LPS-primed CD11b+ leukocytes serve as an effective carrier of Shiga toxin 2 to cause hemolytic uremic syndrome in mice.

Shiga toxin (Stx)-induced hemolytic uremic syndrome (HUS) is a life-threatening complication associated with Stx-producing Escherichia coli infection. One critical barrier of understanding HUS is how Stx transports from infected intestine to kidney to cause HUS. Passive dissemination seems unlikely, while circulating blood cells have been debated to serve as the toxin carrier. Employing a murine model of Stx2-induced HUS with LPS priming (LPS-Stx2), we investigate how Stx causes HUS and identify possible toxin carrier. We show that peripheral white blood cells (WBC), but not other blood cells or cell-free plasma, carry Stx2 in LPS-Stx2-treated mice. The capability of WBC binding to Stx2 is confirmed in brief ex vivo Stx2 incubation, and adoptively transferring these Stx2-bound WBC into mice induces HUS. Cell separation further identifies a subpopulation in the CD11b+ myeloid leukocytes not the CD11b- lymphocytes group act as the toxin carrier, which captures Stx2 upon exposure and delivers the toxin in vivo. Interestingly, LPS-induced inflammation significantly augments these leukocytes for binding to Stx2 and enhances HUS toxicity. Our results demonstrate that a specific fraction of circulating leukocytes carry Stx2 and cause HUS in vivo, and that LPS priming enhances the carrier capacity and aggravates organ damage.

CCR2-dependent Gr1high monocytes promote kidney injury in shiga toxin-induced hemolytic uremic syndrome in mice.

The hemolytic uremic syndrome (HUS) is a life-threatening disease of the kidney that is induced by shiga toxin-producing E.coli. Major changes in the monocytic compartment and in CCR2-binding chemokines have been observed. However, the specific contribution of CCR2-dependent Gr1high monocytes is unknown. To investigate the impact of these monocytes during HUS, we injected a combination of LPS and shiga toxin into mice. We observed an impaired kidney function and elevated levels of the CCR2-binding chemokine CCL2 after shiga toxin/LPS- injection, thus suggesting Gr1high monocyte infiltration into the kidney. Indeed, the number of Gr1high monocytes was strongly increased one day after HUS induction. Moreover, these cells expressed high levels of CD11b suggesting activation after tissue entry. Non-invasive PET-MR imaging revealed kidney injury mainly in the kidney cortex and this damage coincided with the detection of Gr1high monocytes. Lack of Gr1high monocytes in Ccr2-deficient animals reduced neutrophil gelatinase-associated lipocalin and blood urea nitrogen levels. Moreover, the survival of Ccr2-deficient animals was significantly improved. Conclusively, this study demonstrates that CCR2-dependent Gr1high monocytes contribute to the kidney injury during HUS and targeting these cells is beneficial during this disease.

Shiga Toxin Mediated Neurologic Changes in Murine Model of Disease.

Seizures and neurologic involvement have been reported in patients infected with Shiga toxin (Stx) producing E. coli, and hemolytic uremic syndrome (HUS) with neurologic involvement is associated with more severe outcome. We investigated the extent of renal and neurologic damage in mice following injection of the highly potent form of Stx, Stx2a, and less potent Stx1. As observed in previous studies, Stx2a brought about moderate to acute tubular necrosis of proximal and distal tubules in the kidneys. Brain sections stained with hematoxylin and eosin (H&E) appeared normal, although some red blood cell congestion was observed. Microglial cell responses to neural injury include up-regulation of surface-marker expression (e.g., Iba1) and stereotypical morphological changes. Mice injected with Stx2a showed increased Iba1 staining, mild morphological changes associated with microglial activation (thickening of processes), and increased microglial staining per unit area. Microglial changes were observed in the cortex, hippocampus, and amygdala regions, but not the nucleus. Magnetic resonance imaging (MRI) of Stx2a-treated mice revealed no hyper-intensities in the brain, although magnetic resonance spectroscopy (MRS) revealed significantly decreased levels of phosphocreatine in the thalamus. Less dramatic changes were observed following Stx1 challenge. Neither immortalized microvascular endothelial cells from the cerebral cortex of mice (bEnd.3) nor primary human brain microvascular endothelial cells were found to be susceptible to Stx1 or Stx2a. The lack of susceptibility to Stx for both cell types correlated with an absence of receptor expression. These studies indicate Stx causes subtle, but identifiable changes in the mouse brain.

