[53] Assays of hemolytic toxins

Rowe, G E; Welch, Rodney A.

doi:10.1016/0076-6879(94)35179-1

Cited by 113 publications

(94 citation statements)

References 49 publications

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“…Strain WP1 T exhibited a sharp clear zone of b-haemolysis on blood agar. This is of potential importance in the aetiology of white plague type II, because this assay is indicative of cytolytic toxins (Rowe & Welch, 1994). Biochemical testing revealed further that WP1 T is strongly ureolytic, since the urease test was positive after a few hours of incubation.…”

Section: Cultural Physiological and Biochemical Characteristicsmentioning

confidence: 99%

Aurantimonas coralicida gen. nov., sp. nov., the causative agent of white plague type II on Caribbean scleractinian corals

Denner

Smith

Busse

et al. 2003

International Journal of Systematic and Evolutionary Microbiology

209

166

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A bacterium previously isolated from a diseased colony of the scleractinian coral Dichocoenia stokesi (common name elliptical star coral) was subjected to a detailed polyphasic taxonomic characterization. The isolate, designated WP1 T , was halophilic and strictly aerobic and formed golden-orange-pigmented colonies after prolonged incubation. Cells of WP1 T were Gram-negative, rod-shaped and showed a characteristic branching rod morphology. Chemotaxonomically, WP1 T was characterized by having Q-10 as the major respiratory lipoquinone and sym-homospermidine as the main component of the cellular polyamine content. The predominant constituent in the cellular fatty acid profile was C 18 : 1 v7c, along with C 19 : 0 cyclo v8c and C 16 : 0 . Other fatty acids present in smaller amounts were C 17 : 0 , C 18 : 0 , C 16 : 1 v7c, C 20 : 1 v7c and C 18 : 1 2-OH. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine. Minor amounts of diphosphatidylglycerol, phosphatidylmonomethylethanolamine and phosphatidyldimethylethanolamine were present. The G+C content of the genomic DNA was 66?3 mol%. Phylogenetic analysis of the 16S rRNA gene sequence showed that WP1 T represents a separate subline of descent within the order 'Rhizobiales' of the 'Alphaproteobacteria'. The new line of descent falls within the group of families that includes the Rhizobiaceae, Bartonellaceae, Brucellaceae and 'Phyllobacteriaceae', with no particular relative within this group. The 16S rRNA gene sequence similarity to all established taxa within this group was not higher than 92?0 % (to Mesorhizobium mediterraneum). To accommodate this emerging coral pathogen, the creation of a new genus and species is proposed, Aurantimonas coralicida gen. nov., sp. nov. (type strain WP1 T =CIP 107386 T =DSM 14790 T ). INTRODUCTIONIn 1995, Richardson and co-workers documented a dramatic coral epizootic that occurred on reefs of the northern Florida Keys, which spread rapidly to infect 17 of the 43 species of scleractinian corals present. Mortality rates of up to 38 % of the most susceptible coral species, Dichocoenia stokesi (the elliptical star coral), occurred within periods as short as 10 weeks (Richardson et al., 1998a). The disease was designated white plague type II The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of Aurantimonas coralicida strain WP1 T is AJ065627.Abbreviations: DPG, diphosphatidylglycerol; PC, phosphatidylcholine; PDE, phosphatidyldimethylethanolamine; PE, phosphatidylethanolamine; PG, phosphatidylglycerol; PME, phosphatidylmonomethylethanolamine. G 2003 IUMS Printed in Great Britain1115 , 1998a). Beyond it, a replication sequence comparison between the re-determined 16S rRNA gene sequence in the study presented here and the originally deposited sequence revealed that there was no significant degree of similarity (~82 %) between the two sequences.In order to exclude any strain confusion, we have followed the history and distribution of strain WP1 T among our different laborato...

