Discovery of Plant Phenolic Compounds That Act as Type III Secretion System Inhibitors or Inducers of the Fire Blight Pathogen,Erwinia amylovora

Erwinia amylovora causes a devastating disease called fire blight in rosaceous plants. The type III secretion system (T3SS) is one of the important virulence factors utilized by E. amylovora in order to successfully infect its hosts. By using a green fluorescent protein (GFP) reporter construct combined with a high-throughput flow cytometry assay, a library of phenolic compounds and their derivatives was studied for their ability to alter the expression of the T3SS. Based on the effectiveness of the compounds on the expression of the T3SS pilus, the T3SS inhibitors 4-methoxy-cinnamic acid (TMCA) and benzoic acid (BA) and one T3SS inducer, trans-2-(4-hydroxyphenyl)-ethenylsulfonate (EHPES), were chosen for further study. Both the T3SS inhibitors (TMCA and BA) and the T3SS inducer (EHPES) were found to alter the expression of T3SS through the HrpS-HrpL pathway. Additionally, TMCA altered T3SS expression through the rsmB_Ea-RsmA_Ea system. Finally, we found that TMCA and BA weakened the hypersensitive response (HR) in tobacco by suppressing the T3SS of E. amylovora. In our study, we identified phenolic compounds that specifically targeted the T3SS. The T3SS inhibitor may offer an alternative approach to antimicrobial therapy by targeting virulence factors of bacterial pathogens.     

Pilin and Sortase Residues Critical for Endocarditis- and Biofilm-Associated Pilus Biogenesis in Enterococcus faecalis

Enterococci commonly cause hospital-acquired infections, such as infective endocarditis and catheter-associated urinary tract infections. In animal models of these infections, a long hairlike extracellular protein fiber known as the endocarditis- and biofilm-associated (Ebp) pilus is an important virulence factor for Enterococcus faecalis. For Ebp and other sortase-assembled pili, the pilus-associated sortases are essential for fiber formation as they create covalent isopeptide bonds between the sortase recognition motif and the pilin-like motif of the pilus subunits. However, the molecular requirements governing the incorporation of the three pilus subunits (EbpA, EbpB, and EbpC) have not been investigated in E. faecalis. Here, we show that a Lys residue within the pilin-like motif of the EbpC subunit was necessary for EbpC polymerization. However, incorporation of EbpA into the pilus fiber only required its sortase recognition motif (LPXTG), while incorporation of EbpB only required its pilin-like motif. Only the sortase recognition motif would be required for incorporation of the pilus tip subunit, while incorporation of the base subunit would only require the pilin recognition motif. Thus, these data support a model with EbpA at the tip and EbpB at the base of an EbpC polymer. In addition, the housekeeping sortase, SrtA, was found to process EbpB and its predicted catalytic Cys residue was required for efficient cell wall anchoring of mature Ebp pili. Thus, we have defined molecular interactions involved in fiber polymerization, minor subunit organization, and pilus subcellular compartmentalization in the E. faecalis Ebp pilus system. These studies advance our understanding of unique molecular mechanisms of sortase-assembled pilus biogenesis.     

Pilus Gene Pool Variation and the Virulence of Corynebacterium diphtheriae Clinical Isolates during Infection of a Nematode

Toxigenic Corynebacterium diphtheriae strains cause diphtheria in humans. The toxigenic C. diphtheriae isolate NCTC13129 produces three distinct heterotrimeric pili that contain SpaA, SpaD, and SpaH, making up the shaft structure. The SpaA pili are known to mediate bacterial adherence to pharyngeal epithelial cells. However, to date little is known about the expression of different pili in various clinical isolates and their importance in bacterial pathogenesis. Here, we characterized a large collection of C. diphtheriae clinical isolates for their pilin gene pool by PCR and for the expression of the respective pilins by immunoblotting with antibodies against Spa pilins. Consistent with the role of a virulence factor, the SpaA-type pili were found to be prevalent among the isolates, and most significantly, corynebacterial adherence to pharyngeal epithelial cells was strictly correlated with isolates that were positive for the SpaA pili. By comparison, the isolates were heterogeneous for the presence of SpaD- and SpaH-type pili. Importantly, using Caenorhabditis elegans as a model host for infection, we show here that strain NCTC13129 rapidly killed the nematodes, the phenotype similar to isolates that were positive for toxin and all pilus types. In contrast, isogenic mutants of NCTC13129 lacking SpaA-type pili or devoid of toxin and SpaA pili exhibited delayed killing of nematodes with similar kinetics. Consistently, nontoxigenic or toxigenic isolates that lack one, two, or all three pilus types were also attenuated in virulence. This work signifies the important role of pili in corynebacterial pathogenesis and provides a simple host model to identify additional virulence factors.     

