TatC-dependent translocation of pyoverdine is responsible for the microbial growth suppression

Infections are often not caused by a colonization of Pseudomonas aeruginosa alone but by a consortium of other bacteria. Little is known about the impact of P. aeruginosa on the growth of other bacteria upon coinfection. Here, cellree culture supernatants obtained from P. aeruginosa suppressed the growth of a number of bacterial strains such as Corynebacterium glutamicum, Bacillus subtilis, Staphylococcus aureus, and Agrobacterium tumefaciens, but had little effect on the growth of Escherichia coli and Salmonella Typhimurium. The growth suppression effect was obvious when P. aeruginosa was cultivated in M9 minimal media, and the suppression was not due to pyocyanin, a well-known antimicrobial toxin secreted by P. aeruginosa. By performing transposon mutagenesis, PA5070 encoding TatC was identified, and the culture supernatant of its mutant did not suppress the growth. HPLC analysis of supernatants showed that pyoverdine was a secondary metabolite present in culture supernatants of the wild-type strain, but not in those of the PA5070 mutant. Supplementation of FeCl₂ as a source of iron compromised the growth suppression effect of supernatants and also recovered biofilm formation of S. aureus, indicating that pyoverdine-mediated iron acquisition is responsible for the growth suppression. Thus, this study provides the action of TatC-dependent pyoverdine translocation for the growth suppression of other bacteria, and it might aid understanding of the impact of P. aeruginosa in the complex community of bacterial species upon coinfection.     

The Distribution of Prion Protein Allotypes Differs Between Sporadic and Iatrogenic Creutzfeldt-Jakob Disease Patients

Sporadic Creutzfeldt-Jakob disease (sCJD) is the most prevalent of the human prion diseases, which are fatal and transmissible neurodegenerative diseases caused by the infectious prion protein (PrP^Sc). The origin of sCJD is unknown, although the initiating event is thought to be the stochastic misfolding of endogenous prion protein (PrP^C) into infectious PrP^Sc. By contrast, human growth hormone-associated cases of iatrogenic CJD (iCJD) in the United Kingdom (UK) are associated with exposure to an exogenous source of PrP^Sc. In both forms of CJD, heterozygosity at residue 129 for methionine (M) or valine (V) in the prion protein gene may affect disease phenotype, onset and progression. However, the relative contribution of each PrP^C allotype to PrP^Sc in heterozygous cases of CJD is unknown. Using mass spectrometry, we determined that the relative abundance of PrP^Sc with M or V at residue 129 in brain specimens from MV cases of sCJD was highly variable. This result is consistent with PrP^C containing an M or V at residue 129 having a similar propensity to misfold into PrP^Sc thus causing sCJD. By contrast, PrP^Sc with V at residue 129 predominated in the majority of the UK human growth hormone associated iCJD cases, consistent with exposure to infectious PrP^Sc containing V at residue 129. In both types of CJD, the PrP^Sc allotype ratio had no correlation with CJD type, age at clinical onset, or disease duration. Therefore, factors other than PrP^Sc allotype abundance must influence the clinical progression and phenotype of heterozygous cases of CJD.     

Characterization of a bovine pregnancy-associated protein using two-dimensional gel electrophoresis, N-terminal sequencing and mass spectrometry

Bovine pregnancy-associated protein (bPAP) isolated from pregnant bovine urines by two-dimensional electrophoresis (2-DE) was characterized by N-terminal sequencing, internal sequencing, and mass spectrometric analyses using matrix-assisted laser desorption/ionization-time of flight mass spectrometry and tandem mass spectrometry. Database search using the amino acid sequences and the peptide mass profiles showed that the protein is a novel bovine pregnancy-associated protein of which the N-terminus has a high similarity to collagen alpha. The protein has a molecular mass of 21 kDa and a pI of 6.1. The expression profiles of the protein from the urine of 30 pregnant and 20 nonpregnant cows characterized by 2-DE indicated that the expression of bPAP during pregnancy increased to over nmol from the pmol level basal expression of bPAP at the nonpregnant state with < 3% of false negatives and < 10% of false positives. Using the peptide sequence information, polyclonal antibodies against the bPAP protein were generated. The purified polyclonal antibodies against the peptide sequences of bPAP detected the 21 kDa protein on the blots of pregnant cow urine by Western blot analysis. In addition, analysis showed that the expression of bPAP in the urine is associated with pregnancy, but that the urine concentration of bPAP is not correlated with the duration of the pregnancy.     

