Evidence for plasmid-associated crystal toxin production in Bacillus thuringiensis subsp. israelensis

Three crystalliferous (Cry⁺) strains of Bacillus thuringiensis subsp. israelensis (serotype 14) that produce parasporal protein crystals toxic to dipteran larvae and several acrystalliferous (Cry^?) mutants, either induced or spontaneously derived from a single Cry⁺ parent, were examined for the presence of covalently closed circular (CCC) DNA in attempts to correlate toxin production with the presence of a specific plasmid. The plasmid profiles of both Cry⁺ and Cry^? variants were analyzed by both a cleared lysate- and a modified Eckhardt lysateelectrophoresis technique. All of the Cry^? mutants derived from the Cry⁺ parental strain had lost a 4.0- to 4.4-megadalton (Mdal) plasmid. Bioassay data confirmed loss of toxin production by the Cry^? variants. All three Cry⁺ strains, including the parent of the Cry^? strains, contained CCC plasmids DNAs of the following approximate molecular weights: 4.0 to 4.4, 5.2 to 6.0, and 11.4 to 13.0 Mdal. One Cry⁺ strain contained an additional CCC plasmid of 6.7 to 7.2 Mdal. The plasmid patterns for several Cry^? derivatives differed in other respects from the pattern for their parent strain. The various Cry⁺ and Cry^? strains could be distinguished either by phenotypical differences in antibiotic sensitivity, crystal production, and toxicity, or by differences in their plasmid profiles.     

Purification and some properties of Α-amylase from an ectomycorrhizal fungus, <Emphasis Type="Italic">Tricholoma matsutake</Emphasis>

Agr" align="BASELINE" BORDER="0">-Amylase from a still culture filtrate of Tricholoma matsutake, an ectomycorrhizal fungus, was isolated and characterized. The enzyme was purified to a homogeneous preparation with Toyopearl-DEAE, gel filtration, and Mono Q column chromatography. The Agr" align="BASELINE" BORDER="0">-amylase was highly purified (3580 fold) with a recovery of 10.5% and showed a single protein band by SDS-PAGE. The enzyme was most active at pH 5.0–6.0 toward soluble starch and stable within the broad pH range 4.0–10.0. This Agr" align="BASELINE" BORDER="0">-amylase was a relatively thermostable enzyme (optimum temperature, 60°C; thermal stability, 50°C). The molecular mass was 34kDa by size-exclusion chromatography and 46kDa by SDS-PAGE. This enzyme was not inhibited by the Hg²⁺ ion. Measurement of viscosity and TLC and HPLC analysis of the hydrolysates obtained from amylose showed that the amylase from T. matsutake is an endo-type (Agr" align="BASELINE" BORDER="0">-amylase). Substrate specificity was tested using amylose with different polysaccharides. This Agr" align="BASELINE" BORDER="0">-amylase readily hydrolyzed the Agr" align="BASELINE" BORDER="0">-1,4 glucoside bond in soluble starch and amylose A (MW, 2900), but did not hydrolyze the Agr" align="BASELINE" BORDER="0">-1,6 bond and cyclic polysaccharides such as Agr" align="BASELINE" BORDER="0">- and -cyclodextrin.     

反义锁核酸阻断耐甲氧西林金黄色葡萄球菌agr群体感应系统

目的:利用反义策略,设计、筛选针对agrA mRNA的反义锁核酸,阻断agr群体感应系统,降低MRSA毒力因子的表达,减小细菌生存压力,为抗耐药菌感染提供一种新策略。方法:应用Primer Premier 5.0和RNA structure 4.5两种软件,设计、筛选2条针对agrA mRNA的反义寡核苷酸序列,对其进行锁核酸修饰,并与透膜肽(KFF)3K共价连接,将这两条反义序列导入细菌体内。体外与MRSA共培养,观察细菌生存情况。实时定量PCR检测其对agrA及agr群体感受系统的效应分子RNAⅢ和下游毒力基因hla的转录水平的影响,Western blot检测其是否能抑制α-溶血素的表达。结果:两条反义锁核酸序列PLNA34和PLNA522在体外均无抗菌活性,但都能不同程度的抑制agrA,RNAⅢ和hla的转录水平;相比较而言,PLNA34的抑制效果更佳,并能明显抑制α-溶血素的分泌表达。结论:agrA可作为抗MRSA感染的新靶点,为新型抗MRSA药物的研发提供了理论基础。     

Mucispirillum schaedleri Antagonizes Salmonella Virulence to Protect Mice against Colitis

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Interaction between Staphylococcus Agr virulence and neutrophils regulates pathogen expansion in the skin

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Agr2‐interacting Prod1‐like protein Tfp4 from Xenopus laevis is necessary for early forebrain and eye development as well as for the tadpole appendage regeneration

The Agr family genes, Ag1, Agr2, and Agr3, encode for the thioredoxin domain containing secreted proteins and are specific only for vertebrates. These proteins are attracting increasing attention due to their involvement in many physiological and pathological processes, including exocrine secretion, cancer, regeneration of the body appendages, and the early brain development. At the same time, the mode by which Agrs regulate intracellular processes are poorly understood. Despite that the receptor to Agr2, the membrane anchored protein Prod1, has been firstly discovered in Urodeles, and it has been shown to interact with Agr2 in the regenerating limb, no functional homologs of Prod1 were identified in other vertebrates. This raises the question of the mechanisms by which Agrs can regulate regeneration in other lower vertebrates. Recently, we have identified that Tfp4 (three‐fingers Protein 4), the structural and functional homolog of Prod1 in Anurans, interacts with Agr2 in Xenopus laevis embryos. In the present work we show by several methods that the activity of Tfp4 is essential for the tadpole tail regeneration as well as for the early eye and forebrain development during embryogenesis. These data show for the first time the common molecular mechanism of regeneration regulation in amphibians by interaction of Prod1 and Agr2 proteins.