The Helicobacter pylori Protein CagM is Located in the Transmembrane Channel That is Required for CagA Translocation

Helicobacter pylori (H. pylori) is a human gastric pathogen that colonizes the stomach in more than 50 % of the world’s human population. Infection with this bacterium can induce several gastric diseases ranging from gastritis to peptic ulcer and gastric cancer. Virulent H. pylori isolates harboring the cag pathogenicity island (cag PAI), which encodes a Type IV Secretion System (T4SS), form a pilus for the injection of its major virulence protein CagA into gastric cells. Several cag PAI genes have been identified as homologues of T4SS genes from Agrobacterium tumefaciens, while the other members in cag PAI still have no known function. We studied one of such proteins with unknown function, CagM, which was predicted to have a putative N-terminal signal sequence and at least three transmembrane helices. To determine the subcellular localization of CagM, we performed a cell fractionation procedure and produced rabbit anti-CagM polyclonal antibodies for immunoblotting assays. Furthermore, we generated an isogenic ΔcagM mutant to investigate the ability of CagA translocation compared with the wild-type NCTC 11637 strain using GES-1 and MKN-45 cell infection experiments. Our results indicated that CagM was mainly located in the bacterial membrane, partially located in the periplasm, and essential for CagA translocation both in GES-1 and MKN-45 cells, which suggested that CagM was one of the core members of Cag T4SS and localized in the transmembrane channel.     

Secretion of TcpF by the Vibrio cholerae Toxin-Coregulated Pilus Biogenesis Apparatus Requires an N-Terminal Determinant

Type IV pili are important for microcolony formation, biofilm formation, twitching motility, and attachment. We and others have shown that type IV pili are important for protein secretion across the outer membrane, similar to type II secretion systems. This study explored the relationship between protein secretion and pilus formation in Vibrio cholerae. The toxin-coregulated pilus (TCP), a type IV pilus required for V. cholerae pathogenesis, is necessary for the secretion of the colonization factor TcpF (T. J. Kirn, N. Bose, and R. K. Taylor, Mol. Microbiol. 49:81–92, 2003). This phenomenon is not unique to V. cholerae; secreted virulence factors that are dependent on the presence of components of the type IV pilus biogenesis apparatus for secretion have been reported with Dichelobacter nodosus (R. M. Kennan, O. P. Dhungyel, R. J. Whittington, J. R. Egerton, and J. I. Rood, J. Bacteriol. 183:4451–4458, 2001) and Francisella tularensis (A. J. Hager et al., Mol. Microbiol. 62:227–237, 2006). Using site-directed mutagenesis, we demonstrated that the secretion of TcpF is dependent on the presence of selected amino acid R groups at position five. We were unable to find other secretion determinants, suggesting that Y5 is the major secretion determinant within TcpF. We also report that proteins secreted in a type IV pilus biogenesis apparatus-dependent manner have a YXS motif within the first 15 amino acids following the Sec cleavage site. The YXS motif is not present in proteins secreted by type II secretion systems, indicating that this is unique to type IV pilus-mediated secretion. Moreover, we show that TcpF interacts with the pilin TcpA, suggesting that these proteins are secreted by the type IV pilus biogenesis system. These data provide a starting point for understanding how type IV pili can mediate secretion of virulence factors important for bacterial pathogenesis.     

Dual Role for Pilus in Adherence to Epithelial Cells and Biofilm Formation in Streptococcus agalactiae