Development of a Mouse Model of Shiga Toxin 2 (Stx2) Intoxication for Testing Therapeutic Agents Against Hemolytic Uremic Syndrome (HUS).

BACKGROUND: Hemolytic Uremic Syndrome (HUS) caused by infections with Shiga toxin (Stx)-producing E. coli is a life-threatening complication characterized by acute renal failure, thrombocytopenia and hemolytic anemia. Stx is the main pathogenic factor. Therefore, the mouse model by intravenous administration of a single lethal dose of Stx is often used to explore its pathogenic mechanisms. OBJECTIVE: The aim of this work was to develop an alternative mouse model of Stx type 2 (Stx2) intoxication to evaluate new therapeutic strategies. METHODS AND RESULTS: One lethal dose of Stx2 was divided in four daily doses. We observed a dose-dependent toxicity characterized by neutrophilia, leukocytopenia and renal damage. Most importantly, we demonstrated that the polyclonal anti-Stx2 serum was able to protect mice from fatal evolution even when administered together the third dose of Stx2. CONCLUSION: This model would provide an advantage for evaluation of therapeutic strategies. Furthermore, the results presented herein suggest that appropriate treatment with anti-Stx2 agents following the appearance of initial clinical signs may block the ongoing outcome or may alleviate disease in patients who have just been diagnosed with HUS. However, the delay in the onset of therapy would be unsafe.

Shiga Toxin (Stx) Type 1a Reduces the Oral Toxicity of Stx Type 2a.

BACKGROUND: Shiga toxin (Stx) is the primary virulence factor of Stx-producing Escherichia coli (STEC). STEC can produce Stx1a and/or Stx2a, which are antigenically distinct. However, Stx2a-producing STEC are associated with more severe disease than strains producing both Stx1a and Stx2a. METHODS AND RESULTS: To address the hypothesis that the reason for the association of Stx2a with more severe disease is because Stx2a crosses the intestinal barrier with greater efficiency that Stx1a, we covalently labeled Stx1a and Stx2a with Alexa Fluor 750 and determined the ex vivo fluorescent intensity of murine systemic organs after oral intoxication. Surprisingly, both Stxs exhibited similar dissemination patterns and accumulated in the kidneys. We next cointoxicated mice to determine whether Stx1a could impede Stx2a. Cointoxication resulted in increased survival and an extended mean time to death, compared with intoxication with Stx2a only. The survival benefit was dose dependent, with the greatest effect observed when 5 times more Stx1a than Stx2a was delivered, and was amplified when Stx1a was delivered 3 hours prior to Stx2a. Cointoxication with an Stx1a active site toxoid also reduced Stx2a toxicity. CONCLUSIONS: These studies suggest that Stx1a reduces Stx2a-mediated toxicity, a finding that may explain why STEC that produce only Stx2a are associated with more severe disease than strains producing Stx1a and Stx2a.

Protective efficacy of a genetic subunit bacterin against edema disease of swine in mice.

The exotoxin SLT-IIeB from the Escherichia coli Ee strain was expressed in E. coli, and the recombinant protein was purified, mixed with the Ee strain, then emulsified with oil-emulsion adjuvants to obtain a mixed subunit bacterin. Groups of Kunming mice were immunized at weeks 0 and 2, and challenged intraperitoneally with the Ee strain at week 4. Antibodies were detected by ELISA and an agglutination test. After the second immunization, the antibody level increased and the rate of immune protection against the Ee strain was 70 and 91.7% in the subunit bacterin and bacterin groups, respectively. Therefore, the mixed subunit bacterin provided good protection against the homologous Ee strain, which provides a basis for further research, into high-efficacy vaccines against porcine edema disease.

Effects of Shiga toxin type 2 on maternal and fetal status in rats in the early stage of pregnancy.

Shiga toxin type 2 (Stx2), a toxin secreted by Shiga toxin-producing Escherichia coli (STEC), could be one of the causes of maternal and fetal morbimortality not yet investigated. In this study, we examined the effects of Stx2 in rats in the early stage of pregnancy. Sprague-Dawley pregnant rats were intraperitoneally (i.p.) injected with sublethal doses of Stx2, 0.25 and 0.5 ng Stx2/g of body weight (bwt), at day 8 of gestation (early postimplantation period of gestation). Maternal weight loss and food and water intake were analyzed after Stx2 injection. Another group of rats were euthanized and uteri were collected at different times to evaluate fetal status. Immunolocalization of Stx2 in uterus and maternal kidneys was analyzed by immunohistochemistry. The presence of Stx2 receptor (globotriaosylceramide, Gb3) in the uteroplacental unit was observed by thin layer chromatography (TLC). Sublethal doses of Stx2 in rats caused maternal weight loss and pregnancy loss. Stx2 and Gb3 receptor were localized in decidual tissues. Stx2 was also immunolocalized in renal tissues. Our results demonstrate that Stx2 leads to pregnancy loss and maternal morbidity in rats in the early stage of pregnancy. This study highlights the possibility of human pregnancy loss and maternal morbidity mediated by Stx2.

