Two-dimensional Protein Profiles Research Articles

Identification of Stress-related Proteins during the Growth and Development of Piper nigrumL.

Piper nigrum L., also known as the pepper plant, produces pepper berries, which is a spice used worldwide. Changes in the natural environment have forced the plant to undergo acclimatization for plant growth. Consequently, un-adapted plants developed unhealthy, stunted growth, affecting the production of pepper berries, and thus, causing loss to agriculture. Proteomic analysis using two-dimensional polyacrylamide gel electrophoresis (PAGE) coupled with mass spectrometry was used to compare the differentially expressed proteins between healthy and unhealthy pepper plants. The proteins were successfully extracted from the stems and roots of P. nigrum and the two-dimensional protein profiles were compared. Two-dimensional analysis has indicated significant differences between the healthy and unhealthy parts with 41 and 25 differentially expressed proteins in the stems and roots respectively. A total of 10 differentially expressed proteins, characterized as unique (2) and over-expressed (8) proteins, from both plant parts were selected and sent for identification through matrix-assisted laser desorption/ionization, time-of-flight mass spectrometry. The unique and over-expressed proteins in the unhealthy stems were identified as stress-related small heat shock proteins (sHSPs). These proteins act as a chaperone to protect other proteins against a stressful environment for plant defence and adaptation. Meanwhile, over-expressed proteins from the roots were found to be peroxidase 5, malate dehydrogenase, mitochondrial and gamma-glutamyl phosphate reductase which are involved in oxidative stress, photosynthetic activity, and plant development, respectively. The findings of the differentially expressed proteins in this study, therefore, may create opportunities for further investigations of stress management control and help to improve plantation practices in the future.

Two-dimensional electrophoresis protein profiles of HL-60 and CCRF-CEM cell lines treated with epigenetic modification drugs

Leukemia is classified as a malignant disease of hematopoietic stem cells (HSCs) that fails in cell differentiation but preserve their self-renewal. It is caused by genetic alterations and epigenetic modifications resulting in the activation or inactivation of particular genes for transcription. Epigenetic causes changes in gene expression without any alteration in the DNA sequence. The most common epigenetic modifications are DNA methylation and histone acetylation. 5-Azacitidine (5-Aza) is a DNA methytransferase inhibitor (DNMTi) that inhibits DNA methyltransferase enzymes resulting in hypomethylation. Trichostatin A (TSA) is a histone deacetylase inhibitor which inhibits deacetylation of both histone and non-histone proteins resulting in chromatin relaxation. This present study focused on the alteration of proteome profile on 2D gel electrophoresis (2-DE) induced by 5-Aza and TSA in HL-60 and CCRF-CEM cell lines as in vitro model to represent acute promyelocytic leukemia (APL) and T-lymphoblastic leukemia (T-ALL), respectively. Total proteins of untreated and 5-Aza/TSA-treated HL-60 and CCRF-CEM cell lines were extracted using urea/thiourea buffer and stained with Coomassie Blue. Comparative analysis of untreated and 5-Aza/TSA-treated HL-60 and CCRF-CEM was performed by PDQuest software. Qualitative analysis identified 190-659 protein spots detected in untreated, 5-Aza and TSA-treated HL-60 and CCRF-CEM. Quantitative comparison analysis was analyzed by over 2-fold change in 5-Aza/TSA-treated cells compared to untreated. One and eight upregulated proteins were detected in 5-Aza and TSA-treated HL-60, respectively. While five and one upregulated proteins were detected in 5-Aza and TSA-treated CCRF-CEM, respectively. These preliminary results suggested that 5-Aza and TSA induced proteome profiles alterations due to their inhibition effects in HL-60 and CCRF-CEM cell lines.

Protein changes in the shoot-tips of vanilla (Vanilla planifolia) in response to osmoprotective treatments

