Staphylococcal Enterotoxins and Toxic Shock Syndrome Toxin-1 and Their Association among Bacteremic and Infective Endocarditis Patients in Egypt.

PURPOSE: Infective endocarditis (IE) is a major complication in patients with bacteremia of Staphylococcus (S.) aureus infection. Our aim was to determine the association of the major Staphylococcal superantigens (SAgs), including Staphylococcal enterotoxins (SEs) and toxic shock syndrome toxin-1 (TSST-1), among hospitalized patients diagnosed with bacteremia and those with IE. METHODS: This study was conducted on 88 patients; of these, 84 (95.5%) had two positive blood cultures. Eighteen out of the 84 patients (21.4%) were diagnosed based on the modified Duke criteria by a cardiologist to have IE. The recovered isolates were screened phenotypically using ELISA followed by molecular analysis of sea, seb, sec, sed, see, and tsst-1, the major SAg coding genes, and the obtained findings were statistically analyzed. RESULTS: Phenotypic screening for SE production of 26 selected Staphylococci (15 isolated from the IE patients (10 S. aureus and 5 coagulase negative staphylococci (CoNS)) and 11 from bacteremic patients (10 S. aureus and 1 CoNS)) using ELISA revealed that 12/26 (46%) isolates were SE producers. PCR analysis showed that 19 (73%) isolates were PCR positive for SAg genes with the highest prevalence of the sea gene (79%), followed by seb (63%) and tsst-1 (21%). The least frequent gene was sed (5.3%). Statistical correlations between bacteremic and IE isolates with respect to prevalence of SAgs showed no significant difference (P value = 0.139, effect size = 0.572) indicating no specific association between any of the detected SAgs and IE. CONCLUSION: There is high prevalence of SEs among clinical isolates of Staphylococci recovered from patients suffering bacteremia and those with IE. No significant difference was found among Staphylococcal isolates recovered from patients with bacteremia or IE regarding both phenotypic and genotypic detection of the tested SAgs.

Molecular Binding and Simulation Studies of Staphylococcus aureus Superantigens with Flavonoid Compounds.

BACKGROUND: Superantigens of Staphylococcus aureus namely enterotoxin A, exfoliative toxin A, and Toxic shock syndrome toxin-1 cause detrimental effects on the cells of the immune system. METHODS: In this work, the toxins were downloaded from the Protein DataBank database and energies were minimized using KoBaMIN server. Forty flavonoids compounds were identified by pubchem compound database through extensive literature study and their 3D structures were obtained by submitting SMILES to CORINA tool. Based on Lipinski's rule of five, the molecules were filtered that resulted in 27 compounds. Molecular docking was performed for identifying the binding and interaction sites of flavonoids with the toxins using Autodock 4. RESULTS AND CONCLUSION: The docked complexes were then subjected to molecular dynamics simulation using Gromacs. The analysis revealed the stability of the complexes as indicated by three hydrogen bonds formed during the simulation time period of 20 ns.

Interleukin-10 (IL-10) Produced by Mutant Toxic Shock Syndrome Toxin 1 Vaccine-Induced Memory T Cells Downregulates IL-17 Production and Abrogates the Protective Effect against Staphylococcus aureus Infection.

Development of long-term memory is crucial for vaccine-induced adaptive immunity against infectious diseases such as Staphylococcus aureus infection. Toxic shock syndrome toxin 1 (TSST-1), one of the superantigens produced by S. aureus, is a possible vaccine candidate against infectious diseases caused by this pathogen. We previously reported that vaccination with less toxic mutant TSST-1 (mTSST-1) induced T helper 17 (Th17) cells and elicited interleukin-17A (IL-17A)-mediated protection against S. aureus infection 1 week after vaccination. In the present study, we investigated the host immune response induced by mTSST-1 vaccination in the memory phase, 12 weeks after the final vaccination. The protective effect and IL-17A production after vaccination with mTSST-1 were eliminated because of IL-10 production. In the presence of IL-10-neutralizing monoclonal antibody (mAb), IL-17A production was restored in culture supernatants of CD4+ T cells and macrophages sorted from the spleens of vaccinated mice. Vaccinated mice treated with anti-IL-10 mAb were protected against systemic S. aureus infection in the memory phase. From these results, it was suggested that IL-10 produced in the memory phase suppresses the IL-17A-dependent vaccine effect through downregulation of IL-17A production.

