MV140 Mucosal Vaccine Induces Targeted Immune Response for Enhanced Clearance of Uropathogenic E. coli in Experimental Urinary Tract Infection.

MV140 is an inactivated whole-cell bacterial mucosal vaccine with proven clinical efficacy against recurrent urinary tract infections (UTIs). These infections are primarily caused by uropathogenic E. coli (UPEC) strains, which are unique in their virulence factors and remarkably diverse. MV140 contains a non-UPEC strain, suggesting that it may induce an immune response against different UPEC-induced UTIs in patients. To verify this, we experimentally evaluated the cellular and humoral responses to UTI89, a prototypical UPEC strain, in mice vaccinated with MV140, as well as the degree of protection achieved in a UPEC UTI89 model of acute cystitis. The results show that both cellular (Th1/Th17) and antibody (IgG/IgA) responses to UTI89 were induced in MV140-immunized mice. MV140 vaccination resulted in an early increased clearance of UTI89 viable bacteria in the bladder and urine following transurethral infection. This was accompanied by a highly significant increase in CD4+ T cells in the bladder and an increase in urinary neutrophils. Collectively, our results support that MV140 induces cross-reactive humoral and cellular immune responses and cross-protection against UPEC strains.

MV130 in the Prevention of Recurrent Respiratory Tract Infections: A Retrospective Real-World Study in Children and Adults.

Respiratory tract infections (RTIs) are among the most common and important problems in clinical medicine, making antibiotics the gold standard therapeutic option regardless of their frequent viral etiology. Their excessive and inappropriate use contributes to the rapid rise of antibiotic resistance and underscores the need for alternative strategies, especially when dealing with recurrent RTIs. Prevention is the ideal alternative, but specific vaccines targeting a wide range of respiratory pathogens are scarce. MV130 is a sublingual bacterial vaccine that induces trained immunity and provides non-specific protection against respiratory pathogens in various clinical settings according to the concept of TIbV (Trained Immunity-based Vaccine). A retrospective real-world study (RWS) was conducted to evaluate the annual incidence of RTIs and the consumption of antibiotics before and after the administration of MV130, using data sourced from the medical records of 599 patients (186 children and 413 adults) who suffered from recurrent RTIs. The median number of infectious episodes in children was significantly reduced by more than 70% from 5 episodes (interquartile range (IQR) 4.0-6.0) to 1 (IQR, 0.0-2.0) (p < 0.001) after MV130. Similarly, in adults, the median number of episodes before MV130 immunization was 5 (IQR, 4.0-6.0), which dropped by more than 80% to 1 (IQR, 0.0-1.0) during the year following MV130 immunization (p < 0.001). The median number of antibiotic courses also significantly decreased for both children and adults by over 80% (p < 0.001). This RWS showed that MV130 is an effective strategy for the prevention of respiratory infections and the reduction of associated antibiotic consumption.

MV140 mucosal bacterial vaccine improves uropathogenic E. coli clearance in an experimental model of urinary tract infection.

MV140 is a mucosal vaccine of inactivated whole bacteria (E. coli, K. pneumoniae, E. faecalis, P. vulgaris) with clinical efficacy against recurrent urinary tract infections (UTIs). Here, MV140 was evaluated in a murine model of acute uropathogenic E. coli (UPEC)-induced UTI using the UTI89 strain. MV140 vaccination resulted in UPEC clearance, concomitant with increased influx of myeloid cells in urine, CD4+ T cells in the bladder, and a systemic adaptive immune response to both MV140-containing E. coli and UTI89.

Do bacterial vaccines/adjuvants prevent wheezing episodes in children?

PURPOSE OF REVIEW: To discuss recently discovered mechanisms of action of some bacterial vaccines that may account for their clinical benefit in the prevention of recurrent wheezing and asthma exacerbations in infants and early childhood. RECENT FINDINGS: Trained immunity has been shown to confer innate immune cells with a quite long-term nonspecific protection against a broad spectrum of pathogens. Inducers of trained immunity include some bacterial vaccines. Trained immunity-based vaccines (TIbV) of bacterial origin have the capability to induce nonspecific responses to a variety of pathogens, including respiratory viruses, in addition to their nominal bacterial antigens. Clinical data, from epidemiological surveys to well designed randomized clinical trials, indicate that TIbV formulated with bacteria prevent respiratory tract infections of viral cause, such as those associated with recurrent wheezing or asthma exacerbation, in children. Administration of these vaccines by the mucosal route may be important for their outcome in respiratory infections. SUMMARY: Mucosal bacterial immunotherapy, including certain TIbV, confer protection against a broad spectrum of pathogens, such as viruses, through a mechanism mediated by trained immunity. Clinical studies on the use of these preparations against recurrent wheezing reflect these mechanistic effects. These findings open a new avenue for the development of new strategies for this condition.