Streptococcus agalactiae is a common human commensal and a major life-threatening pathogen in neonates. Adherence to host epithelial cells is the first critical step of the infectious process. Pili have been observed on the surface of several gram-positive bacteria including S. agalactiae. We previously characterized the pilus-encoding operon gbs1479-1474 in strain NEM316. This pilus is composed of three structural subunit proteins: Gbs1478 (PilA), Gbs1477 (PilB), and Gbs1474 (PilC), and its assembly involves two class C sortases (SrtC3 and SrtC4). PilB, the bona fide pilin, is the major component; PilA, the pilus associated adhesin, and PilC, are both accessory proteins incorporated into the pilus backbone. We first addressed the role of the housekeeping sortase A in pilus biogenesis and showed that it is essential for the covalent anchoring of the pilus fiber to the peptidoglycan. We next aimed at understanding the role of the pilus fiber in bacterial adherence and at resolving the paradox of an adhesive but dispensable pilus. Combining immunoblotting and electron microscopy analyses, we showed that the PilB fiber is essential for efficient PilA display on the surface of the capsulated strain NEM316. We then demonstrated that pilus integrity becomes critical for adherence to respiratory epithelial cells under flow-conditions mimicking an in vivo situation and revealing the limitations of the commonly used static adherence model. Interestingly, PilA exhibits a von Willebrand adhesion domain (VWA) found in many extracellular eucaryotic proteins. We show here that the VWA domain of PilA is essential for its adhesive function, demonstrating for the first time the functionality of a prokaryotic VWA homolog. Furthermore, the auto aggregative phenotype of NEM316 observed in standing liquid culture was strongly reduced in all three individual pilus mutants. S. agalactiae strain NEM316 was able to form biofilm in microtiter plate and, strikingly, the PilA and PilB mutants were strongly impaired in biofilm formation. Surprisingly, the VWA domain involved in adherence to epithelial cells was not required for biofilm formation.     

Processed VirB2 Is the Major Subunit of the Promiscuous Pilus of Agrobacterium tumefaciens

Previous studies have implicated the obligatory requirement for the vir regulon (or “virulon”) of the Ti plasmid for the transfer of oncogenes from Agrobacterium tumefaciens to plant cells. The machinery used in this horizontal gene transfer has been long thought to be a transformation or conjugative delivery system. Based on recent protein sequence comparisons, the proteins encoded by the virB operon are strikingly similar to proteins involved in the synthesis and assembly of conjugative pili such as the conjugative pilus of F plasmid in Escherichia coli. The F pilus is composed of TraA pilin subunits derived from TraA propilin. In the present study, evidence is provided showing that the counterpart of TraA is VirB2, which like TraA propilin is processed into a 7.2-kDa product that comprises the pilus subunit as demonstrated by biochemical and electron microscopic analyses. The processed VirB2 protein is present exocellularly on medium on which induced A. tumefaciens had grown and appears as thin filaments of 10 nm that react specifically to VirB2 antibody. Exocellular VirB2 is produced abundantly at 19°C as compared with 28°C, an observation that parallels the effect of low temperature on the production of vir gene-specific pili observed previously (K. J. Fullner, L. C. Lara, and E. W. Nester, Science 273:1107–1109, 1996). Export of the processed VirB2 requires other virB genes since mutations in these genes cause the loss of VirB2 pilus formation and result in processed VirB2 accumulation in the cell. The presence of exocellular processed VirB2 is directly correlated with the formation of pili, and it appears as the major protein in the purified pilus preparation. The evidence provides a compelling argument for VirB2 as the propilin whose 7.2-kDa processed product is the pilin subunit of the promiscuous conjugative pilus, hereafter called the “T pilus” of A. tumefaciens.     

Dimeric hydroanthracenes from the unripe seeds of Cassia torosa

From the unripe seeds of Cassia torosa three new dimeric hydroanthracene derivatives were isolated along with stigmasterol, sitosterol, campesterol, physcion-9-anthrone, torosachyrsone and the phlegmacins A₂ and B₂. The structures of the new derivatives were established as physcion-10, 10′-bianthrone, anhydrophlegmacin B₂ [2-(6′-methoxy-3′-methyl-3′, 8′, 9′-trihydroxy-1′-oxo-1′, 2′, 3′, 4′-tetrahydroanthracene-10′-yl)-1, 8-dihydroxy-3-methoxy-6-methyl-9-oxo-9, 10-dihydroanthracene] and torosanin [2-(6′-methoxy-3′-methyl-3′, 8′,9′-trihydroxy-1′-oxo-1′, 2′, 3′,4′-tetrahydroanthracene-5′-yl)-1, 8-dihydroxy-3-methoxy-6-methyl-9-oxo-9, 10-dihydroanthracene], respectively.     

