tissues of Brassica napus

Glucosinolates(GSLs) and their hydrolytic products contribute to the quality traits of rapeseed flowering stalk tissues, such as taste, flavor and anticarcinogenic properties(Glucoraphanin). However, little is known about the genetic mechanisms of GSL accumulation in rapeseed flowering stalks. In this study, the variation and genetic architecture of GSL metabolites in flowering stalk tissues were investigated for the first time among a panel of 107 accessions. All GSL compounds exhibited continuous and wide variations in the present population. Progoitrin,glucobrassicanapin and gluconapin were the most abundant GSL compounds. Five quantitative trait loci(QTL) significantly associated with three GSL compounds were identified by genome-wide association study. GRA_C04 was under selected during modern breeding, in which the ratio of lower GSL haplotype(HAP2) in the accessions bred before 1990(52.56%) was significantly lower than that after 1990(78.95%). Four candidate genes, BnaA01. SOT16, BnaA06. SOT17, Bna A06. MYB51a, and Bna A06. MYB51b, were identified in the GTL_A01 and 4OH_A06 regions.These findings provide new insights into GSL biosynthesis in flowering stalk tissues and facilitate quality improvement in rapeseed flowering stalks.

Glucosinolates and Their Hydrolysis Products in Arabidopsis thaliana Influence Performance and Feeding Choice of Pieris rapae and Spodoptera exigua

Glucosinolates and their hydrolysis products, found in plants of the order Brassicales, are well-known for their defensive properties against insect herbivores. Arabidopsis thaliana (Col-0) genetic lines with mutations that modify the type of glucosinolates (i.e. myb28myb29 and cyp79B2cyp79B3 are deficient in the production of aliphatic and indolyl glucosinolates, respectively) make it possible to test for the specific effects of these secondary chemicals on insect herbivores. The Pad3 mutant (deficient in camalexin), which has a role in resistance to pathogens, was also tested. Likewise, the effects of different glucosinolate hydrolysis products can be evaluated using genetically modified (GM) lines of the wild type Col-0 ecotype, which naturally produces isothiocyanates. These GM lines include the nitrile-producing 35S: ESP and the double knockout tgg1tgg2, which virtually lacks hydrolysis products. In both no-choice and choice experiments, the crucifer specialist Pieris rapae was virtually unaffected by differences in the type of glucosinolates or hydrolysis products. In contrast, the generalist insect Spodoptera exigua had statistically significant increases in pupae/adult weight and faster developmental times when reared on mutants deficient in the production of aliphatic and indolyl glucosinolates and their hydrolysis products. There were no differences in the performance of either insect species when reared on wild type Col-0 or Pad3. Results from feeding choice trials showed that Pieris rapae had no statistically significant preference for any of the genetic lines. In contrast, Spodoptera exigua had a significant feeding preference for the double mutant tgg1tgg2. This study provides evidence that variation in the type of glucosinolates and their hydrolysis products can influence insect performance and feeding choices, and that responses are species-specific.

Effects of Drought Stress Simulated by PEG on Glucosinolates Content in Arabidopsis thaliana

[Objective] The paper aimed to study effects of drought stress simulated by PEG on glucosinolates content in Arabidopsis thaliana.[Method] Drought stress was simulated by PEG-6000,ecological seeds of Arabidopsis thaliana were cultivated by the control group and drought treatment group respectively,Physical signs of Arabidopsis thaliana and contents of glucosinolates were determined after 0,4,5,6,7 d treatment.[Result] The results showed that leaf water content of rosette leaves was obviously decreased,leaf relative conductivity （characterized by membrane permeability） and the concentration of MDA increased,the extent of damage increased with the increased time.Content of total glucosinolate,aliphatic glucosinolate and indole glucosinolate increased got their maximum after 5 days treatment,and rapidly decreased after 6 and 7 days of treatment,even much lower than the control group.Each kind of glucosinolate changed with difference from each other.4MSOB which made the most proportion of the total glucosinolate changed consistently with the total glucosinolate and difference significant.As a whole,aliphatic glucosinolates were more sensitive to drought than indole glucosinolate.The proportion of some kind glucosinolate,like 4MSOB varied had correlation with the content change.[Conclusion] Drought stress had an effects on the contents of total glucosinolate,aliphatic glucosinolate and indole glucosinolate,which made the glucosinolate participated in defense response of plant to the outside of drought stress,but long-term drought stress in leaves was not conducive to the accumulation of glucosinolates.