Trained immunity induction by the inactivated mucosal vaccine MV130 protects against experimental viral respiratory infections.

MV130 is an inactivated polybacterial mucosal vaccine that confers protection to patients against recurrent respiratory infections, including those of viral etiology. However, its mechanism of action remains poorly understood. Here, we find that intranasal prophylaxis with MV130 modulates the lung immune landscape and provides long-term heterologous protection against viral respiratory infections in mice. Intranasal administration of MV130 provides protection against systemic candidiasis in wild-type and Rag1-deficient mice lacking functional lymphocytes, indicative of innate immune-mediated protection. Moreover, pharmacological inhibition of trained immunity with metformin abrogates the protection conferred by MV130 against influenza A virus respiratory infection. MV130 induces reprogramming of both mouse bone marrow progenitor cells and in vitro human monocytes, promoting an enhanced cytokine production that relies on a metabolic shift. Our results unveil that the mucosal administration of a fully inactivated bacterial vaccine provides protection against viral infections by a mechanism associated with the induction of trained immunity.

Sublingual Bacterial Vaccination Reduces Recurrent Infections in Patients With Autoimmune Diseases Under Immunosuppressant Treatment.

Introduction: Conventional or biologic disease-modifying anti-rheumatic drugs (DMARDs) are the mainstay of treatment for systemic autoimmune disease (SAD). Infectious complications are a major concern in their use. Objective: To evaluate the clinical benefit of sublingual mucosal polybacterial vaccines (MV130 and MV140), used to prevent recurrent respiratory and urinary tract infections, in patients with SAD and secondary recurrent infections following conventional or biologic DMARDs. Methods: An observational study in SAD patients with recurrent respiratory tract infections (RRTI) and/or recurrent urinary tract infections (RUTI) was carried out. All patients underwent mucosal (sublingual) vaccination with MV130 for RRTI or with MV140 for RUTI daily for 3 months. Clinical evaluation was assessed during 12 months of follow-up after the first dose, i.e., 3 months under treatment and 9 months once discontinued, and compared with the previous year. Results: Forty-one out of 55 patients completed 1-year follow-up. All patients were on either conventional or biologic DMARDs. A significant decrease in the frequency of RUTI (p<0.001), lower respiratory tract infections (LRTI) (p=0.009) and upper respiratory tract infections (URTI) (p=0.006) at 12-mo with respect to the previous year was observed. Antibiotic prescriptions and unscheduled medical visits decreased significantly (p<0.020) in all groups. Hospitalization rate also declined in patients with RRTI (p=0.019). The clinical benefit demonstrated was concomitant to a significant increase in both anti-S. pneumoniae IgA and IgG antibodies following MV130 vaccination. Conclusions: Sublingual polybacterial vaccines prevent recurrent infections in patients with SAD under treatment with immunosuppressant therapies, supporting a broad non-specific anti-infectious effect in these patients.

Bacterial Mucosal Immunotherapy with MV130 Prevents Recurrent Wheezing in Children: A Randomized, Double-Blind, Placebo-controlled Clinical Trial.