Subinhibitory concentrations of tedizolid potently inhibit extracellular toxin production by methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

PURPOSE: Potent extracellular toxins including alpha-haemolysin, Panton-Valentine leukocidin (PVL) and toxic-shock syndrome toxin 1 (TSST-1) significantly contribute to Staphylococcus aureus pathogenesis, thus, toxin suppression is a primary focus in treatment of staphylococcal disease. S. aureus maintains complex strategies to regulate toxin expression and previous data have demonstrated that subinhibitory concentrations of beta-lactam antibiotics can adversely increase S. aureus exotoxin production. The current study evaluates the effects of subinhibitory concentrations of tedizolid, a second-generation oxazolidinone derivative, on expression of staphylococcal exotoxins in both methicillin-resistant and methicillin-sensitive S. aureus. METHODOLOGY: S. aureus exotoxin expression levels were compared at 12 and 24 h following treatment with tedizolid, linezolid, nafcillin or vehicle control. RESULTS: Our findings show that the level of antibiotic required to alter toxin production was strain-dependent and corresponds with the quantity of toxin produced, but both tedizolid and linezolid could effectively reduce expression of alpha-haemolysin, PVL and TSST-1 toxin at subinhibitory concentrations. In contrast, nafcillin showed less attenuation and, in some S. aureus strains, led to an increase in toxin expression. Tedizolid consistently inhibited toxin production at a lower overall drug concentration than comparator agents. CONCLUSION: Together, our data support that tedizolid has the potential to improve outcomes of infection due to its superior ability to inhibit S. aureus growth and attenuate exotoxin production.

Transcriptional profiling of HERV-K(HML-2) in amyotrophic lateral sclerosis and potential implications for expression of HML-2 proteins.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder. About 90% of ALS cases are without a known genetic cause. The human endogenous retrovirus multi-copy HERV-K(HML-2) group was recently reported to potentially contribute to neurodegeneration and disease pathogenesis in ALS because of transcriptional upregulation and toxic effects of HML-2 Envelope (Env) protein. Env and other proteins are encoded by some transcriptionally active HML-2 loci. However, more detailed information is required regarding which HML-2 loci are transcribed in ALS, which of their proteins are expressed, and differences between the disease and non-disease states. METHODS: For brain and spinal cord tissue samples from ALS patients and controls, we identified transcribed HML-2 loci by generating and mapping HML-2-specific cDNA sequences. We predicted expression of HML-2 env gene-derived proteins based on the observed cDNA sequences. Furthermore, we determined overall HML-2 transcript levels by RT-qPCR and investigated presence of HML-2 Env protein in ALS and control tissue samples by Western blotting. RESULTS: We identified 24 different transcribed HML-2 loci. Some of those loci are transcribed at relatively high levels. However, significant differences in HML-2 loci transcriptional activities were not seen when comparing ALS and controls. Likewise, overall HML-2 transcript levels, as determined by RT-qPCR, were not significantly different between ALS and controls. Indeed, we were unable to detect full-length HML-2 Env protein in ALS and control tissue samples despite reasonable sensitivity. Rather our analyses suggest that a number of HML-2 protein variants other than full-length Env may potentially be expressed in ALS patients. CONCLUSIONS: Our results expand and refine recent publications on HERV-K(HML-2) and ALS. Some of our results are in conflict with recent findings and call for further specific analyses. Our profiling of HML-2 transcription in ALS opens up the possibility that HML-2 proteins other than canonical full-length Env may have to be considered when studying the role of HML-2 in ALS disease.

Epidemiology of toxic shock syndrome toxin-1 harboring Staphylococcus aureus obtained from clinical samples in Iran: A Systematic Review and Meta-analysis.