A translational murine model of sub-lethal intoxication with Shiga toxin 2 reveals novel ultrastructural findings in the brain striatum.

Infection by Shiga toxin-producing Escherichia coli causes hemorrhagic colitis, hemolytic uremic syndrome (HUS), acute renal failure, and also central nervous system complications in around 30% of the children affected. Besides, neurological deficits are one of the most unrepairable and untreatable outcomes of HUS. Study of the striatum is relevant because basal ganglia are one of the brain areas most commonly affected in patients that have suffered from HUS and since the deleterious effects of a sub-lethal dose of Shiga toxin have never been studied in the striatum, the purpose of this study was to attempt to simulate an infection by Shiga toxin-producing E. coli in a murine model. To this end, intravenous administration of a sub-lethal dose of Shiga toxin 2 (0.5 ηg per mouse) was used and the correlation between neurological manifestations and ultrastructural changes in striatal brain cells was studied in detail. Neurological manifestations included significant motor behavior abnormalities in spontaneous motor activity, gait, pelvic elevation and hind limb activity eight days after administration of the toxin. Transmission electron microscopy revealed that the toxin caused early perivascular edema two days after administration, as well as significant damage in astrocytes four days after administration and significant damage in neurons and oligodendrocytes eight days after administration. Interrupted synapses and mast cell extravasation were also found eight days after administration of the toxin. We thus conclude that the chronological order of events observed in the striatum could explain the neurological disorders found eight days after administration of the toxin.

Intraperitoneal administration of Shiga toxin 2 induced neuronal alterations and reduced the expression levels of aquaporin 1 and aquaporin 4 in rat brain.

Shiga toxin-producing Escherichia coli produces watery and hemorrhagic diarrhea, and hemolytic uremic syndrome (HUS) characterized by thrombocytopenia, microangiopathic hemolytic anemia, and acute renal failure. Central nervous system (CNS) complications are observed in around 30% of infant population with HUS. Common signs of severe CNS involvement leading to death include seizures, alteration of consciousness, hemiparesis, visual disturbances, and brain stem symptoms. The purpose of the present work was to study the effects of Shiga toxin 2 (Stx2) in the brain of rats intraperitoneally (i.p.) injected with a supernatant from recombinant E. coli expressing Stx2 (sStx2). Neurological alterations such as postural and motor abnormalities including lethargy, abnormal walking, and paralysis of hind legs, were observed in this experimental model of HUS in rats. Neuronal damage, as well as significant decrease in aquaporin 1 (AQP1) and aquaporin 4 (AQP4) expression levels were observed in the brain of rats, 2 days after sStx2 injection, compared to controls. Downregulation of aquaporin protein levels, and neuronal alterations, observed in brain of rats injected with sStx2, may be involved in edema formation and in neurological manifestations characteristic of HUS.

Protection of piglets against Edema disease by maternal immunization with Stx2e toxoid.

Edema disease (ED) in piglets is caused by Shiga toxin Stx2e-producing Escherichia coli. We show that a genetically disarmed Stx2e toxoid is a safe antigen that generates antiserum protecting piglets against the Stx2e toxin. Immunization of suckling piglets with the Stx2e toxoid was safe, had no adverse effects on growth of the piglets, and resulted in effective prevention of edema disease clinical symptoms after challenge with the Stx2e toxin. Our data showed that maternal immunity against the Stx2e toxoid can be transmitted from the vaccinated sows to the piglets via the colostrum. Very high levels of Stx2e-specific serum antibodies persisted in these piglets until 1 month postweaning, bridging the critical period in which the weaned piglets are most susceptible to edema infection. Challenge with Stx2e toxin resulted in clinical signs of edema disease and death of all control piglets from nonimmunized sows, whereas none of the piglets from immunized sows developed clinical signs of ED.