Rationale: Recurrent wheezing in children represents a severe public health concern. Wheezing attacks (WA), mainly associated with viral infections, lack effective preventive therapies. Objectives: To evaluate the efficacy and safety of mucosal sublingual immunotherapy based on whole inactivated bacteria (MV130) in preventing WA in children. Methods: A Phase 3 randomized, double-blind, placebo-controlled, parallel-group trial including a cohort of 120 children <3 years old with ⩾3 WA during the previous year was conducted. Children with a positive skin test to common aeroallergens in the area where the clinical trial was performed were excluded from the trial. Subjects received MV130 or placebo daily for 6 months. The primary endpoint was the number of WA within 1 year after the first dose comparing MV130 and placebo. Measurements and Main Results: There was a significant lower number of WA in MV130 versus the placebo group, 3.0 (interquartile range [IQR], 2.0-4.0) versus 5.0 (IQR, 3.0-7.0) (P < 0.001). As secondary outcomes, a decrease in the duration of WA and a reduction in symptoms and medication scores in the MV130 versus placebo group were found. No adverse events were reported related to the active treatment. Conclusions: Mucosal bacterial immunotherapy with MV130 shows safety and clinical efficacy against recurrent WA in children.Clinical trial registered with www.clinicaltrials.gov (NCT01734811).

Transcriptional profiling unveils type I and II interferon networks in blood and tissues across diseases.

Understanding how immune challenges elicit different responses is critical for diagnosing and deciphering immune regulation. Using a modular strategy to interpret the complex transcriptional host response in mouse models of infection and inflammation, we show a breadth of immune responses in the lung. Lung immune signatures are dominated by either IFN-γ and IFN-inducible, IL-17-induced neutrophil- or allergy-associated gene expression. Type I IFN and IFN-γ-inducible, but not IL-17- or allergy-associated signatures, are preserved in the blood. While IL-17-associated genes identified in lung are detected in blood, the allergy signature is only detectable in blood CD4+ effector cells. Type I IFN-inducible genes are abrogated in the absence of IFN-γ signaling and decrease in the absence of IFNAR signaling, both independently contributing to the regulation of granulocyte responses and pathology during Toxoplasma gondii infection. Our framework provides an ideal tool for comparative analyses of transcriptional signatures contributing to protection or pathogenesis in disease.

Von Hippel-Lindau Protein Is Required for Optimal Alveolar Macrophage Terminal Differentiation, Self-Renewal, and Function.

The rapid transit from hypoxia to normoxia in the lung that follows the first breath in newborn mice coincides with alveolar macrophage (AM) differentiation. However, whether sensing of oxygen affects AM maturation and function has not been previously explored. We have generated mice whose AMs show a deficient ability to sense oxygen after birth by deleting Vhl, a negative regulator of HIF transcription factors, in the CD11c compartment (CD11cΔVhl mice). VHL-deficient AMs show an immature-like phenotype and an impaired self-renewal capacity in vivo that persists upon culture ex vivo. VHL-deficient phenotype is intrinsic in AMs derived from monocyte precursors in mixed bone marrow chimeras. Moreover, unlike control Vhlfl/fl, AMs from CD11cΔVhl mice do not reverse pulmonary alveolar proteinosis when transplanted into Csf2rb-/- mice, demonstrating that VHL contributes to AM-mediated surfactant clearance. Thus, our results suggest that optimal AM terminal differentiation, self-renewal, and homeostatic function requires their intact oxygen-sensing capacity.

Trained Immunity-Based Vaccines: A New Paradigm for the Development of Broad-Spectrum Anti-infectious Formulations.

Challenge with specific microbial stimuli induces long lasting epigenetic changes in innate immune cells that result in their enhanced response to a second challenge by the same or unrelated microbial insult, a process referred to as trained immunity. This opens a new avenue in vaccinology to develop Trained Immunity-based Vaccines (TIbV), defined as vaccine formulations that induce training in innate immune cells. Unlike conventional vaccines, which are aimed to elicit only specific responses to vaccine-related antigens, TIbV aim to stimulate broader responses. As trained immunity is generally triggered by pattern recognition receptors (PRRs), TIbV should be formulated with microbial structures containing suitable PRR-ligands. The TIbV concept we describe here may be used for the development of vaccines focused to promote host resistance against a wide spectrum of pathogens. Under the umbrella of trained immunity, a broad protection can be achieved by: (i) increasing the nonspecific effector response of innate immune cells (e.g., monocyte/macrophages) to pathogens, (ii) harnessing the activation state of dendritic cells to enhance adaptive T cell responses to both specific and nonrelated (bystander) antigens. This capacity of TIbV to promote responses beyond their nominal antigens may be particularly useful when conventional vaccines are not available or when multiple coinfections and/or recurrent infections arise in susceptible individuals. As the set of PRR-ligands chosen is essential not only for stimulating trained immunity but also to drive adaptive immunity, the precise design of TIbV will improve with the knowledge on the functional relationship among the different PRRs. While the TIbV concept is emerging, a number of the current anti-infectious vaccines, immunostimulants, and even vaccine adjuvants may already fall in the TIbV category. This may apply to increase immunogenicity of novel vaccine design approaches based on small molecules, like those achieved by reverse vaccinology.