Cryogenic storage of vanilla shoot-tips represents the safest biotechnological strategy for the long-term conservation of the vanilla germplasm, but successful cryopreservation depends on its tolerance to both dehydration stress imposed by cryoprotective treatments and thermal stress produced by immersion in liquid nitrogen. In this work, we evaluated the impact of various osmoprotective treatments on protein expression patterns in vanilla (Vanilla planifolia) shoot-tips subjected to successive dehydration steps prior to cryopreservation. Two-dimensional electrophoretic protein profiles of shoot-tips dissected from in vitro grown plants and preconditioned on semisolid media with 0.3 M sucrose for one day, and shoot-tips preconditioned, loaded with a solution of 0.4 M sucrose and 2 M glycerol, and subsequently exposed to plant vitrification solution 3 (50% (w/v) sucrose and 50% (w/v) glycerol), were compared with non-treated dissected shoot-tips. We observed an increase in the expression level of six protein spots (fold change exceeding 1.5) and a decrease (fold change not exceeding 0.6) of ten protein spots after preconditionig treatment, whereas the profiles after preconditioning, loading and exposure to vitrification solution showed an increase in the expression level of 21 protein spots and a decrease in the expression level of 13. Most proteins identified were down-regulated and belonged to groups of biosynthesis, folding, and protein degradation. Many others were related to energetic metabolism, defense, and cell structure. These preliminary results contribute to knowledge of the proteome of this species and partially clarify its sensitivity to osmotic dehydration treatments.

Two-dimensional protein profiles of drought tolerant tomato (Solanum lycopersicum) mutants improved by gamma radiation

Two-dimensional protein profiles of drought tolerant tomato (Solanum lycopersicum) mutants improved by gamma radiation

Differentially expressed proteins associated with Fusarium head blight resistance in wheat.

BackgroundFusarium head blight (FHB), mainly caused by Fusarium graminearum, substantially reduces wheat grain yield and quality worldwide. Proteins play important roles in defense against the fungal infection. This study characterized differentially expressed proteins between near-isogenic lines (NILs) contrasting in alleles of Fhb1, a major FHB resistance gene in wheat, to identify proteins underlining FHB resistance of Fhb1.MethodsThe two-dimensional protein profiles were compared between the Fusarium-inoculated spikes of the two NILs collected 72 h after inoculation. The protein profiles of mock- and Fusarium-inoculated Fhb1+NIL were also compared to identify pathogen-responsive proteins.ResultsEight proteins were either induced or upregulated in inoculated Fhb1+NIL when compared with mock-inoculated Fhb1+NIL; nine proteins were either induced or upregulated in the Fusarium-inoculated Fhb1+NIL when compared with Fusarium-inoculated Fhb1−NIL. Proteins that were differentially expressed in the Fhb1+NIL, not in the Fhb1−NIL, after Fusarium inoculation included wheat proteins for defending fungal penetration, photosynthesis, energy metabolism, and detoxification.ConclusionsCoordinated expression of the identified proteins resulted in FHB resistance in Fhb1+NIL. The results provide insight into the pathway of Fhb1-mediated FHB resistance.

A proteomic view of the response of Paracoccidioides yeast cells to zinc deprivation

Zinc plays a critical role in a diverse array of biochemical processes. However, an excess of zinc is deleterious to cells; therefore, cells require finely tuned homeostatic mechanisms to balance the uptake and the storage of zinc. There is also increasing evidence supporting the importance of zinc during infection. To understand better how Paracoccidioides adapts to zinc deprivation, we compared the two-dimensional (2D) gel protein profile of yeast cells during zinc starvation to yeast cells grown in a zinc rich condition. Protein spots were selected for comparative analysis based on the protein staining intensity, as determined by image analysis. In response to zinc deprivation, a total of 423 out of 845 protein spots showed a significant change in abundance. Quantitative RT-qPCR analysis of RNA from Paracoccidioides grown under zinc restricted conditions validated the correlation between the differentially regulated proteins and transcripts. According to the proteomic data, zinc deficiency may be a stressor to Paracoccidioides, as suggested by the upregulation of a number of proteins related to stress response, cell rescue, and virulence. Other process induced by zinc deprivation included gluconeogenesis. Conversely, the methylcitrate cycle was downregulated. Overall, the results indicate a remodelling of the Paracoccidioides response to the probable oxidative stress induced during zinc deprivation.

Differential expression and sequence polymorphism of the olive pollen allergen Ole e 1 in two Iranian cultivars.

Molecular evidence on the heterogeneity present in the Ole e 1 allergen of the olive pollen is emerging. Such polymorphism is dependent on the cultivar origin of pollen, which also determines wide differences in the expression of this protein. Determination of biochemical and molecular characteristics of Ole e 1 pollen allergen in two Iranian olive cultivars, namely 'Rowghani' and 'Zard' is necessary to assess their allergenicity potential. SDS-PAGE and immunoblotting analysis of pollen extracts showed that both cultivars present high and low expression of Ole e 1, respectively. These protein levels correlated with similarly different levels of transcripts, as determined by RT-PCR. Two-dimensional protein profiles also showed conspicuous differences in the distribution and the level of expression of those spots reacting to an anti-Ole e 1 antibody. Bioinformatic analysis of four Ole e 1 sequences corresponding to 'Rowghani' and two sequences for 'Zard', showed numerous heterogeneities when compared with those Ole e 1 and Ole e 1-like sequences present in databases. Nucleotide substitutions resulted in many cases in changes over the predicted amino acid sequences. A cladistic analysis of the sequences showed Iranian entries in a central position between West-European sequences, and Ole e 1-like sequences from other Oleaceae species. Moreover, amino acid changes affected key epitopes of the protein involved in the recognition of the protein by the human immune system. Putative implications of polymorphism in both the biological role and the allergic reactivity of Ole e 1 are discussed.

