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An antifungal active fraction (AAF) from the coelomic fluid (CF) of the earthworm Dendrobaena veneta was isolated. The aim of the study was to analyze the antifungal activity of the AAF and to carry out chemical characterization of the fraction. The active fraction showed antifungal activity against a clinical C. albicans isolate, C. albicans ATCC 10231, and C. krusei ATCC 6258. It effectively reduced the metabolic activity of C. albicans cells and influenced their morphology after 48 hours of incubation. Scanning electron microscopy (SEM) images revealed loss of integrity of the cell wall induced by the active fraction. Calcofluor White staining showed changes in the structure of the C. albicans cell wall induced by the AAF. The fungal cells died via apoptosis and necrosis after the treatment with the studied fraction. Electrophoresis under native conditions revealed the presence of two compounds in the AAF, while SDS/PAGE gel electrophoresis showed several protein and carbohydrate compounds. The active fraction was analyzed using Raman spectroscopy, MALDI TOF/TOF, and ESI LC-MS. The Raman analysis confirmed the presence of proteins and determined their secondary structure. The MALDI TOF/TOF analysis facilitated detection of four main compounds with a mass of 7694.9 m/z, 12292.3 m/z, 21628.3 m/z, and 42923.2 m/z in the analyzed fraction. The presence of carbohydrate compounds in the preparation was confirmed by nuclear magnetic resonance (NMR) and gas chromatography (GC-MS). The ATR-FTIR spectrum of the AAF exhibited high similarity to the spectrum of egg white lysozyme. The AAF showed no endotoxicity and cytotoxicity towards normal skin fibroblasts (HSF); therefore, it can be used for the treatment of skin and mucous membrane candidiasis in the future. Given its efficient and selective action, the fraction seems to be a promising preparation with antifungal activity against C. albicans. [ABSTRACT FROM AUTHOR]

Ochrobactrum spp. are ubiquitous bacteria attracting growing attention as important members of microbiomes of plants and nematodes and as a source of enzymes for biotechnology. Strain Ochrobactrum sp. A44T was isolated from the rhizosphere of a field-grown potato in Gelderland, the Netherlands. The strain can interfere with quorum sensing (QS) of Gram-negative bacteria through inactivation of N-acyl homoserine lactones (AHLs) and protect plant tissue against soft rot pathogens, the virulence of which is governed by QS. Phylogenetic analysis based on 16S rRNA gene alone and concatenation of 16S rRNA gene and MLSA genes (groEL and gyrB) revealed that the closest relatives of A44T are O. grignonense OgA9aT, O. thiophenivorans DSM 7216T, O. pseudogrignonense CCUG 30717T, O. pituitosum CCUG 50899T, and O. rhizosphaerae PR17T. Genomes of all six type strains were sequenced, significantly expanding the possibility of genome-based analyses in Ochrobactrum spp. Average nucleotide identity (ANIb) and genome-to-genome distance (GGDC) values for A44T and the related strains were below the single species thresholds (95% and 70%, respectively), with the highest scores obtained for O. pituitosum CCUG 50899T (87.31%; 35.6%), O. rhizosphaerae PR17T (86.80%; 34.3%), and O. grignonense OgA9aT (86.30%; 33.6%). Distinction of A44T from the related type strains was supported by chemotaxonomic and biochemical analyses. Comparative genomics revealed that the core genome for the newly sequenced strains comprises 2731 genes, constituting 50–66% of each individual genome. Through phenotype-to-genotype study, we found that the non-motile strain O. thiophenivorans DSM 7216T lacks a cluster of genes related to flagella formation. Moreover, we explored the genetic background of distinct urease activity among the strains. Here, we propose to establish a novel species Ochrobactrum quorumnocens, with A44T as the type strain (= LMG 30544T = PCM 2957T). [ABSTRACT FROM AUTHOR]

In a typing system based on opsonic antibodies against carbohydrate antigens of the cell envelope, 60% of Enterococcus faecalis strains can be assigned to one of four serotypes (CPS-A to CPS-D). The structural basis for enterococcal serotypes, however, is still incompletely understood. Here we demonstrate that antibodies raised against lipoteichoic acid (LTA) from a CPS-A strain are opsonic to both CPS-A and CPS-B strains. LTA-specific antibodies also bind to LTA of CPS-C and CPS-D strains, but fail to opsonize them. From CPS-C and CPS-D strains resistant to opsonization by anti-LTA, we purified a novel diheteroglycan with a repeating unit of →6)-β-Galf-(1→3)- β-D-Glcp-(1→ with O-acetylation in position 5 and lactic acid substitution at position 3 of the Galf residue. The purified diheteroglycan, but not LTA absorbed opsonic antibodies from whole cell antiserum against E. faecalis type 2 (a CPS-C strain) and type 5 (CPS-D). Rabbit antiserum raised against purified diheteroglycan opsonized CPS-C and CPS-D strains and passive protection with diheteroglycan-specific antiserum reduced bacterial counts by 1.4 - 3.4 logs in mice infected with E. faecalis strains of the CPS-C and CPS-D serotype. Diheteroglycanspecific opsonic antibodies were absorbed by whole bacterial cells of E. faecalis FA2-2 (CPS-C) but not by its isogenic acapsular cpsI-mutant and on native PAGE purified diheteroglycan co-migrated with the gene product of the cps-locus, suggesting that it is synthesized by this locus. In summary, two polysaccharide antigens, LTA and a novel diheteroglycan, are targets of opsonic antibodies against typeable E. faecalis strains. These cell-wall associated polymers are promising candidates for active and passive vaccination and add to our armamentarium to fight this important nosocomial pathogen. [ABSTRACT FROM AUTHOR]