BACKGROUND: S. aureus strains, with the capability of producing toxic shock syndrome toxin-1 (TSST-1), are more likely to cause complicated infections. However, due to lack of comprehensive local data on the prevalence of TSST-1, we aimed to determine the prevalence of TSST-1 harboring S. aureus isolates in Iran. METHODS: A systematic search was performed by using PubMed and Scopus databases from papers published by Iranian authors from January 2000 to the end of March 2017. Then, 10 publications which were matched with inclusion criteria were selected for data extraction and analysis by Comprehensive Meta-Analysis Software. RESULTS: The overall prevalence of TSST-1 carrying S. aureus in Iran was 21.3% (95% CI: 7.9%-46.1%), ranging from 0% to 68%. Moreover, from the included studies, the pooled prevalence of TSST-1 producing MRSA isolates was estimated to be 25.2% (95% CI: 13.3%-42.5%), ranging from 0% to 69.8%. From those studies which showed the distribution of toxin-harboring S. aureus it was found that the skin and soft tissue, respiratory and bloodstream infections were the common sites of TSST-1 harboring S. aureus. CONCLUSIONS: In summary, it seems that emergence of MRSA strains leads to higher prevalence of TSST-1 carrying strains in the north of Iran. However, further research is required to elucidate the interplay between the outcome of diseases and TSST-1 producing strains, especially in our country.

The effect of vaginal microbial communities on colonization by Staphylococcus aureus with the gene for toxic shock syndrome toxin 1 (TSST-1): a case-control study.

Menstrual toxic shock syndrome is associated with vaginal colonization by Staphylococcus aureus strains that encode toxic shock syndrome toxin 1 (tst+). Interestingly, a small proportion of women are colonized by S. aureus tst+ but do not have symptoms of toxic shock syndrome. Here we sought to determine if differences in the species composition of vaginal bacterial communities reflect a differential risk of colonization by S. aureus capable of producing toxic shock syndrome toxin 1 (TSST-1). The composition of vaginal communities of women that were or were not colonized with S. aureus tst+ were compared based on terminal restriction fragment length polymorphism (T-RFLP) profiles and sequences of cloned 16S rRNA genes. There were no detectable differences in community composition or species rank abundance between communities of women vaginally colonized with S. aureus tst+ as compared to those that were not. Phylogenetic analysis of cloned 16S rRNA gene sequences showed that the predominant members of communities of women colonized with S. aureus tst+ were indistinguishable from those of other healthy women. The data suggest that the numerically dominant members of vaginal communities do not preclude colonization and proliferation of S. aureus tst+ within indigenous microbial communities of the vagina.

Impact of Currently Marketed Tampons and Menstrual Cups on Staphylococcus aureus Growth and Toxic Shock Syndrome Toxin 1 Production In Vitro.

Fifteen currently marketed intravaginal protection products (11 types of tampon and 4 types of menstrual cup) were tested by the modified tampon sac method to determine their effect on Staphylococcus aureus growth and toxic shock syndrome toxin 1 (TSST-1) production. Most tampons reduced S. aureus growth and TSST-1 production, with differences based on brand and composition, and the level of S. aureus growth was higher in destructured than in unaltered tampons. We observed higher levels of S. aureus growth and toxin production in menstrual cups than in tampons, potentially due to the additional air introduced into the bag by cups, with differences based on cup composition and size.IMPORTANCE Menstrual toxic shock syndrome is a rare but severe disease. It occurs in healthy women vaginally colonized by Staphylococcus aureus producing toxic shock syndrome toxin 1 using intravaginal protection, such as tampons or menstrual cups. Intravaginal protection induces TSS by the collection of catamenial products, which act as a growth medium for S. aureus Previous studies evaluated the impact of tampon composition on S. aureus producing toxic shock syndrome toxin 1, but they are not recent and did not include menstrual cups. This study demonstrates that highly reproducible results for S. aureus growth and TSST-1 production can be obtained by using a simple protocol that reproduces the physiological conditions of tampon and cup usage as closely as possible, providing recommendations for tampon or cup use to both manufacturers and consumers. Notably, our results do not show that menstrual cups are safer than tampons and suggest that they require similar precautions.

