外源硒对低温胁迫下铁皮石斛幼苗的缓解效应及其抗氧化生理特征变化

以3个不同品系的铁皮石斛幼苗为材料,分别于叶面喷施不同浓度的亚硒酸钠溶液处理,并结合人工控温生长条件下的低温胁迫试验,探讨外源硒对铁皮石斛幼苗耐冷性和恢复生长能力的影响作用及其与有关抗氧化生理指标间的关系。结果表明:(1)耐冷性表现较强的品系其幼苗在低温胁迫下的叶绿素、MDA及脯氨酸含量变化幅度相对小于耐冷性较弱的品系。(2)适量浓度(0.05～0.10mg.L-1)外源硒处理对低温胁迫下铁皮石斛幼苗叶绿素降解具有缓解效应,可降低MDA含量,增加脯氨酸含量,同时显著提高GSH-AsA循环活性,从而提高铁皮石斛幼苗的耐冷性,但过高浓度的外源硒处理(>0.20mg.L-1)则效果相反。(3)低温胁迫解除后,0.10mg.L-1的外源硒处理能引起铁皮石斛‘ZD-1’幼苗中AsA含量显著下降并促进其GSH合成积累,有效提升铁皮石斛幼苗在恢复生长期的综合抗氧化防御能力。     

Accumulation and protection activity of protease-resistant heat-stable proteins in Haematococcus pluvialis during high light and nitrogen starvation

Under stress conditions of high light andnitrogen starvation the green motile cellsof the unicellular green algaHaematococcus pluvialis are known tocease growing and transform into inert redcysts, in which the secondary carotenoidastaxanthin accumulates. A study wastherefore made on other effects of suchconditions. A number ofprotease-resistant, heat-stable proteinswith apparent molecular masses of 38 kDa,50 kDa, 62 kDa and 63 kDa accumulated. Thisprotein fraction was effective in theprotection of horseradish peroxidase frominactivation, suggesting a role for theseproteins in H. pluvialis subjected toa stress event.     

Enhancement of growth and polysaccharide production in suspension cultures of protocorm-like bodies from Dendrobium huoshanense by the addition of putrescine

Putrescine at 0.6 mM stimulated protocorm-like body growth and polysaccharide synthesis in suspension cultures of Dendrobium huoshanense. The specific growth rate of protocorm-like body increased from 0.047 to 0.056 day⁻¹, and the maximum dry weight and polysaccharide production reached 33.2 and 2.94 g l⁻¹, respectively, while they were 24.6 and 2.12 g l⁻¹, respectively, in the control. The administration of polyamine inhibitor, α-dl-difluoromethylarginine, at 1 mM, decreased protocorm-like body growth and polysaccharide production to 21.4 and 1.76 g l⁻¹, respectively.     

Effects of in vitro tissue culture conditions and acclimatization on the contents of Rubisco, leaf soluble proteins, photosynthetic pigments, and C/N ratio

The levels of two subunits of chloroplast ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), total soluble proteins, carbon and leaf nitrogen content, and photosynthetic pigments in various plants (avocado, oak, olive, and strawberry) grown in vitro and ex vitro were analysed. Compared to ex vitro grown plants, micropropagated avocado, oak, and strawberry showed a markable decrease in large subunit Rubisco. However, the small subunit only decreased in strawberry and oak. Contrary to this, olive did not reveal any difference in the level of either subunit. The C/N ratio increased significantly in in vitro grown plants, except in the case of olive, where an opposite behaviour was found. Leaf chlorophyll concentration on unit mass basis was higher in all the in vitro plants than in those of greenhouse- grown plants. Only avocado plantlets showed a statistically significant decrease in total soluble proteins. Further, overall data suggest that in vitro cultural conditions have a species-specific influence on large and small subunits of Rubisco, independent of the protein, chlorophyll, or nitrogen level.     

Purification and characterization of carbonyl reductase from Geotrichum candidum

Geotrichum candidum is well known for the reduction of prochiral ketones to chiral alcohol with high yield and excellent enantioselectivity. Carbonyl reductase from G. candidum was purified by ammonium sulphate precipitation, anion exchange and hydrophobic interaction chromatographies. Gel filtration chromatography together with SDS-PAGE revealed this protein to be a dimer of 60 kDa subunits. Maximum enzyme activity was found in acetate buffer at pH 5.4 with t_1/2 of 7.13 h at 30 °C and t_1/2 of 2.8 h at 65 °C. The enzyme was inhibited by p-hydroxymercuribenzoate and hydroxylamine indicating the involvement of thiol and carbonyl groups in the reduction reaction catalyzed by the enzyme. Chelating agents also reduced the enzyme activity indicating the requirement of metal ions as cofactors. The purified carbonyl reductase was found to be highly selective for ketones containing naphthyl ring, whereas aryl or hetero-aryl ketones showed very less or no activity at all.     