Chemical Ionization Mass Spectrometry of Pertrimethylsilylated Desulphoglucosinolates

The chemical ionization mass spectra of ten different glucosinolates isolated from rape seed have been studied. More intense diagnostic ions were identified than in the electron impact mass spectra. The method is suitable for the structure analysis of mixture of glucosinolates.

Solid state fermentation of rapeseed cake with Aspergillus niger for degrading glucosinolates and upgrading nutritional value

Background： Rapeseed cake is a good source of protein for animal feed but its utilization is limited due to the presence of anti-nutritional substances, such as glucosinolates （GIs）, phytic acid, tannins etc. In the present study, a solid state fermentation （SSF） using Aspergillus niger was carried out with the purpose of degrading glucosinolates and improving the nutritional quality of rapeseed cake （RSC）. The effects of medium composition and incubation conditions on the GIs content in fermented rapeseed cake （FRSC） were investigated, and chemical composition and amino acid in vitro digestibility of RSC substrate fermented under optimal conditions were determined. Results： After 72 h of incubation at 34℃, a 76.89% decrease in GIs of RSC was obtained in solid medium containing 70% RSC, 30% wheat bran at optimal moisture content 60% （w/w）. Compared to unfermented RSC, trichloroacetic acid soluble protein （TCA-SP）, crude protein and ether extract contents of the FRSC were increased （P〈 0.05） 103.71, 23.02 and 23.54%, respectively. As expected, the contents of NDF and phytic acid declined （P〈 0.05） by 9.12 and 44.60%, respectively. Total amino acids （TAA） and essential amino acids （EAA） contents as well as AA in vitro digestibility of FRSC were improved significantly （P 〈 0.05）. Moreover, the enzyme activity of endoglucanase, xylanase, acid protease and phytase were increased （P 〈 0.05） during SSF. Conclusions： Our results indicate that the solid state fermentation offers an effective approach to improving the quality of proteins sources such as rapeseed cake.

Content determination of benzyl glucosinolate and anti—cancer activity of its hydrolysis product in Carica papaya L.

Objective:To determine the content of benzyl glueosinolate（BG） in the pulp and the seed and investigate the anti-cuncer activity of its hydrolysis product in Curica papaya L.Methods: Determination of BG was performed on an Hypersil BDS C18 column at the wavelength of 214 nm with 0.1%trifluoroacelic acid（TFA） aqueous solution（A） and 0.1%TFA acelonilrile（B） as the mobile phase.In vitro activity test was adopted with cidtured human lung cancer H69 cell in vitro to investigate the inhibition rate of cell proliferation of benzyl isothiocyanale（BITC） againsl H69 cell.Results:The pulp has more BG before the maturation of papaya and it nearly disappeared after papaya matured,while the seed contains BG at every stage.Activity test demonstrated that the a higher concentration of BITC would have betler inhibition rate of cell proliferation on 1169 cell,and the IC50 was 6.5μmol/L.Conclusions:BG also can be produced in the pulp of papaya and it will be stored in the seed after the fruit has been matured.The hydrolysis product of BG has certain cancer-prevention anti-cancer activities for human.

The Arabidopsis P450 protein CYP82C2 modulates jasmonateinduced root growth inhibition, defense gene expression and indole glucosinolate biosynthesis

Jasmonic acid （JA） is a fatty acid-derived signaling molecule that regulates a broad range of plant defense responses against herbivores and some microbial pathogens. Molecular genetic studies have established that JA also performs a critical role in several aspects of plant development. Here, we describe the characterization of the Arabidopsis mutantjasmonic acid-hypersensitivel-1 （jah1-1）, which is defective in several aspects of JA responses. Although the mutant exhibits increased sensitivity to JA in root growth inhibition, it shows decreased expression of JA-inducible defense genes and reduced resistance to the necrotrophic fungus Botrytis cinerea. Gene cloning studies indicate that these defects are caused by a mutation in the cytochrome P450 protein CYP82C2. We provide evidence showing that the compromised resistance of thejah1-1 mutant to B. cinerea is accompanied by decreased expression of JA-induced defense genes and reduced accumulation of JA-induced indole glucosinolates （IGs）. Conversely, the enhanced resistance to B. cinerea in CYP82C2-overexpressing plants is accompanied by increased expression of JA-induced defense genes and elevated levels of JA-induced IGs. We demonstrate that CYP82C2 affects JA-induced accumulation of the IG biosynthetic precursor tryptophan （Trp）, but not the JA-induced IAA or pathogen-induced camalexin. Together, our results support a hypothesis that CYP82C2 may act in the metabolism of Trp-derived secondary metabolites under conditions in which JA levels are elevated. Thejah1-1 mutant should thus be important in future studies toward understanding the mechanisms underlying the complexity of JA-mediated differential responses, which are important for plants to adapt their growth to the ever-changing environments.

