Effect of a bacterial pheromone peptide on host chemokine degradation in group A streptococcal necrotising soft-tissue infections

Hebrew University of Jerusalem, Yerushalayim, Jerusalem, Israel
The Lancet (Impact Factor: 45.22). 03/2004; 363(9410):696-703. DOI: 10.1016/S0140-6736(04)15643-2
Source: PubMed


Necrotising soft-tissue infections due to group A streptococcus (GAS) are rare (about 0.2 cases per 100000 people). The disease progresses rapidly, causing severe necrosis and hydrolysis of soft tissues. Histopathological analysis of necrotic tissue debrided from two patients (one with necrotising fasciitis and one with myonecrosis) showed large quantities of bacteria but no infiltrating neutrophils. We aimed to investigate whether the poor neutrophil chemotaxis was linked with the ability of group A streptococcus (GAS) to degrade host chemokines.
We did RT-PCR, ELISA, and dot-blot assays to establish whether GAS induces synthesis of interleukin 8 mRNA, but subsequently degrades the released chemokine protein. Class-specific protease inhibitors were used to characterise the protease that degraded the chemokine. We used a mouse model of human soft-tissue infections to investigate the pathogenic relevance of GAS chemokine degradation, and to test the therapeutic effect of a GAS pheromone peptide (SilCR) that downregulates activity of chemokine protease.
The only isolates from the necrotic tissue were two beta-haemolytic GAS strains of an M14 serotype. A trypsin-like protease released by these strains degraded human interleukin 8 and its mouse homologue MIP2. When innoculated subcutaneously in mice, these strains produced a fatal necrotic soft-tissue infection that had reduced neutrophil recruitment to the site of injection. The M14 GAS strains have a missense mutation in the start codon of silCR, which encodes a predicted 17 aminoacid pheromone peptide, SilCR. Growth of the M14 strain in the presence of SilCR abrogated chemokine proteolysis. When SilCR was injected together with the bacteria, abundant neutrophils were recruited to the site of infection, bacteria were cleared without systemic spread, and the mice survived. The therapeutic effect of SilCR was also obtained in mice challenged with M1 and M3 GAS strains, a leading cause of invasive infections.
The unusual reduction in neutrophils in necrotic tissue of people with GAS soft-tissue infections is partly caused by a GAS protease that degrades interleukin 8. In mice, degradation can be controlled by administration of SilCR, which downregulates GAS chemokine protease activity. This downregulation increases neutrophil migration to the site of infection, preventing bacterial spread and development of a fulminant lethal systemic infection.

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Available from: Miriam Ravins, Oct 06, 2015
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    • "A number of animal models of musculoskeletal infection have been reported, most of which simulate osteomyelitis [1]–[6]. Although a few animal models of soft-tissue infection have been reported [7]–[9], these models are not always reproducible or sustainable due to the leakage and spread of bacteria from the inoculated tissues. Furthermore, sequential analyses of the extent of musculoskeletal infection and number of bacteria in these models require sacrifice of the animals at each analysis time point. "
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    ABSTRACT: Musculoskeletal infections, including surgical-site and implant-associated infections, often cause progressive inflammation and destroy areas of the soft tissue. Treating infections, especially those caused by multi-antibiotic resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA) remains a challenge. Although there are a few animal models that enable the quantitative evaluation of infection in soft tissues, these models are not always reproducible or sustainable. Here, we successfully established a real-time, in vivo, quantitative mouse model of soft-tissue infection in the superficial gluteus muscle (SGM) using bioluminescence imaging. A bioluminescent strain of MRSA was inoculated into the SGM of BALB/c adult male mice, followed by sequential measurement of bacterial photon intensity and serological and histological analyses of the mice. The mean photon intensity in the mice peaked immediately after inoculation and remained stable until day 28. The serum levels of interleukin-6, interleukin-1 and C-reactive protein at 12 hours after inoculation were significantly higher than those prior to inoculation, and the C-reactive protein remained significantly elevated until day 21. Histological analyses showed marked neutrophil infiltration and abscesses containing necrotic and fibrous tissues in the SGM. With this SGM mouse model, we successfully visualized and quantified stable bacterial growth over an extended period of time with bioluminescence imaging, which allowed us to monitor the process of infection without euthanizing the experimental animals. This model is applicable to in vivo evaluations of the long-term efficacy of novel antibiotics or antibacterial implants.
    PLoS ONE 09/2014; 9(9):e106367. DOI:10.1371/journal.pone.0106367 · 3.23 Impact Factor
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    • "equisimilis (Belotserkovsky et al., 2009). In the GAS M14 serotype, sil controls virulence as was shown using different animal models of human NF (Hidalgo-Grass et al., 2002, 2004; Kizy and Neely, 2009). Hitherto, it was possible to activate sil by providing the bacterium with a minute quantity of the mature synthetic autoinducer peptide SilCR (Belotserkovsky et al., 2009), but the conditions under which sil is naturally self-activated were not identified. "
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    ABSTRACT: The connection between bacterial pathogens and unfolded protein response (UPR) is poorly explored. In this review we highlight the evidence showing that group A streptococcus (GAS) induces endoplasmic reticulum (ER) stress and UPR through which it captures the amino acid asparagine (ASN) from the host. GAS acts extracellularly and during adherence to host cells it delivers the hemolysin toxins; streptolysin O (SLO) and streptolysin S (SLS). By poorly understood pathways, these toxins trigger UPR leading to the induction of the transcriptional regulator ATF4 and consequently to the upregulation of asparagine synthetase (ASNS) transcription leading to production and release of ASN. GAS senses ASN and alters gene expression profile accordingly, and increases the rate of multiplication. We suggest that induction of UPR by GAS and by other bacterial pathogens represent means through which bacterial pathogens gain nutrients from the host, obviating the need to become internalized or inflict irreversible cell damage.
    Frontiers in Cellular and Infection Microbiology 08/2014; 4:105. DOI:10.3389/fcimb.2014.00105 · 3.72 Impact Factor
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    • "The presence of a bacterial chemokine-degrading enzyme was first described in a patient suffering from necrotizing fasciitis. Histology revealed a striking paucity of neutrophils despite the massive streptococcal infection and was found to be due to IL-8 degradation by GAS (Hidalgo-Grass et al., 2004). The protein responsible for IL-8 degradation was later identified as SpyCEP, a subtilisin-like serine protease present in the supernatant of various GAS blood isolates (Edwards et al., 2005). "
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    ABSTRACT: SpyCEP-mediated chemokine degradation translates into more efficient spreading and increased severity of invasive Group A Streptococcus (GAS) infections, due to impaired neutrophil recruitment to the site of infection. SpyCEP is markedly up-regulated in invasive as compared to colonizing GAS isolates raising the question whether SpyCEP expression hinders bacterial attachment and thus colonization of the host. To address this question we used a molecular approach involving the use of homologous GAS strains either expressing or not SpyCEP or expressing an enzymatically inactive variant of SpyCEP. We found that expression of enzymatically functional SpyCEP lowered GAS adherence and invasion potential toward various epithelial and endothelial cells. SpyCEP also blunted biofilm formation capacity. Our data indicate that expression of SpyCEP decreases colonization and thus might be detrimental for the spreading of GAS.
    Frontiers in Microbiology 07/2014; 5:339. DOI:10.3389/fmicb.2014.00339 · 3.99 Impact Factor
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