Stress-induced accumulation and tissue-specific localization of dehydrins in Arabidopsis thaliana

Stress-induced accumulation of five (COR47, LTI29, ERD14, LTI30 and RAB18) and tissue localization of four (LTI29, ERD14, LTI30 and RAB18) dehydrins in Arabidopsis were characterized immunologically with protein-specific antibodies. The five dehydrins exhibited clear differences in their accumulation patterns in response to low temperature, ABA and salinity. ERD14 accumulated in unstressed plants, although the protein level was up-regulated by ABA, salinity and low temperature. LTI29 mainly accumulated in response to low temperature, but was also found in ABA- and salt-treated plants. LTI30 and COR47 accumulated primarily in response to low temperature, whereas RAB18 was only found in ABA-treated plants and was the only dehydrin in this study that accumulated in dry seeds.Immunohistochemical localization of LTI29, ERD14 and RAB18 demonstrated tissue and cell type specificity in unstressed plants. ERD14 was present in the vascular tissue and bordering parenchymal cells, LTI29 and ERD14 accumulated in the root tip, and RAB18 was localized to stomatal guard cells. LTI30 was not detected in unstressed plants. The localization of LTI29, ERD14 and RAB18 in stress-treated plants was not restricted to certain tissues or cell types. Instead these proteins accumulated in most cells, although cells within and surrounding the vascular tissue showed more intense staining. LTI30 accumulated primarily in vascular tissue and anthers of cold-treated plants.This study supports a physiological function for dehydrins in certain plant cells during optimal growth conditions and in most cell types during ABA or cold treatment. The differences in stress specificity and spatial distribution of dehydrins in Arabidopsis suggest a functional specialization for the members of this protein family.     

Accumulation of seed storage proteins and the taxonomy ofPoaceae

The accumulation of specific seed proteins is a taxonomically valuable feature and can be used to additionally characterize plant taxa. To date, mainly crop proteins have been analysed in thePoaceae. In this investigation seed proteins from 147 species were screened with emphasis on legumin-like proteins and prolamins. The groups resulting from evaluation of the protein profiles correspond with well-known subfamilies and tribes.Panicoideae are clearly separated fromPooideae. WithinPooideae, theBromeae plusTriticeae tribes revealed obvious similarities.Lolium, Festuca andVulpia, generally included in the tribeFestuceae, revealed a protein profile similar to the profile of theBromeae/Triticeae. Legumin-like proteins are accumulated abundantly inBambusoideae andPooideae exceptBromeae/Triticeae, however, only the species included in theAveninae subtribe produce soluble (globulin-type) legumins as already known fromAvena sativa. Dedicated to emer. Univ.-Prof. DrFriedrich Ehrendorfer on the occasion of his 70th birthday     

In vitro Propagation and Isozyme Polymorphism of the Medicinal Plant Hypericum brasiliense

Responses of 20 d-old plants of two Brassica napus L. cultivars Dunkeld and Cyclon to NaCl salinized soil [electrical conductivity 2.4 (control), 4.0, 8.0 or 12.0 dS m⁻¹] were examined. The salt tolerant line Dunkeld had significantly higher fresh and dry masses of shoots, and seed yield than salt sensitive line Cyclon in all salinities. The effect of salt stress on reduction in total leaf soluble sugars was markedly greater in Dunkeld as compared to that in Cyclon. No effect of salt stress was observed on leaf soluble proteins but there was a slight increase in total free amino acids of both cultivars. Leaf proline content increased markedly in both cultivars and Dunkeld had greater proline content than Cyclon at all salinities. Salt stress had no significant effect on seed oil content and erucic acid content of seed oil, however, content of glucosinolates in the seed meal increased and Cyclon had greater content of glucosinolates than Dunkeld. This revised version was published online in July 2006 with corrections to the Cover Date.     