Effect of Nitrogen and Sulfur Supply on Glucosinolates in Brassica campestris ssp.chinensis

Glucosinolates （GSs） are a group of plant secondary metabolites containing abundant nitrogen （N） and sulfur （S） mainly in Brassica and have the beneficial effects on human health including anti-carcinogenic, cholesterol-reducing and other pharmacological effects. The objective of this study was to investigate the effect of different concentrations of N （5, 10, and 20 mmol L-a, denoted by N5, N10 and N20） and S （0,5, 1, and 2 mmol L^-1, denoted by S0.5, S1 and S2） on the yield and GSs in pakchoi （Brassica campestris L. ssp. chinensis var. communis） in hydroponics. Results showed that N10 and N20 significantly enhanced the yield compared with N5, however, N20 had a negative effect relative to N10. Only with N10 and N20 low S supply （S0.5） reduced the yield. The concentrations of aliphatic GSs, aromatic GS and total GSs were enhanced by N5 and indolyl GSs were enhanced by N20. S2 enhanced the concentration of individual GS and total GSs. The concentrations of indolyl GSs were maximized in N20S2 treatment, whereas the highest concentrations of aliphatic GSs, aromatic GS and total GSs were found in N5S2 treatment. Effects of N and S on aliphatic GSs were higher than on indolyl GSs. The results suggest that the accumulation of aliphatic GSs and aromatic GS could be enhanced by low N and high S and restricted by high N while that of indolyl GSs could be enhanced by high N and high S.

Identification of MAM1s in Regulation of 3C Glucosinolates Accumulation in Allopolyploid Brassica juncea

Allopolyploid Brassica juncea is particularly enriched in sinigrin,a kind of 3C aliphatic glucosinolates(GSLs),giving rise to characteristic taste after picking.However,the molecular mechanism underlying 3C aliphatic GSLs biosynthesis in this species remains unknown.In this study,we genome-widely identified GSLs metabolic genes,indicating different evolutionary rate of GSLs metabolic genes between subgenomes of B.juncea.Eight methythioalkylmalate synthase(MAMs)homologs were identified from B.juncea,in which six MAM1s were located in chloroplast and the other two were not detected with any expression.Furthermore,BjMAM1-4,BjMAM1-5,and BjMAM1-6 displayed higher expression levels in leaves than other tissues.Silenced expression analysis revealed that BjMAM1-4 and BjMAM1-6 function in 3C and 4C aliphatic GSLs accumulation.The specificity of the substrate selection for the second cycle reaction is much lower than that of the first cycle,suggesting these genes may preferentially catalyze 3C aliphatic GSLs biosynthesis.Our study provides insights into the molecular mechanism underlying the accumulation of 3C aliphatic GSLs,thereby facilitating the manipulation of aliphatic GSLs content in B.juncea.

Degradation of glucosinolates in rapeseed meal by Lactobacillus delbrueckii and Bacillus subtilis

Lactobacillus delbrueckii and Bacillus subtilis were employed as a new combination of strains to treat rapeseed meal by solid-state fermentation,aiming to efficiently degrade the glucosinolates,which are the main toxin in the meal.Single-factor tests and Response surface methodology(RSM)were used to optimize the fermentation parameters.Under the optimum fermentation parameters of 15%total injection volume of the mixture of Lactobacillus delbrueckii and Bacillus subtilis with a ratio of 2:1,bran content of16%,feed to water ratio of 1:1.5,fermentation temperature of 36°C and fermentation time of 72 h,the content of glucosinolates in rapeseed meal was decreased from 64.558μmol/g to 3.473μmol/g,reaching a high degradation rate(94.62%).The high detoxification rate by a consortium of Lactobacillus delbrueckii and Bacillus subtilis provides a bright application prospect in feed utilization of rapeseed meal.