Human dendritic cells activated with MV130 induce Th1, Th17 and IL-10 responses via RIPK2 and MyD88 signalling pathways.

Recurrent respiratory tract infections (RRTIs) are the first leading cause of community- and nosocomial-acquired infections. Antibiotics remain the mainstay of treatment, enhancing the potential to develop antibiotic resistances. Therefore, the development of new alternative approaches to prevent and treat RRTIs is highly demanded. Daily sublingual administration of the whole heat-inactivated polybacterial preparation (PBP) MV130 significantly reduced the rate of respiratory infections in RRTIs patients, however, the immunological mechanisms of action remain unknown. Herein, we study the capacity of MV130 to immunomodulate the function of human dendritic cells (DCs) as a potential mechanism that contribute to the clinical benefits. We demonstrate that DCs from RRTIs patients and healthy controls display similar ex vivo immunological responses to MV130. By combining systems biology and functional immunological approaches we show that MV130 promotes the generation of Th1/Th17 responses via receptor-interacting serine/threonine-protein kinase-2 (RIPK2)- and myeloid-differentiation primary-response gene-88 (MyD88)-mediated signalling pathways under the control of IL-10. In vivo BALB/c mice sublingually immunized with MV130 display potent systemic Th1/Th17 and IL-10 responses against related and unrelated antigens. We elucidate immunological mechanisms underlying the potential way of action of MV130, which might help to design alternative treatments in other clinical conditions with high risk of recurrent infections.

Lung CD103+ dendritic cells restrain allergic airway inflammation through IL-12 production.

DCs are necessary and sufficient for induction of allergic airway inflammation. CD11b+ DCs direct the underlying Th2 immunity, but debate surrounds the function of CD103+ DCs in lung immunity and asthma after an allergic challenge. We challenged Batf3-/- mice, which lacked lung CD103+ DCs, with the relevant allergen house dust mite (HDM) as a model to ascertain their role in asthma. We show that acute and chronic HDM exposure leads to defective Th1 immunity in Batf3-deficient mice. In addition, chronic HDM challenge in Batf3-/- mice results in increased Th2 and Th17 immune responses and exacerbated airway inflammation. Mechanistically, Batf3 absence does not affect induction of Treg or IL-10 production by lung CD4+ T cells following acute HDM challenge. Batf3-dependent CD103+ migratory DCs are the main source of IL-12p40 in the mediastinal lymph node DC compartment in the steady state. Moreover, CD103+ DCs selectively increase their IL-12p40 production upon HDM administration. In vivo IL-12 treatment reverts exacerbated allergic airway inflammation upon chronic HDM challenge in Batf3-/- mice, restraining Th2 and Th17 responses without triggering Th1 immunity. These results suggest a protective role for lung CD103+ DCs to HDM allergic airway inflammation through the production of IL-12.

A proteasome inhibitor produced by Burkholderia pseudomallei modulates intracellular growth.

The NRPS/PKS cluster encodes the enzymes necessary for glidobactin synthesis it is partially conserved in various members of the Burkholderia genus including B. pseudomallei. In this study we have shown that the insertional inactivation or deletion of glbC in this cluster in B. pseudomallei could reduce the ability of the bacterium to survive or grow in murine macrophages or in human neutrophils. Exogenously added proteasome inhibitors were able to chemically complement the mutation. The insertional inactivation or deletion of glbC increased virulence in an acute model of infection in Balb/c or C57BL/6 mice but virulence in a chronic model of infection was similar to that of the wild type. Our findings contrast with the previous finding that inactivation of the glb gene cluster in B. pseudomallei strain 1026b resulted in marked attenuation, and provides evidence of differential roles for some genes in virulence of different strains of B. pseudomallei.

Immunisation with proteins expressed during chronic murine melioidosis provides enhanced protection against disease.