Epidemiological Trends Observed from Molecular Characterization of Methicillin-Resistant Staphylococcus aureus Isolates from Blood Cultures at a Japanese University Hospital, 2012-2015.

Despite increasing reports of skin and soft tissue infections caused by community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) in Japan, the extent to which these strains cause nosocomial infections remains unknown, and this is especially true for bloodstream infections. In this study, we molecularly characterized MRSA isolates from Japanese blood samples. Among the 151 MRSA isolates collected from 53 medical facilities in 2011, 115 (76%) and 30 (20%) were classified as staphylococcal cassette chromosome mec (SCCmec) types II and IV, respectively, while the Panton-Valentine leukocidin (PVL) gene was detected in only two isolates. Among 66 MRSA isolates collected from Tokyo Medical University Hospital between 2012 and 2015, 43 (65%) and 20 (30%) were classifiable as SCCmec types II and IV, respectively. In 2015, highly virulent strains, such as the SCCmec type IV/PVL and SCCmec type IV/ toxic shock syndrome toxin-1 clonal types, increased in number. Therefore, the SCCmec type IV clone may cause invasive infections not only in community settings but also in healthcare settings in Japan.

Characterization of Toxin Genes and Antimicrobial Susceptibility of Staphylococcus aureus Isolates in Fishery Products in Iran.

Staphylococcus aureus is one of the most common causes of seafood-borne diseases worldwide, which are attributable to the contamination of food by preformed enterotoxins. In this study, a total of 206 (34.3%) Staphylococcus aureus strains were obtained from 600 fish and shrimp samples and were tested for their antimicrobial susceptibility. We assessed the prevalence of the genes responsible for the staphylococcal enterotoxins (SEA, SEB) and toxic shock syndrome toxin 1 (TSST-1) genes. The results indicated that 34% of aqua food samples were contaminated with S. aureus, and 23.8% of these isolates were mec-A-positive. Sixty-four percent of the strains isolated from contaminated seafood was enterotoxigenic S. aureus, and 28.2% of SEs were MRSA-positive. The most prevalent genotype was characterized by the presence of the sea gene (45.2%), followed by the seb gene (18.5%), and the tst gene encoding TSST-1 was found in eight strains (3.9%). Of the 206 S. aureus isolates, 189 strains (84.9%) were resistant to at least one antibiotic. Given the frequent outbreaks of enterotoxigenic MRSA, it is necessary to make revisions to mandatory programmes to facilitate improved hygiene practices during fishing, aquaculture, processing, and sales to prevent the contamination of fishery products in Iran.

Significant Differences in Antigen-Induced Transendothelial Migration of Human CD8 and CD4 T Effector Memory Cells.

OBJECTIVE: Circulating human T effector memory cell (TEM) recognition of nonself MHC (major histocompatibility complex) molecules on allograft endothelial cells can initiate graft rejection despite elimination of professional antigen-presenting cells necessary for naive T-cell activation. Our previous studies of CD4 TEM have established that engagement of the T-cell receptor not only activates T cells but also triggers transendothelial migration (TEM) by a process that is distinct from that induced by activating chemokine receptors on T cells, being slower, requiring microtubule-organizing center-directed cytolytic granule polarization to and release from the leading edge of the T cell, and requiring engagement of proteins of the endothelial cell lateral border recycling compartment. Although CD4 TEM may contribute to acute allograft rejection, the primary effectors are alloreactive CD8 TEM. Whether and how T-cell receptor engagement affects TEM of human CD8 TEM is unknown. APPROACH AND RESULTS: We modeled TEM of CD8 TEM across cultured human microvascular endothelial cells engineered to present superantigen under conditions of venular shear stress in vitro in a flow chamber. Here, we report that T-cell receptor engagement can also induce TEM of this population that similarly differs from chemokine receptor-driven TEM with regard to kinetics, morphological manifestations, and microtubule-organizing center dynamics as with CD4 TEM. However, CD8 TEM do not require either cytolytic granule release or interactions with proteins of the lateral border recycling compartment. CONCLUSIONS: These results imply that therapeutic strategies designed to inhibit T-cell receptor-driven recruitment based on targeting granule release or components of the lateral border recycling compartment will not affect CD8 TEM and are unlikely to block acute rejection in the clinic.