A sulfotransferase gene BnSOT-like1 has a minor genetic effect on seed glucosinolate content in Brassica napus

The theory and associated selection methods of classical quantitative genetics are based on the multifactorial or polygene hypothesis.Major genes or quantitative trait loci(QTL)in modern quantitative genetics based on a“major gene plus polygenes”genetic system have been paid much attention in genetic studies.However,it remains unclear how the numerous minor genes act,although the polygene theory has sustained genetic improvement in plants and animals for more than a hundred years.In the present study,we identified a novel minor gene,BnSOT-like1(BnaA09g53490D),which is a sulfotransferase(SOT)gene catalyzing the formation of the core glucosinolate(GSL)structure in Brassica napus.This gene has been occasionally found during investigations of plant height-related genes,but has not been identified by QTL mapping because of its small phenotypic effects on GSL content.The overexpression of BnSOT-like1 up-regulated the expression of aliphatic GSL-associated genes,leading to a high seed aliphatic GSL content,and the overexpression of the allelic gene Bnsot-like1 did not increase seed GSL content.These findings suggest that the SOT gene has a marked effect on a quantitative trait from a reverse genetics standpoint,but a minor effect on the quantitative trait in its natural biological state.Because of the redundancy of GSL biosynthetic genes in the allotetraploid species B.napus,mutations of a single functional gene in the pathway will not result in significant phenotypic changes,and that the genes in biosynthetic pathways such as BnSOT-like1 in our study have minor effects and may be called polygenes in contrast to the reported three regulatory genes(BnHAG1s)which strongly affect GSL content in B.napus.The present study has shed light on a minor gene for a quantitative trait.

Inheritance of Erucic Acid,Glucosinolate,and Oleic Acid Contents in Rapeseed (Brassica napus L.)

This study was conducted to verify the inheritance of certain characters of rapeseed including erucic acid, glucosinolate and oleic acid contents by using generation mean analysis. The cross of lines Ⅲ174×Zi20 （F1）, F2, BC1 （F1×P1）, BC2 （F1×P2）, and parents （P1 and P2） were evaluated in the field. Data were measured on individual plants for oleic acid, erucic acid, and glucosinolate contents. Transgressive variations in F2 population were observed for oleic acid content, indicating that dominance and recessive genes distributed in both parents. Scaling test indicated that the effects of genes controlling these characters did not follow the additivedominance model. The data for three characters were analyzed using six parameter models and found that one or more types of epistatic gene effects were important for glucosinolate content. High broad sense heritabilities were obtained for erucic acid, oleic acid, and glucosinolate contents with the values of 98.97%, 93.68%, and 86.17%, respectively. Two major gene pairs were found to control the expression of erucic acid and oleic acid contents, while three major gene pairs were detected to control glucosinolate content.

Impact of the Glucosinolate Sinigrin on Bacterial Communities in Pieris rapae

Dynamics in animal-associated microbiota can be difficult to study due to community complexity. Previous work showed that microbial communities in the midguts of Pieris rapae larvae contain relatively few members. In this study, we used P. rapae to test hypotheses related to how diet impacts gastrointestinal microbiota. More specifically, we investigated how the concentration of sinigrin, a glucosinolate in the natural diet of this insect, alters microbial community structure. Larvae were fed either sterile wheat germ diet alone or amended with 3.0 mg/ml, 6.0 mg/ml, or 9.0 mg/ml of sinigrin. In order to determine shifts in the gut microbial community, 16S rRNA genes from midguts were subjected to pyrosequencing and analyzed. Sinigrin had a significant impact on microbial communities in fourth instar P. rapae larvae, but this was dependent on concentration. The predominant phyla in all treatment groups were Proteobacteria and Firmicutes. Significant difference in beta diversity was typically observed when sinigrin 6 mg/ml and the control treatment groups were compared. The impact of sinigrin on the structure of the midgut microbiota is dependent on concentration, but not in a linear fashion. This may indicate that types and concentrations of glucosinolates have varied impact on midgut microbial community.