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Section: Cultural Physiological and Biochemical Characteristicsmentioning

confidence: 99%

Aurantimonas coralicida gen. nov., sp. nov., the causative agent of white plague type II on Caribbean scleractinian corals

Denner

Smith

Busse

et al. 2003

International Journal of Systematic and Evolutionary Microbiology

209

166

View full text Add to dashboard Cite

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“…The hemolytic activity of ClyA and ClyA-GFP was measured by a liquid hemolysis assay adopted from Rowe and Welch (42). Sheep erythrocytes (Becton Dickinson) were washed and diluted 1:100 in PBS.…”

Section: Methodsmentioning

confidence: 99%

Delivery of foreign antigens by engineered outer membrane vesicle vaccines

Chen

Osterrieder

Metzger

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

256

247

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As new disease threats arise and existing pathogens grow resistant to conventional interventions, attention increasingly focuses on the development of vaccines to induce protective immune responses. Given their admirable safety records, protein subunit vaccines are attractive for widespread immunization, but their disadvantages include poor immunogenicity and expensive manufacture. We show here that engineered Escherichia coli outer membrane vesicles (OMVs) are an easily purified vaccine-delivery system capable of greatly enhancing the immunogenicity of a low-immunogenicity protein antigen without added adjuvants. Using green-fluorescent protein (GFP) as the model subunit antigen, genetic fusion of GFP with the bacterial hemolysin ClyA resulted in a chimeric protein that elicited strong anti-GFP antibody titers in immunized mice, whereas immunization with GFP alone did not elicit such titers. Harnessing the specific secretion of ClyA to OMVs, the ClyA-GFP fusion was found localized in OMVs, resulting in engineered recombinant OMVs. The anti-GFP humoral response in mice immunized with the engineered OMV formulations was indistinguishable from the response to the purified ClyA-GFP fusion protein alone and equal to purified proteins absorbed to aluminum hydroxide, a standard adjuvant. In a major improvement over current practice, engineered OMVs containing ClyA-GFP were easily isolated by ultracentrifugation, effectively eliminating the need for laborious antigen purification from cell-culture expression systems. With the diverse collection of heterologous proteins that can be functionally localized with OMVs when fused with ClyA, this work signals the possibility of OMVs as a robust and tunable technology platform for a new generation of prophylactic and therapeutic vaccines.protein subunit | adjuvant

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“…HlyE activity was measured essentially as described by Rowe & Welch (1994). Briefly, HlyE protein (isolated as previously described; Atkins et al, 2000) was incubated with defibrinated sterile horse blood (TCS Microbiology) in Tris-buffered saline (50 mM Tris/HCl, pH 8.0, containing 150 mM NaCl).…”

Section: Methodsmentioning

confidence: 99%

The formation and structure of Escherichia coli K-12 haemolysin E pores

et al. 2008

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Some enteric bacteria synthesize a pore-forming toxin, HlyE, which is cytolytic and cytotoxic to host cells. Measurement of HlyE binding to erythrocyte ghosts and the kinetics of HlyE-mediated erythrocyte lysis suggests that interaction with target membranes is not the rate-limiting step in the formation of HlyE pores, but that there is a temperature-dependent lag phase before a functional pore is formed. Circular dichroism and fluorescence energy transfer analyses show that HlyE protomers retain an a-helical structure when oligomerized to form a pore consisting of parallel HlyE protomers. Comparison of the proteolytic sensitivities of the water-soluble and oligomeric forms of HlyE identifies inner and outer surfaces of the pore. This new information has been used to constrain a model of the HlyE pore, which allows a more detailed interpretation of previous low-resolution 3D reconstructions and suggests a novel mechanism for insertion of HlyE into target membranes.

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[53] Assays of hemolytic toxins

Cited by 113 publications

References 49 publications

Aurantimonas coralicida gen. nov., sp. nov., the causative agent of white plague type II on Caribbean scleractinian corals

Aurantimonas coralicida gen. nov., sp. nov., the causative agent of white plague type II on Caribbean scleractinian corals

Delivery of foreign antigens by engineered outer membrane vesicle vaccines

The formation and structure of Escherichia coli K-12 haemolysin E pores

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