赤杨对辽东落叶松人工林土壤氨基糖积累的影响

微生物残留物是土壤稳定性碳库的重要组成部分,然而其对固氮树种的响应还不清楚。以辽东山区日本落叶松(Larix kaempferi)人工纯林和落叶松-赤杨混交林为研究对象,以氨基糖作为微生物残留物的生物标识物,研究了赤杨对落叶松人工林根际和非根际土壤氨基糖积累的影响。结果表明,混交林中落叶松根际和非根际各氨基糖单体含量均显著高于纯林,说明赤杨引入有利于土壤微生物残留物的积累。其中,混交林落叶松根际及非根际土壤氨基糖葡萄糖含量分别比纯林高出99.5%(P0.01)和154%(P0.01);胞壁酸含量分别比纯林高出66.1%(P0.01)和132.3%(P0.01)。赤杨引入对氨基葡萄糖/胞壁酸比值的影响不显著,但显著增加氨基糖对土壤有机碳的贡献。冗余度分析表明全氮是驱动土壤中氨基糖发生变化的主要因子。表明赤杨引入显著提高微生物固持的土壤碳库和有机碳稳定性,这对落叶松人工林合理经营具有重要意义。     

Rapid degradation of starch in chloroplasts and concomitant accumulation of soluble sugars associated with ABA-induced freezing tolerance in the moss Physcomitrella patens

Abscisic acid (ABA) has been postulated to play a role in the development of freezing tolerance during the cold acclimation process in higher plants, but its role in cold tolerance in tower land plants has not been elucidated. The moss Physcomitrella patens rapidly developed freezing tolerance when its protonemata were grown in a medium containing ABA, with dramatic changes in the LT50 value from -2 degrees C to over -10 degrees C. We examined physiological and morphological alterations in protonema cells caused by ABA treatment to elucidate early cellular events responsible for rapid enhancement of freezing tolerance. Microscopic observations revealed that ABA treatment for 1 day resulted in a dramatic alteration in the appearance of intracellular organelles. ABA-treated cells had slender chloroplasts, with a reduced amount of starch grains, in comparison with those of non-treated cells. The ABA-treated cells also had several segmented vacuoles while many of non-treated cells had one central vacuole. When frozen to -4 degrees C, freezing injury-associated ultrastructural changes such as formation of aparticulate domains and fracture-jump lesions were frequently observed in the plasma membrane of non-treated protonema cells but not in that of ABA-treated cells. The ABA treatment increased the osmotic concentration of the protonema cells, in correlation with accumulation of free soluble sugars. These results suggest that ABA-induced accumulation of soluble sugars, associated with morphological changes in organelles, mitigated freezing-induced structural damage in the plasma membrane, eventually leading to enhancement of freezing tolerance in the protonema cells.     

弱光对长春花（Catharanthus roseus）幼苗中可溶性糖、生物碱及激素含量的影响

阐述了低光照强度和正常光照条件下长春花（Catharanthus roseus）幼苗叶片中可溶性糖、生物碱及内源激素等几种生理活性物质含量的动态变化,以揭示弱光胁迫对长春花生理代谢的影响及上述生理代谢对植物适应弱光环境的生态学意义。结果显示,弱光培养提高了长春花在自身支撑结构的投入,如叶柄长和节间长都显著增加,同时抑制了有性生殖。在弱光培养的第1周,长春花叶片中果糖（fructose,Fru）、葡萄糖（glucose,Glc）和蔗糖（sucrose,Suc）的含量分别由处理前的（0．62±0．01）,（1．86±0．12）,（0．24±0．01）mg／g FW下降为（0．38±0．02）,（0．60±0．03）,（0．17±0．02）mg／g FW,均显著低于对照（P〈0．01）,表明碳同化水平下降。长春花叶片中文朵灵（vindoline,VIN）和长春质碱（catharanthine,CAT）含量在弱光条件下呈显著增加趋势,处理结束时是对照的3倍左右,而它们的耦合产物长春碱（vinblastine,VLB）合成和积累受到抑制,其含量在第3周时仅为对照的50％。弱光条件处理前2周促进了长春花叶片中脱落酸（abscisic acid,ABA）、赤霉素（gibberelline,GA3）和吲哚乙酸（indole-3-acetic acid,IAA）含量水平的积累。这些结果表明,在弱光条件下长春花幼苗的上述生理代谢都发生着显著的变化,可能在植物适应弱光胁迫过程中发挥着积极的调控作用。     