Preparation and In Vitro Evaluation of Antitumor Activity of TGFαL3-SEB as a Ligand-Targeted Superantigen.

Tumor-targeted superantigens (TTSs) have been used to treat a variety of tumors in preclinical studies. The TTS utilizes the powerful T-cell activation strategy by means of staphylococcal enterotoxins (SEs) as superantigens (Sags) to target tumor cells. Monoclonal antibodies and tumor-related ligands have been used as targeting molecules of Sag. In this study, we assessed the antitumor potency of tumor-targeted superantigen (TTS) strategy to design and produce fusion protein as a new antitumor candidate. The third loop (L3) of transforming growth factor α (TGF-α) was genetically conjugated to staphylococcal enterotoxin type B (TGFαL3-SEB), and its in vitro antitumor activity against murine breast cancer cells (A431 cell line) was evaluated. We designed and prepared TGFαL3-SEB chimeric protein and evaluated superantigenic activity, binding property to cancer cells, overexpression of epidermal growth factor receptor (EGFR), and in vitro antitumor activities. Cloning of tgfαl3-seb was confirmed by colony-polymerase chain reaction, enzymatic digestion, and sequencing. The recombinant TGFαL3-SEB fusion protein with molecular weight of 31 kDa was expressed and confirmed by anti-His Western-blot analysis. The TGFαL3-SEB fusion protein attached to A431 cell line with proper affinity and induced dose-dependent cytotoxicity against EGFR-expressing cancer cells in vitro. The TGFαL3-SEB chimeric protein exhibited potent in vitro antitumor activity. Our findings indicated that TGFαL3-SEB may be a promising anticancer candidate in cancer immunotherapy, and further studies are required to explore its potential in vivo therapeutic applications.

Superantigens produced by catheter-associated Staphylococcus aureus elicit systemic inflammatory disease in the absence of bacteremia.

SAgs, produced by Staphylococcus aureus, play a major role in the pathogenesis of invasive staphylococcal diseases by inducing potent activation of the immune system. However, the role of SAgs, produced by S. aureus, associated with indwelling devices or tissues, are not known. Given the prevalence of device-associated infection with toxigenic S. aureus in clinical settings and the potency of SAgs, we hypothesized that continuous exposure to SAgs produced by catheter-associated S. aureus could have systemic consequences. To investigate these effects, we established a murine in vivo catheter colonization model. One centimeter long intravenous catheters were colonized with a clinical S. aureus isolate producing SAgs or isogenic S. aureus strains, capable or incapable of producing SAg. Catheters were subcutaneously implanted in age-matched HLA-DR3, B6, and AE(o) mice lacking MHC class II molecules and euthanized 7 d later. There was no evidence of systemic infection. However, in HLA-DR3 transgenic mice, which respond robustly to SSAgs, the SSAg-producing, but not the nonproducing strains, caused a transient increase in serum cytokine levels and a protracted expansion of splenic CD4(+) T cells expressing SSAg-reactive TCR Vß8. Lungs, livers, and kidneys from these mice showed infiltration with CD4(+) and CD11b(+) cells. These findings were absent in B6 and AE(o) mice, which are known to respond poorly to SSAgs. Overall, our novel findings suggest that systemic immune activation elicited by SAgs, produced by S. aureus colonizing foreign bodies, could have clinical consequences in humans.

[ENTEROTOXIGENICITY OF STAPHYLOCOCCUS AUREUS STRAINS, ISOLATED FROM BREAST MILK OF WOMEN, FEEDING CHILDREN WITH INFECTIOUS PATHOLOGY].