Mouse model for hemolytic uremic syndrome induced by outer membrane vesicles of Escherichia coli O157:H7.

Hemolytic uremic syndrome (HUS) is characterized by acute renal failure in children and is typically complicated with thrombocytopenia and hemolytic anemia. Although mouse models of HUS have been evaluated using Shiga toxin (STx) combined with or without lipopolysaccharide (LPS), no HUS model has been tested using purified outer membrane vesicles (OMVs) from STx-producing Escherichia coli (STEC) O157:H7. Accordingly, we investigated whether OMVs of STEC O157:H7 conveying STx2 and LPS can cause HUS-like symptoms in mice inoculated intraperitoneally. Three types of OMVs differing in LPS acylation status and STx2 amount were used to compare their ability to induce HUS-like symptoms. Native OMVs (nOMV) with fully hexa-acylated LPS caused HUS-like symptoms at 72-96 h after dually divided injections of 1 µg nOMV per animal. However, elevated doses of modified OMVs (mOMV) carrying mainly penta-acylated LPS were required to induce similar HUS signs. Moreover, mitomycin-C-induced OMVs (mcOMV) that were enriched with STx2 induced HUS-like symptoms similar to those of nOMV at the same dose. These results suggest that the OMVs of STEC O157:H7 potentiated with STx2 and fully hexa-acylated LPS can be utilized for inducing HUS-like symptoms in mice and could be the causative material involved in the development of HUS.

Ingested Shiga toxin 2 (Stx2) causes histopathological changes in kidney, spleen, and thymus tissues and mortality in mice.

The Shiga toxin (Stx)-producing bacterial strain, Escherichia coli O157:H7, colonizes the distal small intestine and the colon, initiating serious illness, including hemolytic-uremic syndrome (HUS), characterized by microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. Although intravenous administration of purified Stx to primates has been able to reproduce the features of HUS, it has not been conclusively established as to whether ingestion of Stx alone without the bacterium poses a potential health risk. To help answer this question, in this study, we fed Shiga toxin 2 (Stx2) directly into the stomachs of mice via gavage. Our data show that ingestion of Stx2 at a concentration of 50 µg/mouse induces weight loss and kills the mice at 3-5 days post-gavage. Additional studies revealed that the toxin retains activity at low pH, that its activity is neutralized by treatment with toxin-specific antibody, and that about 1% of the fed toxin is absorbed into the blood circulation. Lethality by intraperitoneal (IP) injection of Stx2 occurred at much lower doses than by ingestion. Detailed histopathological evaluation of stained tissues by light microscopy revealed severe histopathological changes in kidneys, spleen, and thymus but not in the pancreas, lymph nodes, heart, lungs, trachea, esophagus, stomach, duodenum, jejunum, ileum, cecum, and colon. The pathological changes in the kidney appeared similar to those seen in humans with HUS. The cited data suggest that (a) most but not all of the toxin is inactivated in the digestive tract, (b) part of the oral-ingested toxin is absorbed from the digestive tract into the circulation, (c) enough active toxin reaches susceptible organs to induce damage, and (d) Stx2 in the absence of toxin-producing bacteria can be harmful to mice. The results are clinically relevant for food safety because we also found that heat treatments (pasteurization) that destroy bacteria did not inactivate the heat-resistant toxin produced and secreted by the bacteria.

Fusion expression and immunogenicity of EHEC EspA-Stx2Al protein: implications for the vaccine development.

Shiga toxin 2 (Stx2) is a major virulence factor for enterohemorrhagic Escherichia coli (EHEC), which is encoded by lambda lysogenic phage integrated into EHEC chromosome. Stx2Al, Al subunit of Stx2 toxin has gathered extensive concerns due to its potential of being developed into a vaccine candidate. However, the substantial progress is hampered in part for the lack of a suitable in vitro expression system. Here we report use of the prokaryotic system pET-28a::espA-Stx2Al/BL21 to carry out the fusion expression of Stx2Al which is linked to E. coli secreted protein A (EspA) at its N-terminus. Under the IPTG induction, EspA-Stx2Al fusion protein in the form of inclusion body was obtained successfully, whose expression level can reach about 40% of total bacterial protein at 25 degrees C, much higher than that at 37 degrees C. Western blot test suggested the refolded fusion protein is of excellent immuno-reactivity with both monoclonal antibodies, which are specific to EspA and Stx2Al, respectively. Anti-sera from Balb/c mice immunized with the EspA-Stx2Al fusion protein were found to exhibit strong neutralization activity and protection capability in vitro and in vivo. These data have provided a novel feasible method to produce Stx2Al in large scale in vitro, which is implicated for the development of multivalent subunit vaccines candidate against EHEC 0157:H7 infections.