There is an urgent need for an effective vaccine against human disease caused by Burkholderia pseudomallei, and although a wide range of candidates have been tested in mice none provide high level protection. We considered this might reflect the inability of these vaccine candidates to protect against chronic disease. Using Q-RT PCR we have identified 6 genes which are expressed in bacteria colonising spleens and lungs of chronically infected mice. Three of the genes (BPSL1897, BPSL3369 and BPSL2287) have been expressed in Escherichia coli and the encoded proteins purified. We have also included BPSL2765, a protein known to induce immune responses associated with a reduced incidence of chronic/recurrent disease in humans. Immunisation of mice with a combination of these antigens resulted in the induction of antibody responses against all of the proteins. Compared with mice immunised with capsular polysaccharide or LolC protein, mice immunised with the combination of chronic stage antigens showed enhanced protection against experimental disease in mice.

The Blood Transcriptome of Experimental Melioidosis Reflects Disease Severity and Shows Considerable Similarity with the Human Disease.

Melioidosis, a severe human disease caused by the bacterium Burkholderia pseudomallei, has a wide spectrum of clinical manifestations ranging from acute septicemia to chronic localized illness or latent infection. Murine models have been widely used to study the pathogenesis of infection and to evaluate novel therapies or vaccines, but how faithfully they recapitulate the biology of human melioidosis at a molecular level is not known. In this study, mice were intranasally infected with either high or low doses of B. pseudomallei to generate either acute, chronic, or latent infection and host blood and tissue transcriptional profiles were generated. Acute infection was accompanied by a homogeneous signature associated with induction of multiple innate immune response pathways, such as IL-10, TREM1, and IFN signaling, largely found in both blood and tissue. The transcriptional profile in blood reflected the heterogeneity of chronic infection and quantitatively reflected the severity of disease. Genes associated with fibrosis and tissue remodeling, including matrix metalloproteases and collagen, were upregulated in chronically infected mice with severe disease. Transcriptional signatures of both acute and chronic melioidosis revealed upregulation of iNOS in tissue, consistent with the expression of IFN-γ, but also Arginase-1, a functional antagonist of the iNOS pathway, and was confirmed by immunohistochemistry. Comparison of these mouse blood datasets by pathway and modular analysis with the blood transcriptional signature of patients with melioidosis showed that many genes were similarly perturbed, including Arginase-1, IL-10, TREM1, and IFN signaling, revealing the common immune response occurring in both mice and humans.

The condition-dependent transcriptional landscape of Burkholderia pseudomallei.

Burkholderia pseudomallei (Bp), the causative agent of the often-deadly infectious disease melioidosis, contains one of the largest prokaryotic genomes sequenced to date, at 7.2 Mb with two large circular chromosomes (1 and 2). To comprehensively delineate the Bp transcriptome, we integrated whole-genome tiling array expression data of Bp exposed to >80 diverse physical, chemical, and biological conditions. Our results provide direct experimental support for the strand-specific expression of 5,467 Sanger protein-coding genes, 1,041 operons, and 766 non-coding RNAs. A large proportion of these transcripts displayed condition-dependent expression, consistent with them playing functional roles. The two Bp chromosomes exhibited dramatically different transcriptional landscapes--Chr 1 genes were highly and constitutively expressed, while Chr 2 genes exhibited mosaic expression where distinct subsets were expressed in a strongly condition-dependent manner. We identified dozens of cis-regulatory motifs associated with specific condition-dependent expression programs, and used the condition compendium to elucidate key biological processes associated with two complex pathogen phenotypes--quorum sensing and in vivo infection. Our results demonstrate the utility of a Bp condition-compendium as a community resource for biological discovery. Moreover, the observation that significant portions of the Bp virulence machinery can be activated by specific in vitro cues provides insights into Bp's capacity as an "accidental pathogen", where genetic pathways used by the bacterium to survive in environmental niches may have also facilitated its ability to colonize human hosts.

Role of RelA and SpoT in Burkholderia pseudomallei virulence and immunity.

