Characterization of the haemolytic activity of Streptococcus equi.
Abstract: The haemolytic activity of Streptococcus equi, the cause of equine strangles, was characterized. Production of haemolysin in Todd Hewitt broth was dependent on an equine serum supplement and the logarithmic phase of growth after which activity declined sharply. RNA core also induced haemolysin production from cells harvested at the end of the logarithmic phase of growth. Haemolysis was not affected by cholesterol, was only slightly increased in reducing conditions and was completely inactivated by trypan blue, identifying the haemolytic activity as streptolysin S-like (SLS-like). Purification by hydroxyapatite and Sephacryl column chromatography yielded proteins of molecular weights of approximately 6000 and 17 000-22 000 Da with a 64-fold increase in specific activity. Low molecular weight proteins from the RNA core were still present in the purified toxin. Two non-haemolytic mutants were derived by conjugation with an Enterococcus faecalis-carrying transposon Tn916. Southern blots of HindIII digests of DNA revealed that one of the mutants contained three transposon insertions and the other just one. A lambda phage library of S. equi contained plaques whose haemolytic activity was enhanced by reducing conditions and inhibited by cholesterol, suggesting a streptolysin O-like (SLO-like) activity. However, haemolysin in culture sonicates of host E. coli in which the lambda phage insert was subcloned into plasmid (pUC18), was not affected by these conditions. Seven isolates of S. equi in medium without SLS-like inducers showed no SLO-like activity and no evidence for an SLO-like toxin could be found by immunoblotting with pneumolysin antiserum and monoclonal antibodies or by polymerase chain reaction with primers derived from sequences conserved between the SLO genes of Lancefield group A, C and G streptococci. S. equi does not appear to possess a streptolysin O but does make a streptolysin S-like toxin whose production can be interrupted at just one genetic locus.
Copyright 1998 Academic Press Limited.
Publication Date: 1998-05-30 PubMed ID: 9533893DOI: 10.1006/mpat.1997.0190Google Scholar: Lookup
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- Journal Article
Summary
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This study reveals the properties of a blood-dissolving bacteria, Streptococcus equi, which causes an equine disease. Researchers purified the bacteria, identified its proteins and its production capabilities, and discovered it doesn’t possess one type of toxin but does generate a similar one.
Properties and Production of Haemolysin
- Haemolysin is a substance produced by the bacteria Streptococcus equi, which is known to cause equine strangles, a severe infection in horses.
- The production of haemolysin in the study was found to depend on an equine serum supplement and its growth increases during the logarithmic phase which then sharply declines after this phase. Logarithmic phase of growth refers to a period when bacteria grow and reproduce rapidly.
- The study revealed that the component known as RNA core could induce further production of haemolysin from cells that were harvested at the end of the logarithmic phase.
Characteristics of Haemolysis
- The haemolysis process was not affected by cholesterol and was slightly increased under reducing conditions – conditions where oxidation is prevented.
- However, the use of trypan blue, a compound often used to stain cells, completely disabled the haemolytic activity, thus identifying the haemolysis as akin to the streptolysin S (SLS) activity.
Investigations involving Bacterial Toxins
- Purification methods were used to isolate proteins of specific molecular weights. Low molecular weight proteins from the RNA core remained present in the toxin after it was purified.
- Various testing methodologies, including conjugation with a different bacterium and exploration of a bacteriophage library, were used to identify non-hemolytic mutants and related activities.
- These tests indicated there was no Streptolysin O-like (SLO-like) activity, a different kind of bacterial toxin, in the S. equi bacteria samples.
- However, the researchers affirmed the presence of SLS-like toxins. According to genetic evidence, production of this toxin can be halted at a specific genetic location.
Cite This Article
APA
Flanagan J, Collin N, Timoney J, Mitchell T, Mumford JA, Chanter N.
(1998).
Characterization of the haemolytic activity of Streptococcus equi.
Microb Pathog, 24(4), 211-221.
https://doi.org/10.1006/mpat.1997.0190 Publication
Researcher Affiliations
- Animal Health Trust, Lanwades Park, Kentford, Newmarket, Suffolk, CB8 7UU, U.K.