AIM: Determination of enterotoxigenicity and ability to synthesize TSST-1 in S. aureus strains, isolated from breast milk of women, feeding children with infectious pathology. MATERIALS AND METHODS: 35 S. aureus strains, isolated from breast milk of women feeding children with varying infectious pathology in hospitals and as outpatients were studied for the presence of staphylococci enterotoxins (SE) of types A and B and toxic shock syndrome toxin (TSST-1). Determination of SEA, SEB and TSST-1 was carried out by enzyme immunoassay. RESULTS: Toxins were detected in 94.2% of S. aureus strains. SEB was synthesized by 86.7%, SEA--34.3%, TSST-1--42.8% of S. aureus strains. Toxins were detected with equal frequencies in healthy women and women with inflammatory diseases of breasts. Differences in frequency of colonization of intestines of children receiving breast milk, infected with toxigenic and non-toxigenic staphylococci strains was not detected. CONCLUSION: A high frequency of occurrence of enterotoxins and TSST-1 in S. aureus, isolated from breast milk of the mother during infectious pathology in the child was discovered. Enterotoxigenic strains can be detected in breast milk in healthy women. Study of the role of breast milk, infected with S. aureus, producing SEA, SEB And TSST-1 in development of child pathology is necessary.

Regulation of Staphylococcal Superantigen-Like Gene, ssl8, Expression in Staphylococcus aureus strain, RN6390.

Staphylococcal superantigen-like (SSL) proteins, which are encoded by a cluster of eleven ssl genes, contribute to the Staphylococcus aureus virulence. Recently we reported ssl8 expression profiles in seven clinically important strains-MW2, USA300FPR3757, MSSA476, Newman, RN6390, Mu50, and N315-and showed the differential expression of ssl8 in Newman, RN6390, and USA300FPR3757 strains, despite harboring identical allelic forms of ssl8, suggesting the roles for different regulatory elements for this gene in different S. aureus strains. In this communication, using RN6390, a common laboratory S. aureus strain and its isogenic knockout mutant strains of agr, sae, sarA, sigB, rot, and the agr-/sigB (-) double mutant, we showed that SarA and Rot are inducer and repressor, respectively, for ssl8 expression in RN6390. This is in contrast to the Newman strain, where ssl8 is positively regulated by Sae but negatively by Agr, indicating the variable expression of ssl8 in clinical strains is more likely due to strain-specific regulatory elements.

Cloning, expression, crystallization and preliminary X-ray diffraction studies of staphylococcal superantigen-like protein 1 (SSL1).

Staphylococcus aureus produces a family of exotoxins which are structural homologues of superantigens and thus are called staphylococcal superantigen-like proteins (SSLs). Amongst the 14 SSL genes, ssl1 (SAOUHSC_00383) has been cloned in the pQE30 expression vector, overexpressed in Escherichia coli M15 (pREP4) cells and the protein purified to homogeneity. The protein was crystallized using 6% Tacsimate pH 6.0, 0.1 M MES pH 6.0, 25%(w/v) polyethylene glycol 3350, 100 mM NDSB 256 at 298 K by the sitting-drop vapour-diffusion method. The crystals belonged to space group P21, with unit-cell parameters a = 77.9, b = 70.5, c = 126.5 Å, ß = 106.2°. X-ray diffraction data were collected and processed to a maximum resolution of 2.5 Å. The crystal contains six molecules in the asymmetric unit.

Novel antimicrobial peptides that inhibit gram positive bacterial exotoxin synthesis.

Gram-positive bacteria, such as Staphylococcus aureus, cause serious human illnesses through combinations of surface virulence factors and secretion of exotoxins. Our prior studies using the protein synthesis inhibitor clindamycin and signal transduction inhibitors glycerol monolaurate and α-globin and ß-globin chains of hemoglobin indicate that their abilities to inhibit exotoxin production by S. aureus are separable from abilities to inhibit growth of the organism. Additionally, our previous studies suggest that inhibition of exotoxin production, in absence of ability to kill S. aureus and normal flora lactobacilli, will prevent colonization by pathogenic S. aureus, while not interfering with lactobacilli colonization. These disparate activities may be important in development of novel anti-infective agents that do not alter normal flora. We initiated studies to explore the exotoxin-synthesis-inhibition activity of hemoglobin peptides further to develop potential agents to prevent S. aureus infections. We tested synthesized α-globin chain peptides, synthetic variants of α-globin chain peptides, and two human defensins for ability to inhibit exotoxin production without significantly inhibiting S. aureus growth. All of these peptides were weakly or not inhibitory to bacterial growth. However, the peptides were inhibitory to exotoxin production with increasing activity dependent on increasing numbers of positively-charged amino acids. Additionally, the peptides could be immobilized on agarose beads or have amino acid sequences scrambled and still retain exotoxin-synthesis-inhibition. The peptides are not toxic to human vaginal epithelial cells and do not inhibit growth of normal flora L. crispatus. These peptides may interfere with plasma membrane signal transduction in S. aureus due to their positive charges.