An 18 kDa Scaffold Protein Is Critical for Staphylococcus epidermidis Biofilm Formation

Virulence of the nosocomial pathogen Staphylococcus epidermidis is crucially linked to formation of adherent biofilms on artificial surfaces. Biofilm assembly is significantly fostered by production of a bacteria derived extracellular matrix. However, the matrix composition, spatial organization, and relevance of specific molecular interactions for integration of bacterial cells into the multilayered biofilm community are not fully understood. Here we report on the function of novel 18 kDa Small basic protein (Sbp) that was isolated from S. epidermidis biofilm matrix preparations by an affinity chromatographic approach. Sbp accumulates within the biofilm matrix, being preferentially deposited at the biofilm–substratum interface. Analysis of Sbp-negative S. epidermidis mutants demonstrated the importance of Sbp for sustained colonization of abiotic surfaces, but also epithelial cells. In addition, Sbp promotes assembly of S. epidermidis cell aggregates and establishment of multilayered biofilms by influencing polysaccharide intercellular-adhesin (PIA) and accumulation associated protein (Aap) mediated intercellular aggregation. While inactivation of Sbp indirectly resulted in reduced PIA-synthesis and biofilm formation, Sbp serves as an essential ligand during Aap domain-B mediated biofilm accumulation. Our data support the conclusion that Sbp serves as an S. epidermidis biofilm scaffold protein that significantly contributes to key steps of surface colonization. Sbp-negative S. epidermidis mutants showed no attenuated virulence in a mouse catheter infection model. Nevertheless, the high prevalence of sbp in commensal and invasive S. epidermidis populations suggests that Sbp plays a significant role as a co-factor during both multi-factorial commensal colonization and infection of artificial surfaces.     

Structural Basis of Pilus Anchoring by the Ancillary Pilin RrgC of Streptococcus pneumoniae

Pili are surface-attached, fibrous virulence factors that play key roles in the pathogenesis process of a number of bacterial agents. Streptococcus pneumoniae is a causative agent of pneumonia and meningitis, and the appearance of drug-resistance organisms has made its treatment challenging, especially in developing countries. Pneumococcus-expressed pili are composed of three structural proteins: RrgB, which forms the polymerized backbone, RrgA, the tip-associated adhesin, and RrgC, which presumably associates the pilus with the bacterial cell wall. Despite the fact that the structures of both RrgA and RrgB were known previously, structural information for RrgC was still lacking, impeding the analysis of a complete model of pilus architecture. Here, we report the structure of RrgC to 1.85 Å and reveal that it is a three-domain molecule stabilized by two intradomain isopeptide bonds. RrgC does not depend on pilus-specific sortases to become attached to the cell wall; instead, it binds the preformed pilus to the peptidoglycan by employing the catalytic activity of SrtA. A comprehensive model of the type 1 pilus from S. pneumoniae is also presented.     

Lipophilic C-methylflavonoids with no B-ring oxygenation in Metrosideros species (Myrtaceae)

Five unusual C-methylflavonoids lacking B-ring oxygenation (2′,4′-dihydroxy-3′,5′-dimethyl-6′-methoxychalcone, 2′,4′-dihydroxy-3′-methyl-6′-methoxychalcone, 2′,6′-dihydroxy-3′-methyl-4′-methoxychalcone, 2′-hydroxy-3′-methyl-4′,6′-dimethoxychalcone and 5,7-dihydroxy-6,8-dimethylflavanone) were found for the first time in Metrosideros excelsa. The flavanone was the major constituent in leaves, whereas 2′,6′-dihydroxy-3′-methyl-4′-methoxychalcone dominated all other aerial plant parts studied. Other Metrosideros species were investigated for these five flavonoids. C₁₉–C₃₆ aldehydes and C₂₂–C₃₂ alcohols were also identified from the dried seed capsules of M. excelsa.     

Quantitative estimates of steric effects: Intramolecular strain energy effects on the activation energy for aquation of some trans-dichlorotetraaminecobalt(III) complexes