MeSH Terms
- Antibodies, Bacterial
- Bacterial Proteins
- Cholesterol / pharmacology
- Conjugation, Genetic
- DNA Transposable Elements
- DNA, Bacterial / genetics
- Enterococcus faecalis / genetics
- Hemolytic Plaque Technique
- Molecular Weight
- Mutagenesis, Insertional
- Streptococcus equi / chemistry
- Streptococcus equi / immunology
- Streptolysins / analysis
- Streptolysins / biosynthesis
- Streptolysins / chemistry
- Streptolysins / genetics
- Streptolysins / isolation & purification
Citations
This article has been cited 15 times.- Kerstens J, Durmus B, Lambrecht S, Baar I, Ieven MM, Van Der Zijden T, Parizel PM, Menovsky T, Lammens MMY, Jorens PG. Meningoencephalitis with Streptococcus equi Subspecies equi Leading to a Dural Arteriovenous Fistula. Case Rep Neurol Med 2021;2021:9898364.
- Draberova L, Tumova M, Draber P. Molecular Mechanisms of Mast Cell Activation by Cholesterol-Dependent Cytolysins. Front Immunol 2021;12:670205.
- Burkhart BJ, Schwalen CJ, Mann G, Naismith JH, Mitchell DA. YcaO-Dependent Posttranslational Amide Activation: Biosynthesis, Structure, and Function. Chem Rev 2017 Apr 26;117(8):5389-5456.
- Molloy EM, Casjens SR, Cox CL, Maxson T, Ethridge NA, Margos G, Fingerle V, Mitchell DA. Identification of the minimal cytolytic unit for streptolysin S and an expansion of the toxin family. BMC Microbiol 2015 Jul 24;15:141.
- Asam D, Mauerer S, Spellerberg B. Streptolysin S of Streptococcus anginosus exhibits broad-range hemolytic activity. Med Microbiol Immunol 2015 Apr;204(2):227-37.
- Molloy EM, Cotter PD, Hill C, Mitchell DA, Ross RP. Streptolysin S-like virulence factors: the continuing sagA. Nat Rev Microbiol 2011 Aug 8;9(9):670-81.
- Shimomura Y, Okumura K, Murayama SY, Yagi J, Ubukata K, Kirikae T, Miyoshi-Akiyama T. Complete genome sequencing and analysis of a Lancefield group G Streptococcus dysgalactiae subsp. equisimilis strain causing streptococcal toxic shock syndrome (STSS). BMC Genomics 2011 Jan 11;12:17.
- Lannergård J, Flock M, Johansson S, Flock JI, Guss B. Studies of fibronectin-binding proteins of Streptococcus equi. Infect Immun 2005 Nov;73(11):7243-51.
- Alouf JE. Molecular features of the cytolytic pore-forming bacterial protein toxins. Folia Microbiol (Praha) 2003;48(1):5-16.
- Wescombe PA, Tagg JR. Purification and characterization of streptin, a type A1 lantibiotic produced by Streptococcus pyogenes. Appl Environ Microbiol 2003 May;69(5):2737-47.
- Fuller JD, Camus AC, Duncan CL, Nizet V, Bast DJ, Thune RL, Low DE, De Azavedo JC. Identification of a streptolysin S-associated gene cluster and its role in the pathogenesis of Streptococcus iniae disease. Infect Immun 2002 Oct;70(10):5730-9.
- Harrington DJ, Greated JS, Chanter N, Sutcliffe IC. Identification of lipoprotein homologues of pneumococcal PsaA in the equine pathogens Streptococcus equi and Streptococcus zooepidemicus. Infect Immun 2000 Oct;68(10):6048-51.
- Whatmore AM, King SJ, Doherty NC, Sturgeon D, Chanter N, Dowson CG. Molecular characterization of equine isolates of Streptococcus pneumoniae: natural disruption of genes encoding the virulence factors pneumolysin and autolysin. Infect Immun 1999 Jun;67(6):2776-82.
- Wahlenmayer ER, Hammers DE. Streptococcal peptides and their roles in host-microbe interactions. Front Cell Infect Microbiol 2023;13:1282622.
- Turner CE, Bubba L, Efstratiou A. Pathogenicity Factors in Group C and G Streptococci. Microbiol Spectr 2019 May;7(3).
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