Intraperitoneal administration of shiga toxin type 2 in rats in the late stage of pregnancy produces premature delivery of dead fetuses.

Infection associated with Shiga toxin-producing Escherichia coli (STEC) and subsequent Hemolytic-Uremic Syndrome (HUS) have become relevant in public health since STEC is considered as one of the most important emergent pathogens. STEC infection may either be asymptomatic or begin with watery diarrhea associated with hemorrhagic colitis and HUS. The major virulence factor of STEC is Shiga toxin type 1 or 2 (Stx1, Stx2) although strains that express only Stx2 are highly prevalent. Up to now, it has not been established whether STEC infection affect pregnant women. In this study, we evaluated the effect of Stx2 on maternal lethality, fetal status and delivery time by injecting Stx2 in rats in the late stage of pregnancy. Stx2 induced fetal resorption, placental abruption, intrauterine hemorrhage and fetal death at 1-2 days post-injection in a dose-dependent manner. With 2ng Stx2/g body weight, placentas and fetuses presented extensive necrotic areas, while uteri and kidneys showed normal histology. Immunolocalization of Stx2 was observed in placentas and fetuses. With 4 and 6ng Stx2/g body weight maternal death was also observed. Those rats that survived after Stx2-treatment were able to become pregnant and deliver normal pups at term. Our results show, for the first time, that the preterm labor with fetal death observed in treated rats may be a consequence of the action of Stx2 on the feto-maternal unit. Although there are no reports of Stx2 effects in human pregnancy, we speculate that STEC infections could be one of the causes not yet determined of fetal morbimortality.

Intracerebroventricular administration of Shiga toxin type 2 altered the expression levels of neuronal nitric oxide synthase and glial fibrillary acidic protein in rat brains.

Shiga toxin (Stx) from enterohemorrhagic Escherichia coli (STEC) is the main cause of hemorrhagic colitis which may derive into Hemolytic Uremic Syndrome (HUS) and acute encephalopathy, one of the major risk factors for infant death caused by the toxin. We have previously demonstrated that intracerebroventricular administration of Stx2 causes neuronal death and glial cell damage in rat brains. In the present work, we observed that the intracerebroventricular administration of Stx2 increased the expression of glial fibrillary acidic protein (GFAP) leading to astrogliosis. Confocal microscopy showed reactive astrocytes in contact with Stx2-containing neurons. Immunocolocalization of increased GFAP and Stx2 in astrocytes was also observed. This insult in the brain was correlated with changes in the expression and activity of neuronal nitric oxide synthase (nNOS) by using the NADPH-diaphorase histochemical technique (NADPH-d HT). A significant decrease in NOS/NADPH-d-positive neurons and NOS/NADPH-d activity was observed in cerebral cortex and striatum, whereas an opposite effect was found in the hypothalamic paraventricular nucleus. We concluded that the i.c.v. administration of Stx2 promotes a typical pattern of brain injury showing reactive astrocytes and an alteration in the number and activity of nNOS/NADPH-d. According to the functional state of nNOS/NADPH-d and to brain cell morphology data, it could be inferred that the i.c.v. administration of Stx2 leads to either a neurodegenerative or a neuroprotective mechanism in the affected brain areas. The present animal model resembles the encephalopathy developed in Hemolytic Uremic Syndrome (HUS) patients by STEC intoxication.

Shiga-toxin-induced firm adhesion of human leukocytes to endothelium is in part mediated by heparan sulfate.

BACKGROUND: Shiga toxin (Stx) is the main pathogenic factor in the haemolytic-uraemic syndrome (HUS). Stx damages the renal endothelium, which leads to inflammation and coagulation. Endothelial heparan sulfate proteoglycans (HSPG), and heparan sulfate in particular, play an important role in the inflammatory process by acting as a ligand for l-selectin. Furthermore, leukocytes are able to interact with chemokines bound to HSPG (examples are IL-8, RANTES and MCP-1). This leads to an activation of integrins on leukocytes and results in more stable leukocyte-endothelial wall adhesion. In this study, we have evaluated the effect of a subtoxic dose of Stx1 and Stx2 on the HSPG and its role in adhesion of leukocytes. METHODS: Primary human umbilical venous endothelial cells (HUVEC) and primary human glomerular microvascular endothelial cells (GMVEC) were incubated for 24 h with a subtoxic dose of Stx1 or Stx2. Then, cells were treated with heparan sulfate-degrading enzyme heparitinase I or left untreated, followed by determination of binding leukocytes to endothelial cells in a parallel plate flow chamber. RESULTS: In both cell types, Stx increased the amount of firmly adherent leukocytes. After removal of endothelial heparan sulfate, the number of adhering leukocytes decreased. CONCLUSIONS: HSPG have a distinctive role in adhesion of leukocytes to endothelial cells stimulated by a subtoxic dose of Stx.