Alteration of protein patterns in black rock inhabiting fungi as a response to different temperatures

Rock inhabiting fungi are among the most stress tolerant organisms on Earth. They are able to cope with different stressors determined by the typical conditions of bare rocks in hot and cold extreme environments. In this study first results of a system biological approach based on two-dimensional protein profiles are presented. Protein patterns of extremotolerant black fungi –Coniosporium perforans, Exophiala jeanselmei – and of the extremophilic fungus –Friedmanniomyces endolithicus – were compared with the cosmopolitan and mesophilic hyphomycete Penicillium chrysogenum in order to follow and determine changes in the expression pattern under different temperatures. The 2D protein gels indicated a temperature dependent qualitative change in all the tested strains. Whereas the reference strain P. chrysogenum expressed the highest number of proteins at 40 °C, thus exhibiting real signs of temperature induced reaction, black fungi, when exposed to temperatures far above their growth optimum, decreased the number of proteins indicating a down-regulation of their metabolism. Temperature of 1 °C led to an increased number of proteins in all of the analysed strains, with the exception of P. chrysogenum. These first results on temperature dependent reactions in rock inhabiting black fungi indicate a rather different strategy to cope with non-optimal temperature than in the mesophilic hyphomycete P. chrysogenum.

Liver-specific loss of glucose-regulated protein 78 perturbs the unfolded protein response and exacerbates a spectrum of liver diseases in mice

The endoplasmic reticulum (ER) chaperone protein glucose-regulated protein 78 (GRP78)/binding immunoglobulin protein is a master regulator of ER homeostasis and stress responses, which have been implicated in the pathogenesis of metabolic disorders. By applying the locus of X-over P1-cyclization recombination strategy, we generated mice with liver-specific GRP78 loss. Our studies using this novel mouse model revealed that liver GRP78 was required for neonatal survival, and a loss of GRP78 in the adult liver greater than 50% caused an ER stress response and dilation of the ER compartment, which was accompanied by the onset of apoptosis. This suggested the critical involvement of GRP78 in maintaining hepatocyte ER homeostasis and viability. Furthermore, these mice exhibited elevations of serum alanine aminotransferase and fat accumulation in the liver, and they were sensitized to a variety of acute and chronic hepatic disorders by alcohol, a high-fat diet, drugs, and toxins. These disorders were alleviated by the simultaneous administration of the molecular chaperone 4-phenylbutyrate. A microarray analysis and a two-dimensional protein profile revealed major perturbations of unfolded protein response targets, common enzymes/factors in lipogenesis, and new factors possibly contributing to liver steatosis or fibrosis under ER stress (e.g., major urinary proteins in the liver, fatty acid binding proteins, adipose differentiation-related protein, cysteine-rich with epidermal growth factor-like domains 2, nuclear protein 1, and growth differentiation factor 15). Our findings underscore the importance of GRP78 in managing the physiological client protein load and suppressing apoptosis in hepatocytes, and they support the pathological role of ER stress in the evolution of fatty liver disease under adverse conditions (i.e., drugs, diet, toxins, and alcohol).

Proteome profiles in medaka ( Oryzias melastigma) liver and brain experimentally exposed to acute inorganic mercury

Mercury is a widespread and persistent pollutant occurring in a variety of forms in freshwater and marine ecosystems. Using the proteomic approach, this study examined the protein profiles of the medaka ( Oryzias melastigma) liver and brain exposed to an acute mercuric chloride (HgCl 2) concentration (1000 μg/L) for 8 h. The results showed that acute exposure of medaka to inorganic mercury enhanced metal accumulation in both the liver and brain, and a higher content of mercury was detected in the latter. Comparison of the two-dimensional electrophoresis protein profiles of HgCl 2-exposed and non-exposed group revealed that altered protein expression was quantitatively detected in 20 spots in the brain and 27 in the liver. The altered protein spots were subjected to matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry analysis, with the resultant identification of 46 proteins. The proteins identified were involved in oxidative stress, cytoskeletonal assembly, signal transduction, protein modification, metabolism and other related functions (e.g. immune response, ionoregulation and transporting), highlighting the fact that inorganic mercury toxicity in fish seems to be complex and diverse. This study provided basic information to aid our understanding of the possible molecular mechanisms of acute inorganic mercury toxicity in aquatic organisms, as well as potential protein biomarker candidates for aquatic environmental monitoring.

