Phytoplasmas are cell wall-less bacteria inhabiting the phloem and utilizing it for their spread. Infected plants often show changes in growth pattern and a reduced crop yield. A quantitative real-time polymerase chain reaction (Q-PCR) assay and a bioimaging method were developed to quantify and localize phytoplasmas in situ. According to the Q-PCR assay, phytoplasmas accumulated disproportionately in source leaves of Euphorbia pulcherrima and, to a lesser extent, in petioles of source leaves and in stems. However, phytoplasma accumulation was small or nondetectable in sink organs (roots and sink leaves). For bioimaging, infected plant tissue was stained with vital fluorescence dyes and examined using confocal laser scanning microscopy. With a DNA-sensitive dye, the pathogens were detected exclusively in the phloem, where they formed dense masses in sieve tubes of Catharanthus roseus. Sieve tubes were identified by counterstaining with aniline blue for callose and multiphoton excitation. With a potentiometric dye, not all DNA-positive material was stained, suggesting that the dye stained metabolically active phytoplasmas only. Some highly infected sieve tubes contained phytoplasmas that were either inactive or dead upon staining.

Roseobacter strain 27-4 has been isolated from a turbot larval rearing unit and is capable of reducing mortality in turbot egg yolk sac larvae. Here, we demonstrate that the supernatant of Roseobacter 27-4 is lethal to the larval pathogens Vibrio anguillarum and Vibrio splendidus in a buffer system and inhibited their growth in marine broth. Liquid chromatography (LC) with both UV spectral detection and high-resolution mass spectrometry (HR-MS) identified the known antibacterial compound thiotropocin or its closely related precursor tropodithietic acid in the bioactive fractions. Antibacterial activity correlated with the appearance of a brownish pigment and was only formed in marine broth under static growth conditions. A thick biofilm of multicellular star-shaped aggregated cells formed at the air-liquid interface under static growth conditions. Here, the bioactive compound was the base peak in the LC-UV chromatograms of the extracts where it constituted 15% of the total peak area. Aerated conditions results in 10-fold-higher cell yield, however, cultures were nonpigmented, did not produce antibacterial activity, and grew as single cells. Production of antibacterial compounds may be quorum regulated, and we identified the acylated homoserine lactone (3-hydroxy-decanoyl homoserine lactone) from cultures of Roseobacter 27-4 using LC-HR-MS. The signal molecule was primarily detected in stagnant cultures. Roseobacter 27-4 grew between 10 and 30°C but died rapidly at 37°C. Also, the antibacterial compounds was sensitive to heat and was inactivated at 37°C in less than 2 days and at 25°C in 8 days. Using Roseobacter 27-4 as a probiotic culture will require that is be established in stagnant or adhered conditions and, due to the temperature sensitivity of the active compound, constant production must be ensured.The dramatic growth in the aquaculture sector (an increment of 9 to 10% per year over the last 8 to 10 years) has emphasized the importance of fish disease control. Bacterial diseases are important constraints and may be treated with antibiotics. However, due to the risk of development and transfer of antibiotic resistance, alternative disease control measures must be implemented. Vaccines have been very successful also in fish farming; however, vaccines are not efficient at the larval stages. Several studies have demonstrated that probiotic bacteria may be used to control pathogenic organisms in fish larval rearing (17,18,32), and one of several promising candidates are bacteria of the marine Roseobacter clade.The ability of Roseobacter to inhibit other bacteria was noted already by who used Roseobacter gallaeciensis strain BS107 as a probiotic treatment of scallop larvae (32). Roseobacter strains have also been isolated from turbot larval farms, and they were selected from this environment due to their strong anti-Vibrio activity (18). These strains appeared to constitute a relatively stable community since the same DNA subtypes were isolated over several months (19). Antagonistic Roseobacter were especiall...

Serum YKL-40 varies according to disease activity in RA, but provides in some respect information different from conventional markers. Our previous studies are consistent with a local release of YKL-40 in the arthritic joint followed by a secondary increase in serum YKL-40. YKL-40 may prove to be a new tool for the study of disease activity and pathophysiology of RA.

Pathogens induce the expression of many genes encoding plant transcription factors, though specific knowledge of the biological function of individual transcription factors remains scarce. NAC transcription factors are encoded in plants by a gene family with proposed functions in both abiotic and biotic stress adaptation, as well as in developmental processes. In this paper, we provide convincing evidence that a barley NAC transcription factor has a direct role in regulating basal defence. The gene transcript was isolated by differential display from barley leaves infected with the biotrophic powdery mildew fungus, Blumeria graminis f.sp. hordei (Bgh). The full-length cDNA clone was obtained using 5'-RACE and termed HvNAC6, due to its high similarity to the rice homologue, OsNAC6. Gene silencing of HvNAC6 during Bgh inoculation compromises penetration resistance in barley epidermal cells towards virulent Bgh. Complementing the effect of HvNAC6 gene silencing, transient overexpression of HvNAC6 increases the occurrence of penetration resistant cells towards Bgh attack. Quantitative RT-PCR shows the early and transient induction of HvNAC6 in barley epidermis upon Bgh infection. Additionally, our results show that the Arabidopsis HvNAC6 homologue ATAF1 is also induced by Bgh and the ataf1-1 mutant line shows decreased penetration resistance to this non-host pathogen. Collectively, these data suggest a conserved role of HvNAC6 and ATAF1 in the regulation of penetration resistance in monocots and dicots, respectively.