Recombinant Dendroides canadensis antifreeze proteins as potential ingredients in cryopreservation solutions

Expanding cryopreservation methods to include a wider range of cell types, such as those sensitive to freezing, is needed for maintaining the viability of cell-based regenerative medicine products. Conventional cryopreservation protocols, which include use of cryoprotectants such as dimethylsulfoxide (Me₂SO), have not prevented ice-induced damage to cell and tissue matrices during freezing. A family of antifreeze proteins (AFPs) produced in the larvae of the beetle, Dendroides canadensis allow this insect to survive subzero temperatures as low as −26 °C. This study is an assessment of the effect of the four hemolymph D. canadensis AFPs (DAFPs) on the supercooling (nucleating) temperature, ice structure patterns and viability of the A10 cell line derived from the thoracic aorta of embryonic rat. Cryoprotectant solution cocktails containing combinations of DAFPs in concentrations ranging from 0 to 3 mg/mL in Unisol base mixed with 1 M Me₂SO were first evaluated by cryomicroscopy. Combining multiple DAFPs demonstrated significant supercooling point depressing activity (∼9 °C) when compared to single DAFPs and/or conventional 1 M Me₂SO control solutions. Concentrations of DAFPs as low as 1 μg/mL were sufficient to trigger this effect. In addition, significantly improved A10 smooth muscle cell viability was observed in cryopreservation experiments with low DAFP-6 and DAFP-2 concentrations in combination with Me₂SO. No significant improvement in viability was observed with either DAFP-1 or DAFP-4. Low and effective DAFP concentrations are advantageous because they minimize concerns regarding cell cytotoxicity and manufacturing cost. These findings support the potential of incorporating DAFPs in solutions used to cryopreserve cells and tissues.     

WCS120 protein family and proteins soluble upon boiling in cold-acclimated winter wheat

The amount of proteins soluble upon boiling (especially WCS120 proteins) and the ability to develop frost tolerance (FT) after cold acclimation was studied in two frost-tolerant winter wheat cultivars, Mironovskaya 808 and Bezostaya 1. Protein gel blot analysis, mass spectrometry (MS) and image analysis of two-dimensional gel electrophoresis (2-DE) gels were used to identify and/or quantify the differences in protein patterns before (non-acclimated, NA) and after 3 weeks of cold acclimation (CA) of the wheats, when FT increased from -4 degrees C (lethal temperature (LT(50)), for both cultivars) to -18.6 degrees C in Bezostaya 1 and -20.8 degrees C in Mironovskaya 808. Only WCS120 protein was visible in NA leaves while all five WCS120 proteins were induced in the CA leaves. Mironovskaya 808 had higher accumulation of three members of WCS120 proteins (WCS120, WCS66 and WCS40) than Bezostaya 1. MS analysis of total sample of proteins soluble upon boiling showed seven COR proteins in the CA samples and only three COR proteins in the NA samples of cultivar Mironovskaya 808 (MIR). In conclusion, the level of the accumulation of WCS120, WCS66 and WCS40 distinguished our two frost-tolerant winter wheat cultivars. Moreover, the differences of CA and NA samples of the MIR were shown by liquid chromatography (LC)-tandem mass spectrometry (MS/MS).     

Legumin- and vicilin-like proteins from spores of the fern Matteuccia struthiopteris

Legumin- and vicilin-like proteins have been isolated from spores of the fern Matteuccia struthiopteris. Their relationship with seed legumin and vicilin was demonstrated by cross-reactivities of antibodies directed against respective storage globulins from Vicia faba as evidenced by Western blotting. The Matteuccia legumin-like protein was characterised as a 300-340 kDa holoprotein preferentially consisting of a 32 kDa alpha-chain and a 24 kDa beta-chain. Patterns of limited proteolysis of the spore legumin-like protein and seed legumins were similar as well. In contrast to seed legumins, the Matteuccia legumin-like protein is devoid of disulfide bridges between alpha- and beta-chains. A 52 kDa polypeptide of the Matteuccia vicilin-like protein, first detected by SDS gel electrophoresis, is probably encoded by a vicilin-like gene specifically expressed in Matteuccia struthiopteris spores (Shutov et al. 1998). The vicilin-like holoprotein was found to form a complex of 600 kDa apparent molecular mass, presumably composed of four vicilin-like trimers.     