In an effort to quantitatively estimate steric contributions to the aquation rates of a series of structurally related cobalt(III) tetraamine complexes, strain energy minimization calculations have been performed on the reactant and some plausible transition state structures. Free energies of activation ΔG*_obs, are factored as: ΔG*_obs, = ΔG*_bb + ΔG*_strain + Δ_G*_CF + ΔG*_solvation + … where ΔG*_bb is the free energy change associated with bond breaking, ΔG*_solvation is the solvation free energy difference between the reactant and a proposed transition stare, ΔG*_CF is the difference in crystal field stabilization between the reactant and a proposed transition state, and ΔG*_strain is the strain energy difference between the reactant complex and a proposed transition state. The activation energy for the aquation of a hypothetical ‘strain free’ complex is defined as ΔG*_int and reflects the energy required for the bond breaking step with all other terms. For the cations trans-(RR,SS)-dichloro-1,8- diamino-3,6-diazaoctanecobalt(III)(trans [Co(2,2,2- tet)Cl₂]⁺), trans-(RR,SS)- or trans-(RS)-dichloro-1.9- diamino-3,7-diazanonanecobalt(III)(trans [Co(2,3,2- tet)Cl₂]⁺ and trans-(RS)-dichloro-1,10-diamino-4,7- diazadecanecobalt(III)(trans[Co(3,2,3-tet)Cl₂]⁺) ΔG*_int is found to be a constant 123 kJ/mol. For the trans-dichlorocobalt(III) complexes with the ligands 1,4,7,10-tetraazacyclotridecane([13]-ane-N₄), 1,4,8, 11-tetraazacyclotetradecane([14]-ane-N₄), 1,4,8,12- tetraazacyclopentadecane([15]-ane-N₄) and 1,5,9,13- tetraazacyclohexadecane([16]-ane-N₄), ΔG*_int lies in the range 133–139 kJ/mol.     

Dibenzocyclooctadiene lignans from Magnolia and Talauma (Magnoliaceae): Their absolute configuration ascertained by circular dichroism and X-ray crystallography and re-evaluation of previously published pyramidatin structures

Twelve pyramidatins, i.e., dibenzocyclooctadiene-type lignans, together with Machilin G, were isolated from the dichloromethane extracts of aerial material of Talauma gloriensis, Magnolia fraseri, and Magnolia pyramidata (Magnoliaceae). These lignans contain a highly oxidized 7,9′-epoxy-2,2′-cyclolignane skeleton. Their structures were established using NMR spectroscopy (1D and 2D experiments) and mass spectrometry. The absolute configurations of five pairs of atropisomers (S_a/R_a-pyramidatins) and two single atropisomers (S_a-pyramidatins) were determined by experimental and calculated circular dichroism (CD). In addition, the absolute configuration of (S_a)-3,3′,4,4′,5,5′-hexamethoxypyramidatin was confirmed using X-ray crystallography.Five pyramidatins, (R_a)-3,3′,4,4′,5,5′-hexamethoxypyramidatin, (R_a)-3,3′-dimethoxy-4,5:4′,5′-bis(methylenedioxy)pyramidatin, (S_a)-3,3′,4,5′-tetramethoxy-4,5-methylenedioxypyramidatin, (R_a)-3,3′,4,5′-tetramethoxy-4,5-methylenedioxypyramidatin, and (R_a)-3,3′,4,5-tetramethoxy-4′,5′-methylenedioxypyramidatin are reported herein for the first time. In the current dataset, NMR values are in accordance with the observed and calculated CD values. These values are herein reported with particular reference to previously described data of pyramidatins, which have to be revised.     

Functional Identification of Conserved Residues Involved in Lactobacillus rhamnosus Strain GG Sortase Specificity and Pilus Biogenesis

In Gram-positive bacteria, sortase-dependent pili mediate the adhesion of bacteria to host epithelial cells and play a pivotal role in colonization, host signaling, and biofilm formation. Lactobacillus rhamnosus strain GG, a well known probiotic bacterium, also displays on its cell surface mucus-binding pilus structures, along with other LPXTG surface proteins, which are processed by sortases upon specific recognition of a highly conserved LPXTG motif. Bioinformatic analysis of all predicted LPXTG proteins encoded by the L. rhamnosus GG genome revealed a remarkable conservation of glycine residues juxtaposed to the canonical LPXTG motif. Here, we investigated and defined the role of this so-called triple glycine (TG) motif in determining sortase specificity during the pilus assembly and anchoring. Mutagenesis of the TG motif resulted in a lack or an alteration of the L. rhamnosus GG pilus structures, indicating that the TG motif is critical in pilus assembly and that they govern the pilin-specific and housekeeping sortase specificity. This allowed us to propose a regulatory model of the L. rhamnosus GG pilus biogenesis. Remarkably, the TG motif was identified in multiple pilus gene clusters of other Gram-positive bacteria, suggesting that similar signaling mechanisms occur in other, mainly pathogenic, species.     