A model of an integrated immune system pathway in Homo sapiens and its interaction with superantigen producing expression regulatory pathway in Staphylococcus aureus: comparing behavior of pathogen perturbed and unperturbed pathway.

Response of an immune system to a pathogen attack depends on the balance between the host immune defense and the virulence of the pathogen. Investigation of molecular interactions between the proteins of a host and a pathogen helps in identifying the pathogenic proteins. It is necessary to understand the dynamics of a normally behaved host system to evaluate the capacity of its immune system upon pathogen attack. In this study, we have compared the behavior of an unperturbed and pathogen perturbed host system. Moreover, we have developed a formalism under Flux Balance Analysis (FBA) for the optimization of conflicting objective functions. We have constructed an integrated pathway system, which includes Staphylococcal Superantigen (SAg) expression regulatory pathway and TCR signaling pathway of Homo sapiens. We have implemented the method on this pathway system and observed the behavior of host signaling molecules upon pathogen attack. The entire study has been divided into six different cases, based on the perturbed/unperturbed conditions. In other words, we have investigated unperturbed and pathogen perturbed human TCR signaling pathway, with different combinations of optimization of concentrations of regulatory and signaling molecules. One of these cases has aimed at finding out whether minimization of the toxin production in a pathogen leads to the change in the concentration levels of the proteins coded by TCR signaling pathway genes in the infected host. Based on the computed results, we have hypothesized that the balance between TCR signaling inhibitory and stimulatory molecules can keep TCR signaling system into resting/stimulating state, depending upon the perturbation. The proposed integrated host-pathogen interaction pathway model has accurately reflected the experimental evidences, which we have used for validation purpose. The significance of this kind of investigation lies in revealing the susceptible interaction points that can take back the Staphylococcal Enterotoxin (SE)-challenged system within the range of normal behavior.

Target expression of Staphylococcus enterotoxin A from an oncolytic adenovirus suppresses mouse bladder tumor growth and recruits CD3+ T cell.

We recently engineered an oncolytic adenovirus (PPE3-SEA) that expresses the superantigen, Staphylococcus enterotoxin A (SEA), and that has enhanced tumor specificity because the telomerase reverse transcriptase and hypoxia-inducible factor promoters regulate expression of E1A and E1B genes, respectively. Here, we evaluated the PPE3-SEA adenovirus anti-tumor activity against MB49 mouse bladder cancer cell proliferation in vitro and in vivo. PPE3-SEA infection of murine MB49 cells in vitro induced cytopathic effects, and significant expression of SEA mRNA and protein, as measured by RT-PCR and western blot, respectively. Subcutaneous MB29 bladder tumors were established in syngeneic C57BL/6 mice. After 10 days, tumors were injected with either oncolytic virus or PBS. Tumor dimensions were measured on days 1, 3, 5, 7, 9, and 11 post-treatment and tumor volumes were calculated. One of eight PPE3-SEA-treated mice had no tumor by day 9. PPE3-SEA treated group had significantly lower mean tumor volume beginning on day 5 post-treatment (p < 0.01), more fibrous tissue in the tumor, and increased presence of infiltrating CD3+ T cells than those of the control group. Gross appearance and histologic sections from the livers and kidneys of the PPE3-SEA-treated group were similar to those of the control group. In conclusion, oncolytic adenoviruses can provide a novel delivery vehicle for SEA to tumor sites, and PPE3-SEA warrants further study as a potential anti-tumor agent for bladder cancer.