Sperm cryopreservation of the critically endangered olive barb (Sarpunti) Puntiussarana (Hamilton, 1822)

The present study focused on development of a sperm cryopreservation protocol for the critically endangered olive barb Puntiussarana (Hamilton, 1822) collected from two stocks within Bangladesh and reared in the Fisheries Field Laboratory, Bangladesh Agricultural University (BAU). The sperm were collected in Alsever’s solution prepared at 296 mOsmol kg⁻¹. Sperm were activated with distilled water (24 mOsmol kg⁻¹) to characterize motility. Maximum motility (90%) was observed within 15 s after activation, and sperm remained motile for 35 s. Sperm activation was evaluated in different osmolalities and motility was completely inhibited when osmolality of the extender was ?287 mOsmol kg⁻¹. To evaluate cryoprotectant toxicity, sperm were equilibrated with 5%, 10% and 15% each of dimethyl sulfoxide (DMSO) and methanol. Sperm motility was noticeably reduced within 10 min, when sperm were equilibrated with 15% DMSO, indicating acute toxicity to spermatozoa and therefore this concentration was excluded in further trials. Sperm were cryopreserved using DMSO at concentrations of 5% and 10% and methanol at 5%, 10% and 15%. The one-step freezing protocol (from 5 °C to −80 °C at 10 °C/min) was carried out in a computer-controlled freezer (FREEZE CONTROL® CL-3300; Australia) and 0.25-ml straws containing spermatozoa were stored in liquid nitrogen for 7–15 days at −196 °C. The highest motility in thawed sperm 61 ± 8% (mean ± SD) was obtained with 10% DMSO. The fertilization and hatching rates were 70% and 37% for cryopreserved sperm, and 72% and 62% for fresh sperm. The protocol reported here can be useful for hatchery-scale production of olive barb. The use of cryopreserved sperm can facilitate hatchery operations, and can provide for long-term conservation of genetic resources to contribute in the recovery of critically endangered fish such as the olive barb.     

The transport and mediation mechanisms of the common sugars in Escherichia coli

Escherichia coli can uptake and utilize many common natural sugars to form biomass or valuable target bio-products. Carbon catabolite repression (CCR) will occur and hamper the efficient production of bio-products if E. coli strains are cultivated in a mixture of sugars containing some preferred sugar, such as glucose. Understanding the transport and metabolism mechanisms of the common and inexpensive sugars in E. coli is important for further improving the efficiency of sugar bioconversion and for reducing industrial fermentation costs using the methods of metabolic engineering, synthetic biology and systems biology. In this review, the transport and mediation mechanisms of glucose, fructose, sucrose, xylose and arabinose are discussed and summarized, and the hierarchical utilization principles of these sugars are elucidated.     

Elicitation of alkaloids in in vitro PLB (protocorm-like body) cultures of Pinellia ternata

The objective of this study was to determine the effect of two endophytic bacterial elicitors (Pseudomonas sp. and Enterobacter sp.) on the production of alkaloids in protocorm-like bodies (PLBs) of Pinellia ternata Breit. Both bacterial strains increased the growth rate of P. ternata PLBs. Pseudomonas sp. promoted the differentiation of the PLBs, whereas Enterobacter sp. inhibited PLB differentiation. The bacterial strains increased guanosine production in PLBs by 9–166%, inosine production by 2–33%, and trigonelline production by 114–1140% compared to the control. For Pseudomonas sp., guanosine and trigonelline production was greater when bacterial extracts were added to the PLB suspension cultures rather than living cells (co-culture treatment). Inosine production was similar in both the bacterial extract and co-culture treatments. For the Enterobacter sp., guanosine, inosine, and trigonelline production tended to be greatest when living cells were added to the PLB suspension cultures rather than bacterial extracts. These results suggest that Pseudomonas sp. and Enterobacter sp. could increase alkaloid yield from P. ternata under field or tissue culture conditions. We also observed that Pseudomonas sp. and Enterobacter sp. produced some of the same alkaloids as their host plants. Additional study needs to be done to determine if these endophytic bacteria could be used to produce alkaloids in the fermentation industry.     