Protein profiles in zebrafish ( Danio rerio) brains exposed to chronic microcystin-LR

Microcystin-LR (MCLR) is a commonly encountered blue-green algal hepatotoxin and a known inhibitor of cellular protein phosphatase (PP), however, little is known about its neurotoxicity. This study investigated the protein profiles of zebrafish ( Danio rerio) brains chronically exposed to MCLR concentrations (2 or 20 μg L −1) using the proteomic approach. The results showed that MCLR strikingly enhanced toxin accumulation and the PP activity in zebrafish brains after 30 d exposure. Comparison of two-dimensional electrophoresis protein profiles of MCLR exposed and non-exposed zebrafish brains revealed that the abundance of 30 protein spots was remarkably altered in response to MCLR exposure. These proteins are involved in cytoskeleton assembly, macromolecule metabolism, oxidative stress, signal transduction, and other functions (e.g. transporting, protein degradation, apoptosis and translation), indicating that MCLR toxicity in the fish brain is complex and diverse. The chronic neurotoxicity of MCLR might initiate the PP pathway via an upregulation of PP2C in the zebrafish brain, in addition to the reactive oxygen species pathway. Additionally, the increase of vitellogenin abundance in MCLR exposed zebrafish brains suggested that MCLR might mimic the effects of endocrine disrupting chemicals. This study demonstrated that MCLR causes neurotoxicity in zebrafish at the proteomic level, which provides a new insight into MCLR toxicity in aquatic organisms and human beings.

ClpP of Streptococcus mutans Differentially Regulates Expression of Genomic Islands, Mutacin Production, and Antibiotic Tolerance

Streptococcus mutans is the primary etiological agent of human dental caries and, at times, of infective endocarditis. Within the oral cavity, the pathogen is subjected to conditions of stress. A well-conserved protein complex named ClpP (caseinolytic protease) plays a vital role in adaptation under stress conditions. To gain a better understanding of the global role of the ClpP protease in cellular homeostasis, a transcriptome analysis was performed using a DeltaclpP mutant strain. The expression levels of more than 100 genes were up- or downregulated in the DeltaclpP mutant compared to the wild type. Notably, the expression of genes in several genomic islands, such as TnSmu1 and TnSmu2, was differentially modulated in the DeltaclpP mutant strain. ClpP deficiency also increased the expression of genes associated with a putative CRISPR locus. Furthermore, several stress-related genes and genes encoding bacteriocin-related peptides and many transcription factors were also found to be altered in the DeltaclpP mutant strain. A comparative analysis of the two-dimensional protein profile of the wild type and the DeltaclpP mutant strains showed altered protein profiles. Comparison of the transcriptome data with the proteomic data identified four common gene products, suggesting that the observed altered protein expression of these genes could be due to altered transcription. The results presented here indicate that ClpP-mediated proteolysis plays an important global role in the regulation of several important traits in this pathogen.

Development of handed body asymmetry in mammals.

We have proposed a three step model for the specification of left-right in mammalian embryos. The fundamental assumption is that handedness is imparted by an asymmetrical molecule. Conversion of molecular asymmetry to the cellular level gives a property to one side of the embryo to bias an otherwise random generation of an asymmetrical gradient which can be interpreted by developing organs. Rat embryos, treated at discrete stages, show a window of sensitivity for disruption of handedness, which may reflect the time of conversion/biasing. Heat shock and several chemicals cause left-right inversion in up to 50% of embryos exposed during neural groove formation. Earlier stages are less sensitive; no treatment begun after foregut pocket formation influences asymmetry. Evidence for cellular interactions in left-right specification comes from the apparent rescue of iv/iv mutant embryos in chimeras. We are looking for molecular left-right disparity before morphological asymmetry but detect no differences in two-dimensional protein profiles. Using an indirect measure, we find a right-left gradient of tissue oxygen in embryos at the 20-30 somite stage. This may reflect asymmetrical vasculature, as we have suggested to explain drug-induced asymmetrical limb malformations.