Intracerebroventricular administration of Shiga toxin type 2 induces striatal neuronal death and glial alterations: an ultrastructural study.

Shiga toxin (Stx) from enterohemorrhagic Escherichia coli (STEC) is the main cause of hemorrhagic colitis which may derive to hemolytic-uremic syndrome (HUS). HUS is characterized by acute renal failure, thrombocytopenia and microangiopathic hemolytic anemia. Mortality in the acute stage has been lower than 5% of total affected argentine children with endemic HUS. Common signs of severe CNS involvement leading to death included seizures, alteration of consciousness, hemiparesis, visual disturbances, and brainstem symptoms. The main purpose of the present work was to study the direct involvement of Stx2 in brain cells by intracerebroventricular (i.c.v.) administration of Stx2. Immunodetection of Stx2 was confirmed by immunoelectron cytochemistry in different subsets and compartments of affected caudate putamen cells of corpus striatum. Transmission electron microscopy (TEM) studies revealed apoptotic neurons, glial ultrastructural alterations and demyelinated fibers. The i.c.v. microinfusion was applied for the first time in rats to demonstrate the direct action of Stx2 in neurons and glial cells. The toxin may affect brain neuroglial cells without the involvement of proinflammatory or systemic neurotoxic elements.

Experimental infection of enterotoxemic Escherichia coli associated with porcine edema disease and its pathologic characteristics in the intestine.

Edema disease (ED) has become frequent in Japan, but no effective method for experimental infection has been developed. We report here the use of a capsule that resistant against gastric digestion to induce the ED in piglets. Four 21-day-old piglets were used. Shiga toxin 2e-producing Escherichia coli (STEC) cell pellet was encapsulated and administered orally. Two pigs received 1.0x10(10) CFU for two days, and the others received 3.9x10(8) CFU for three days. The high-dose group caused the typical clinical ED signs (palpebral edema or neurologic impairment). Eosinophil infiltration, swollen lymphoid follicles, and edema were observed in the ileum. The kidney had the thrombus in the glomerulus.

Oral immunization with a shiga toxin B subunit: rotavirus NSP4(90) fusion protein protects mice against gastroenteritis.

A fusion protein containing the shiga toxin-1 B subunit (STB) linked to a 90 amino acid peptide (aa residues 86--175) from simian rotavirus (SA--11) nonstructural protein NSP4 was synthesized in Escherichia coli. Mice orally inoculated with 60 microg of STB::NSP4(90) fusion protein per dose generated higher humoral and intestinal antibody titers than mice inoculated with 30 microg of NSP4 alone. Serum anti-NSP4 IgG2a isotype titers were substantially greater than IgG1 titers, suggesting a dominant Th1 immune response. ELISA measurement of cytokines secreted from splenocytes isolated from immunized mice confirmed the STB::NSP4(90) fusion protein stimulation of a strong Th1 cell mediated immune response. Diarrhea in SA-11 rotavirus challenged neonates suckling from STB::NSP4 immunized dams was significantly reduced in severity and duration in comparison with virus challenged neonates from unimmunized mice. Together, our experiments demonstrate for the first time that the shiga toxin B subunit provides ligand mediated delivery of virus antigens to the gut-associated lymphoid tissues for enhanced stimulation of humoral and cellular responses against rotavirus gastroenteritis.

Shiga toxin 2 induces macrophage-granulocyte colonies from human bone marrow and cord blood stem cells.

Addition of Shiga toxin 2 to human bone marrow or cord blood cell culture induced macrophage-granulocyte colonies. Although Shiga toxin 2 alone induced colonies mainly composed of macrophages, it induced colonies mainly consisting of granulocytes when combined with physiological doses of interleukin-1beta, granulocyte colony-stimulating factor, or stem cell factor with interleukin-3.