Induction of PR proteins and resistance by the biocontrol agent Clonostachys rosea in wheat plants infected with Fusarium culmorum

Clonostachys rosea (CR) is a common worldwide saprophyte with destructive effect against several plant pathogenic fungi showing antagonistic features against a wide variety of pathogens. We recently isolated a strain of C. rosea, named CR47, from wheat crown infected with Fusarium culmorum (FC); this strain proved to be effective against Fusarium seed borne diseases of cereals under field condition. In this paper the function of C. rosea applied as seed treatment on wheat seedling growth was investigated. In addition, we investigated the expression pattern of peroxidases and chitinases as well as PR4 proteins following both CR treatments of seeds and FC infection and also in the three-component system pathogen–antagonist–wheat. Several chitinase isoforms were induced by CR-treatment both in coleoptiles and roots, whereas some peroxidase isoforms were induced only in the presence of both antagonist and pathogen. In the latter case, it seems that CR-treatment by itself promotes plant growth and reduces the peroxidase expression, while enhances some chitinase isoforms probably involved in cell wall disruption. Moreover, both the antagonist and the pathogen studied induced PR4 protein expression, which probably exerts its role on the invading microorganisms by a translation-inhibitory process that could be ascribed to their ribonuclease activity.     

Motility and cryopreservation of spermatozoa of European common frog, Rana temporaria

Motility and cryopreservation of testicular sperm of European common frog, Rana temporaria were investigated. Collected testicular spermatozoa were immotile in solutions of high osmolalities: 300 mmol/l sucrose and motility inhibiting saline solution-MIS. Full sperm motility could be activated in distilled water or in a solution of 50 mmol/l NaCl, = 90 mosmol/kg, with 75-90% motility and 14-16 μm s⁻¹ swimming velocity. Spermatozoa activated in distilled water and kept at room temperature ceased the motility within a period of 1 h. But when they were kept at 4 °C, no significant decrease in sperm motility and velocity occurred over a period of 1 h. Incubation of testicular sperm diluted 1:2 with MIS containing 10% DMSO, 5% glycerol, 10% methanol, or 10% propandiol for a period of 40 min at 4 °C showed that propandiol was the most toxic cryoprotectant for spermatozoa of European common frog R. temporaria. However, methanol was not toxic to spermatozoa during the 40 min incubation period, it failed to protect spermatozoa during the freezing and thawing process. DMSO and glycerol were useful penetrating cryoprotectants that interacted with sperm diluents in cryodiluent efficacy. In combination with the sucrose diluent, DMSO was a better cryoprotectant than glycerol, while in combination with MIS, DMSO and glycerol were similarly useful. Sperm was frozen at two freezing levels above the surface of liquid nitrogen. Sperm frozen 5 cm above the surface of liquid nitrogen resulted in immotile and non-viable spermatozoa. However, sperm frozen at 10 cm above the surface of liquid nitrogen showed 40-45% viability and 30-35% motility, compared to the untreated freshly collected testicular sperm. Addition of hen egg yolk had no positive effect on the post-thaw sperm motility, viability and hatching rate when added to sucrose cryodiluents. However, addition of 5% egg yolk to the MIS containing 5% glycerol and 2.5% sucrose significantly improved the hatching rate than all other treatments. Therefore, we conclude that, MIS and 300 mmol/l sucrose are suitable diluents for immotile storage of testicular semen. For cryopreservation, dilution to a final concentration of 5-6 × 10⁶/ml in MIS with 5% glycerol, 2.5% sucrose and 5% egg yolk, frozen in liquid nitrogen vapour at 10 cm above its surface, and thawed at 22 °C for 40 s is a useful cryopreservation protocol for R. temporaria sperm. Further research is needed to determine the motility parameters and cryopreservation of spermatic urine of R. temporaria.     

A two-dimensional electrophoretic profile of the proteins secreted by Herbaspirillum seropedicae strain Z78

Herbaspirillum seropedicae is an endophytic bacterium that associates with rice, sugarcane and other economically important crops. Secreted proteins play a key role in the plant–bacterial interaction. Using 2D electrophoresis and peptide mass fingerprint mass spectrometry, 63 protein spots representing 41 different secreted proteins were identified during growth of H. seropedicae under nitrogen-sufficient conditions. In silico analysis showed that 25.4% of the proteins had signal peptides and 15.9% were predicted to be non-classically secreted. Among the most abundant were flagellar components and ABC-type transport system proteins. Nine secreted proteins had also been identified in the cellular proteome, suggesting that they also play a role in the extracellular environment. No type III secreted proteins were detected by comparison of the wild type strain with an hrcN mutant strain.