Proteomic Analysis of Seed Dormancy in Arabidopsis

The mechanisms controlling seed dormancy in Arabidopsis (Arabidopsis thaliana) have been characterized by proteomics using the dormant (D) accession Cvi originating from the Cape Verde Islands. Comparative studies carried out with freshly harvested dormant and after-ripened non-dormant (ND) seeds revealed a specific differential accumulation of 32 proteins. The data suggested that proteins associated with metabolic functions potentially involved in germination can accumulate during after-ripening in the dry state leading to dormancy release. Exogenous application of abscisic acid (ABA) to ND seeds strongly impeded their germination, which physiologically mimicked the behavior of D imbibed seeds. This application resulted in an alteration of the accumulation pattern of 71 proteins. There was a strong down-accumulation of a major part (90%) of these proteins, which were involved mainly in energetic and protein metabolisms. This feature suggested that exogenous ABA triggers proteolytic mechanisms in imbibed seeds. An analysis of de novo protein synthesis by two-dimensional gel electrophoresis in the presence of [(35)S]-methionine disclosed that exogenous ABA does not impede protein biosynthesis during imbibition. Furthermore, imbibed D seeds proved competent for de novo protein synthesis, demonstrating that impediment of protein translation was not the cause of the observed block of seed germination. However, the two-dimensional protein profiles were markedly different from those obtained with the ND seeds imbibed in ABA. Altogether, the data showed that the mechanisms blocking germination of the ND seeds by ABA application are different from those preventing germination of the D seeds imbibed in basal medium.

Comparative proteomic analysis of metabolically labelled proteins from Plasmodium falciparum isolates with different adhesion properties

The virulence of Plasmodium falciparum relates in part to the cytoadhesion characteristics of parasitized erythrocytes but the molecular basis of the different qualitative and quantitative binding phenotypes is incompletely understood. This paucity of information is due partly to the difficulty in working with membrane proteins, the variant nature of these surface antigens and their relatively low abundance. To address this two-dimensional (2D) protein profiles of closely related, but phenotypically different laboratory strains of P. falciparum have been characterized using proteomic approaches. Since the mature erythrocyte has no nucleus and no protein synthesis capability, metabolic labelling of proteins was used to selectively identify parasite proteins and increase detection sensitivity.A small number of changes (less than 10) were observed between four different P. falciparum laboratory strains with distinctive cytoadherence properties using metabolic labelling, with more parasite protein changes found in trophozoite iRBCs than ring stage. The combination of metabolic labelling and autoradiography can therefore be used to identify parasite protein differences, including quantitative ones, and in some cases to obtain protein identifications by mass spectrometry. The results support the suggestion that the membrane protein profile may be related to cytoadherent properties of the iRBCs. Most changes between parasite variants were differences in iso-electric point indicating differential protein modification rather than the presence or absence of a specific peptide.

Two-dimensional electrophoresis protein profile of the phytopathogenic fungus Botrytis cinerea

Botrytis cinerea is a phytopathogenic fungi causing disease in a number of important crops. It is considered a very complex species in which different populations seem to be adapted to different hosts. In order to characterize fungal virulence factors, a proteomic research was started. A protocol for protein extraction from mycelium tissue, with protein separation by 2-DE and MS analysis, was optimised as a first approach to defining the B. cinerea proteome. Around 400 spots were detected in 2-DE CBB-stained gels, covering the 5.4-7.7 pH and 14-85 kDa ranges. The averages of analytical and biological coefficients of variance for 64 independent spots were 16.1% and 37.5%, respectively. Twenty-two protein spots were identified by MALDI-TOF or ESI IT MS/MS, with some of them corresponding to forms of malate dehydrogenase and glyceraldehyde-3-phosphate dehydrogenase. Two more spots matched a cyclophilin and a protein with an unknown function.