Identification and characterization of the adenosine 3′,5′-cyclic monophosphate binding proteins appearing during the development of Dictyostelium discoideum

A photosensitive, radioactive analogue of cyclic adenosine monophosphate, 8-azido-adenosine 3′,5′-[³²P]monophosphate (8-N₃-cyclic AMP), was used to label the cyclic AMP binding proteins of Dictyostelium discoideum. During development cytosolic proteins appear which are specifically labeled by the photoaffinity agent. The proteins are developmentally regulated since they are only found in starved, developing cells. Unlabeled cyclic AMP competes specifically with the labeled analogue for protein binding sites in contrast to unlabeled 5′-AMP which does not compete. A mutant which develops spores but is deficient in stalk cell production produces a different set of cyclic AMP binding proteins from the parent strain.     

Contribution of Individual Ebp Pilus Subunits of Enterococcus faecalis OG1RF to Pilus Biogenesis,Biofilm Formation and Urinary Tract Infection

The endocarditis and biofilm-associated pilus (Ebp) operon is a component of the core genome of Enterococcus faecalis that has been shown to be important for biofilm formation, adherence to host fibrinogen, collagen and platelets, and in experimental endocarditis and urinary tract infection models. Here, we created single and double deletion mutants of the pilus subunits and sortases; next, by combining western blotting, immunoelectron microscopy, and using ebpR in trans to increase pilus production, we identified EbpA as the tip pilin and EbpB as anchor at the pilus base, the latter attached to cell wall by the housekeeping sortase, SrtA. We also confirmed EbpC and Bps as the major pilin and pilin-specific sortase, respectively, both required for pilus polymerization. Interestingly, pilus length was increased and the number of pili decreased by deleting ebpA, while control overexpression of ebpA in trans restored wild-type levels, suggesting a dual role for EbpA in both initiation and termination of pilus polymerization. We next investigated the contribution of each pilin subunit to biofilm formation and UTI. Significant reduction in biofilm formation was observed with deletion of ebpA or ebpC (P<0.001) while ebpB was found to be dispensable; a similar result was seen in kidney CFUs in experimental UTI (ΔebpA, ΔebpC, P≤0.0093; ΔebpB, non-significant, each vs. OG1RF). Hence, our data provide important structural and functional information about these ubiquitous E. faecalis pili and, based on their demonstrated importance in biofilm and infection, suggest EbpA and EbpC as potential targets for antibody-based therapeutic approaches.     

Characterization and Proteome Analysis of Inosine 5-Monophosphate Dehydrogenase in Epidemic Streptococcus suis Serotype 2

Streptococcus suis serotype 2 (SS2) is an important zoonotic pathogen that causes severe disease symptoms in pigs and humans. In the present study, we found one isogenic mutant lacking inosine 5-monophosphate dehydrogenase (IMPDH) ΔZY05719 was attenuated in pigs compared with the wild-type SS2 strain ZY05719. Comparative proteome analysis of the secreted proteins expression profiles between ZY05719 and ΔZY05719 allowed us to identify Triosephosphate isomerase (TPI) and glyceraldehyde phosphate dehydrogenase (GAPDH), which were down expressed in the absence of the IMPDH. Both of them are glycolytic enzymes participating in the glycolytic pathway. Compared with ZY05719, ΔZY05719 lost the ability of utilize mannose, which might relate to down expression of TPI and GAPDH. In addition, GAPDH is a well-known factor that involved in adhesion to host cells, and we demonstrated ability of adhesion to HEp-2 and PK15 by ΔZY05719 was significantly weakened, in contrast to ZY05719. The adhesion to host cells is the crucial step to cause infection for pathogen, and the reduction adhesion of ΔZY05719, to some extent illustrates the attenuated virulence of ΔZY05719.     

Isolation and identification of 2-hydroxyplectaniaxanthin from Rhodotorula aurantiaca

A new trihydroxyl carotenoid has been isolated from the yeast Rhodotorula aurantiaca (Saito) Lodder C.B.S. 317 and identified as 2-hydroxyplectaniaxanthin (3′,4′-didehydro,1′,2′-dihydro-β, ψ-caroten-2,1′,2′-triol). Its m.p., partition coefficient, _Rf, extinction coefficient, ms and NMR spectra are reported. Since the hydroxyl group at C-2 of the β-ionone ring is unusual, a possible mechanism for the biosynthesis of this carotenoid has been proposed.