Geojedonia litorea gen. nov., sp. nov., a member of the family Flavobacteriaceae isolated from coastal seawater

A Gram-negative, non-flagellated, non-gliding and rod-shaped, coccoid or filamentous bacterial strain, designated YCS-16^T, was isolated from coastal seawater from a seaweed farm on the South Sea, South Korea, and its taxonomic position was investigated by using a polyphasic study. Strain YCS-16^T was observed to grow optimally at 30 °C, at pH 7.0–7.5 and in the presence of 2 % (w/v) NaCl. Strain YCS-16^T exhibited the highest 16S rRNA gene sequence similarity values to the type strains of Bizionia echini (96.1 %), Formosa spongicola (95.8 %), Bizionia algoritergicola (95.5 %) and Psychroserpens mesophilus (95.4 %). Neighbour-joining and maximum-parsimony phylogenetic trees based on 16S rRNA gene sequences revealed that strain YCS-16^T joined the cluster comprising the type strains of Psychroserpens species. Strain YCS-16^T was found to contain MK-6 as the predominant menaquinone and iso-C_17:0 3-OH, iso-C_15:1 G and iso-C_15:0 as the major fatty acids. The major polar lipids detected in strain YCS-16^T were phosphatidylethanolamine and one unidentified lipid. The DNA G+C content of strain YCS-16^T was determined to be 35.7 mol%. The phylogenetic analysis, chemotaxonomic data and other phenotypic properties revealed that strain YCS-16^T constitutes a new genus and species within the family Flavobacteriaceae, for which the name Geojedonia litorea gen. nov., sp. nov. is proposed. The type strain of Geojedonia litorea is YCS-16^T (=KCTC 32260^T = CCUG 63682^T).     

Akt/PKB kinase phosphorylates separately Thr212 and Ser214 of tau protein in vitro

Microtubule-associated protein tau contains a consensus motif for protein kinase B/Akt (Akt), which plays an essential role in anti-apoptotic signaling. The motif encompasses the AT100 double phospho-epitope (Thr212/Ser214), a specific marker for Alzheimer's disease (AD) and other neurodegenerations, raising the possibility that it could be generated by Akt. We studied Akt-dependent phosphorylation of tau protein in vitro. We found that Akt phosphorylated both Thr212 and Ser214 in the longest and shortest tau isoforms as determined using phospho site-specific antibodies against tau. Akt did not phosphorylate other tau epitopes, including Tau-1, AT8, AT180, 12E8 and PHF-1. The Akt-phosphorylated tau retained its initial electrophoretic mobility. Immunoprecipitation studies with phospho-specific Thr212 and Ser214 antibodies revealed that only one of the two sites is phosphorylated per single tau molecule, resulting in tau immunonegative for AT100. Mixed kinase studies showed that prior Ser214 phosphorylation by Akt blocked protein kinase A but not GSK3beta activity. On the other hand, GSK3beta selectively blocked Ser214 phosphorylation, which was prevented by lithium. The results suggest that Akt may be involved in AD-specific phosphorylation of tau at the AT100 epitope in conjunction with other kinases. Our data suggest that phosphorylation of tau by Akt may play specific role(s) in Akt-mediated anti-apoptotic signaling, particularly relevant to AD and other neurodegenerations.     

Simultaneous degradation of p-nitrophenol and phenol by a newly isolated Nocardioides sp

A p-nitrophenol (PNP)- and phenol-mineralizing bacterium (strain NSP41) was isolated from an industrial wastewater and identified as a member of the genus Nocardioides. PNP was degraded via a hydroquinone pathway, and phenol was degraded through a catechol pathway in strain NSP41. Both enzyme systems for the degradation of PNP and phenol were induced simultaneously in the presence of both compounds. Although both enzyme systems were induced at the same time, PNP and phenol were degraded by the hydroquinone and catechol pathway, respectively. However, during the simultaneous degradation in the low phenol concentration, after the exhaustion of phenol, some PNP was transformed by the catechol pathway and 4-nitrocatechol was transiently accumulated. Kinetically, the addition of phenol greatly enhanced the apparent PNP degradation rate, which may be due to the increased cell mass by the assimilation of phenol.     

Rapid discrimination of CMS cybrid lines between <Emphasis Type="Italic">Brassica oleracea</Emphasis> var. <Emphasis Type="Italic">capitata</Emphasis> and <Emphasis Type="Italic">Raphanus sativus</Emphasis> L. using Fourier transform infrared (FT-IR) spectroscopy of genomic DNA

The purpose of this study is to establish the rapid discrimination system of cybrid callus lines by Fourier transform infrared (FT-IR) spectroscopy without genetic fingerprinting analysis. Genomic DNA isolated from two parental lines (Brassica oleracea var. capitata and Raphanus sativus L.) and their cybrid callus lines were analyzed by FT-IR spectroscopy in the spectral region from 4000 to 400 cm^?1. Several spectral differences between the two parental lines were detected in the frequency regions of N–H stretching (amide I), C=O stretching vibrations (amide II), and PO₂^? ionized asymmetric and symmetric stretching. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were used to discriminate cybrid cell lines from the two parental species at the genomic DNA level. PLS-DA analysis provided more clear discrimination between the two parental lines and their progeny cybrid lines in the score plot. PCA loading values also showed that obvious spectral differences played a significant role in discrimination between the two parental lines and their cybrid lines. These spectral differences might be directly related to subtle changes in the base functional groups and backbone structures of genomic DNA. Considering these results, this technique could provide a research foundation for the FT-IR spectral-based diagnosis, selection, and discrimination of parental lines and their cybrids. Furthermore, this technique could be applied in the hybrid seed industry for rapid screening with high heterogeneity.