Proteomic Analysis of Different Mutant Genotypes of Arabidopsis Led to the Identification of 11 Proteins Correlating with Adventitious Root Development

A lack of competence to form adventitious roots by cuttings or explants in vitro occurs routinely and is an obstacle for the clonal propagation and rapid fixation of elite genotypes. Adventitious rooting is known to be a quantitative genetic trait. We performed a proteomic analysis of Arabidopsis (Arabidopsis thaliana) mutants affected in their ability to develop adventitious roots in order to identify associated molecular markers that could be used to select genotypes for their rooting ability and/or to get further insight into the molecular mechanisms controlling adventitious rooting. Comparison of two-dimensional gel electrophoresis protein profiles resulted in the identification of 11 proteins whose abundance could be either positively or negatively correlated with endogenous auxin content, the number of adventitious root primordia, and/or the number of mature adventitious roots. One protein was negatively correlated only to the number of root primordia and two were negatively correlated to the number of mature adventitious roots. Two putative chaperone proteins were positively correlated only to the number of primordia, and, interestingly, three auxin-inducible GH3-like proteins were positively correlated with the number of mature adventitious roots. The others were correlated with more than one parameter. The 11 proteins are predicted to be involved in different biological processes, including the regulation of auxin homeostasis and light-associated metabolic pathways. The results identify regulatory pathways associated with adventitious root formation and represent valuable markers that might be used for the future identification of genotypes with better rooting abilities.

Evaluation of the role of sigma B in Mycobacterium smegmatis

The alternate sigma factor, sigB, is known to play a crucial role in maintaining the stationary phase in mycobacteria. In this communication, we have studied the proteomics of Mycobacterium smegmatis mc 2155 and its two derivatives, one of which has a disrupted sigB gene and the other, PMVSigB, which contains a multicopy plasmid containing sigB. We have identified by two-dimensional gel analyses, several proteins that are over-expressed in PMVSigB compared to mc 2155. These proteins are either stress proteins or participate actively in different metabolic pathways of the organisms. On the other hand, when sigB deleted mycobacteria were grown until the stationary phase and its two-dimensional protein profile was compared to that of mc 2155, few DNA binding proteins were found to be up-regulated. We have shown recently that upon over-expressing sigB, the cell surface glycopeptidolipids of M. smegmatis are hyperglycosylated, a situation similar to what was observed for nutritionally starved bacteria. Gene expression profile through quantitative PCR presented here identified a Rhamnosyltransferase responsible for this hyperglycosylation.

Abscisic acid promoted changes in the protein profiles of rice seedling by proteome analysis

This study investigates the influence of exogenously applied abscisic acid (ABA) on the leaves and leaf sheaths of two-week-old rice seedling at the level of the proteome. Significant differences were observed in the two-dimensional polyacrylamide gel electrophoresis protein profiles between control and ABA treated samples. Amino-acid sequence analysis of affected proteins revealed that ABA caused drastic changes in the major photosynthetic protein, ribulose 1,5-bisphosphate carboxylase/oxygenase and accumulation of certain defense/stress-related proteins. Moreover, cutting or treating leaf sheaths with jasmonic acid (JA) rapidly increased the endogenous level of ABA, suggesting a role for ABA during the defense/stress-response. Comparative study indicated a potential overlap between ABA and JA as detected at the level of the proteome. Furthermore, in vitro protein phosphorylation experiments and in-gel kinase assays also revealed considerable changes in the phosphorylation status of some proteins, and differential effects on myelin basic protein and calcium-dependent protein kinase activities by ABA treatment, which suggests involvement of kinase in the downstream signaling cascade. These results provide evidence at proteome level for the involvement of ABA in stress-response in rice seedling.

Characterization of Ferroplasma isolates and Ferroplasma acidarmanus sp. nov., extreme acidophiles from acid mine drainage and industrial bioleaching environments.

Three recently isolated extremely acidophilic archaeal strains have been shown to be phylogenetically similar to Ferroplasma acidiphilum Y(T) by 16S rRNA gene sequencing. All four Ferroplasma isolates were capable of growing chemoorganotrophically on yeast extract or a range of sugars and chemomixotrophically on ferrous iron and yeast extract or sugars, and isolate "Ferroplasma acidarmanus" Fer1(T) required much higher levels of organic carbon. All four isolates were facultative anaerobes, coupling chemoorganotrophic growth on yeast extract to the reduction of ferric iron. The temperature optima for the four isolates were between 35 and 42 degrees C and the pH optima were 1.0 to 1.7, and "F. acidarmanus" Fer1(T) was capable of growing at pH 0. The optimum yeast extract concentration for "F. acidarmanus" Fer1(T) was higher than that for the other three isolates. Phenotypic results suggested that isolate "F. acidarmanus" Fer1(T) is of a different species than the other three strains, and 16S rRNA sequence data, DNA-DNA similarity values, and two-dimensional polyacrylamide gel electrophoresis protein profiles clearly showed that strains DR1, MT17, and Y(T) group as a single species. "F. acidarmanus" Fer1(T) groups separately, and we propose the new species "F. acidarmanus" Fer1(T) sp. nov.