Mechanisms controlling microbial degradation of dissolved organic matter (DOM) in aquatic environments are poorly understood, although microbes are crucial to global nutrient cycling. Bacterial cell wall components may be one of the keys in understanding the presence of slowly degrading DOM in nature. We found that dominant components of bacterial cell walls (D-amino acids (D-AA), glucosamine (GluA) and diaminopimelic acid (DAPA)) comprised up to 11.4% of the dissolved organic nitrogen in 50 diverse rivers entering the Baltic Sea. Occurrence of DAPA, a characteristic component of Gram-negative (G(-)) bacteria, in the rivers suggests that G(-) bacteria rather than Gram-positive (G(+)) were the major source of the cell wall material. In laboratory studies, the degradation of whole bacterial cells, cell wall material and purified peptidoglycan was studied to characterize degradation of cell wall material by natural aquatic bacteria. Addition of whole killed G(-) and G(+) bacteria to cultures of estuarine bacteria demonstrated fragmentation and loss of cell structure of the G(+) bacteria, while the G(-) bacteria maintained an intact cell shape during the entire 69-day period. In another experiment, estuarine bacteria degraded 39-69% of GluA, D-AA and DAPA in added cell wall material of a representative G(-) bacterial species during 8 days, as compared to a 72-89% degradation of GluA, D-AA and DAPA in cell material of a G(+) bacterial species. When cultures of estuarine bacteria were enriched with purified G(-) and G(+) peptidoglycan (1 mg l(-1)), at least 49% (G(-)) and 58% (G(+)) of D-AA in the peptidoglycan was degraded. No major changes in GluA were obvious. Interpretation of the results was difficult as a portion of the purified peptidoglycan was of similar size to the bacteria and could not be differentiated from cells growing in the cultures. Addition of the purified peptidoglycan stimulated the bacterial growth, and after 6 days the cell density in the enriched cultures was 4-fold higher than in the controls. A regrowth of bacteria after addition of L-broth at 105 days caused a 50- to 75-fold increase in dissolved D-AA and GluA. Most of the D-AA and GluA were taken up during the following 10 days, indicating that cell wall constituents are dynamic compounds. Our results show that a variable portion of peptidoglycan in G(-) and G(+) bacteria can be degraded by natural bacteria, and that peptidoglycan in G(-) bacteria is more resistant to bacterial attack than that in G(+) bacteria. Thus, the presence of cell wall constituents in natural DOM may reflect the recalcitrant nature of especially G(-) peptidoglycan.

In recent years, aquaporin biomimetic membranes (ABMs) for water separation have gained considerable interest. Although the first ABMs are commercially available, there are still many challenges associated with further ABM development. Here, we discuss the interplay of the main components of ABMs: aquaporin proteins (AQPs), block copolymers for AQP reconstitution, and polymer-based supporting structures. First, we briefly cover challenges and review recent developments in understanding the interplay between AQP and block copolymers. Second, we review some experimental characterization methods for investigating AQP incorporation including freeze-fracture transmission electron microscopy, fluorescence correlation spectroscopy, stopped-flow light scattering, and small-angle X-ray scattering. Third, we focus on recent efforts in embedding reconstituted AQPs in membrane designs that are based on conventional thin film interfacial polymerization techniques. Finally, we describe some new developments in interfacial polymerization using polyhedral oligomeric silsesquioxane cages for increasing the physical and chemical durability of thin film composite membranes.

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A fluorescence in situ hybridization (FISH) technique based on binding of a rhodamine-labelled oligonucleotide probe to 16S rRNA was used to estimate the numbers of ribosome-rich bacteria in soil samples. Such bacteria, which have high cellular rRNA contents, were assumed to be active (and growing) in the soil. Hybridization to an rRNA probe, EUB338, for the domain Bacteria was performed with a soil slurry, and this was followed by collection of the bacteria by membrane filtration (pore size, 0.2 μm). A nonsense probe, NONEUB338 (which has a nucleotide sequence complementary to the nucleotide sequence of probe EUB338), was used as a control for nonspecific staining. Counting and size classification into groups of small, medium, and large bacteria were performed by fluorescence microscopy. To compensate for a difference in the relative staining intensities of the probes and for binding by the rhodamine part of the probe, control experiments in which excess unlabelled probe was added were performed. This resulted in lower counts with EUB338 but not with NONEUB338, indicating that nonspecific staining was due to binding of rhodamine to the bacteria. A value of 4.8 × 108 active bacteria per g of dry soil was obtained for bulk soil incubated for 2 days with 0.3% glucose. In comparison, a value of 3.8 × 108 active bacteria per g of dry soil was obtained for soil which had been air dried and subsequently rewetted. In both soils, the majority (68 to 77%) of actively growing bacteria were members of the smallest size class (cell width, 0.25 to 0.5 μm), but the active (and growing) bacteria still represented only approximately 5% of the total bacterial population determined by DAPI (4′,6-diamidino-2-phenylindole) staining. The FISH technique in which slurry hybridization is used holds great promise for use with phylogenetic probes and for automatic counting of soil bacteria.