Burkholderia pseudomallei is a Gram-negative soil bacterium and the causative agent of melioidosis, a disease of humans and animals. It is also listed as a category B bioterrorism threat agent by the U.S. Centers for Disease Control and Prevention, and there is currently no melioidosis vaccine available. Small modified nucleotides such as the hyperphosphorylated guanosine molecules ppGpp and pppGpp play an important role as signaling molecules in prokaryotes. They mediate a global stress response under starvation conditions and have been implicated in the regulation of virulence and survival factors in many bacterial species. In this study, we created a relA spoT double mutant in B. pseudomallei strain K96243, which lacks (p)ppGpp-synthesizing enzymes, and investigated its phenotype in vitro and in vivo. The B. pseudomallei ΔrelA ΔspoT mutant displayed a defect in stationary-phase survival and intracellular replication in murine macrophages. Moreover, the mutant was attenuated in the Galleria mellonella insect model and in both acute and chronic mouse models of melioidosis. Vaccination of mice with the ΔrelA ΔspoT mutant resulted in partial protection against infection with wild-type B. pseudomallei. In summary, (p)ppGpp signaling appears to represent an essential component of the regulatory network governing virulence gene expression and stress adaptation in B. pseudomallei, and the ΔrelA ΔspoT mutant may be a promising live-attenuated vaccine candidate.

Development of vaccines against burkholderia pseudomallei.

Burkholderia pseudomallei is a Gram-negative bacterium which is the causative agent of melioidosis, a disease which carries a high mortality and morbidity rate in endemic areas of South East Asia and Northern Australia. At present there is no available human vaccine that protects against B. pseudomallei, and with the current limitations of antibiotic treatment, the development of new preventative and therapeutic interventions is crucial. This review considers the multiple elements of melioidosis vaccine research including: (i) the immune responses required for protective immunity, (ii) animal models available for preclinical testing of potential candidates, (iii) the different experimental vaccine strategies which are being pursued, and (iv) the obstacles and opportunities for eventual registration of a licensed vaccine in humans.

Low-dose exposure of C57BL/6 mice to burkholderia pseudomallei mimics chronic human melioidosis.

Burkholderia pseudomallei is the etiological agent of human melioidosis, a disease with a broad spectrum of clinical manifestations ranging from fatal septicemia to chronic localized infection or asymptomatic latent infection. Most clinical and immunological studies to date have focused on the acute disease process; however, little is known about pathology and immune response in chronic melioidosis. Here, we have developed a murine model of chronic disease by challenging C57BL/6 mice intranasally with a low dose of B. pseudomallei and monitoring them up to 100 days postinfection. Bacterial burdens were heterogeneous in different animals at all time points, consistent with the spectrum of clinical severity observed in humans. Proinflammatory cytokines such as gamma interferon (IFN-γ), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), and tumor necrosis factor-α (TNF-α) were induced during chronic infection, and histopathological analysis showed features in common with human melioidosis. Interestingly, many of these features were similar to those induced by Mycobacterium tuberculosis in humans, such as development of a collagen cord that encapsulates the lesions, the presence of multinucleated giant cells, and granulomas with a caseous necrotic center, which may explain why chronic melioidosis is often misdiagnosed as tuberculosis. Our model now provides a relevant and practical tool to define the immunological features of chronic melioidosis and aid in the development of more effective treatment of this disease in humans.

Automatic quantification of histological studies in allergic asthma.

The evaluation of new therapies to treat allergic asthma makes frequent use of histological studies. Some of them are based on microscope observation of stained paraffin lung sections to quantify cellular infiltrate, an effect directly related to allergic processes. Currently, there is no software tool available for doing this quantification automatically. This paper presents a methodology and a software tool for the quantification of cellular infiltrate in lung tissue images in an allergic asthma mouse model. The image is divided into regions of equal size, which are then classified by means of a segmentation algorithm based on texture analysis. The classification uses three discriminant functions, built from parameters derived from the histogram and the co-occurrence matrix. These functions were calculated by means of a stepwise discriminant analysis on 79 samples from a training set. Results provided a correct classification of 96.8% on an independent test set of 251 samples labeled manually. Regression analysis showed a good agreement between automatic and manual methods. A reliable and easy to implement method has been developed to provide an automatic method for quantifying microscopy images of lung histological studies. Results showed similar accuracy to that provided by an expert, while allowing analyzing a much larger number of fields in a repeatable way.