Fusion to chicken C3d enhances the immunogenicity of the M2 protein of avian influenza virus

Background

Infectious hematopoietic necrosis virus (IHNV) is the type species of the genus Novirhabdovirus, within the family Rhabdoviridae, infecting several species of wild and hatchery reared salmonids. Similar to other rhabdoviruses, IHNV has a linear single-stranded, negative-sense RNA genome of approximately 11,000 nucleotides. The IHNV genome encodes six genes; the nucleocapsid, phosphoprotein, matrix protein, glycoprotein, non-virion protein and polymerase protein genes, respectively. This study describes molecular characterization of the virulent IHNV strain 220-90, belonging to the M genogroup, and its phylogenetic relationships with available sequences of IHNV isolates worldwide.

Results

The complete genomic sequence of IHNV strain 220-90 was determined from the DNA of six overlapping clones obtained by RT-PCR amplification of genomic RNA. The complete genome sequence of 220-90 comprises 11,133 nucleotides (GenBank GQ413939) with the gene order of 3'-N-P-M-G-NV-L-5'. These genes are separated by conserved gene junctions, with di-nucleotide gene spacers. An additional uracil nucleotide was found at the end of the 5'-trailer region, which was not reported before in other IHNV strains. The first 15 of the 16 nucleotides at the 3'- and 5'-termini of the genome are complementary, and the first 4 nucleotides at 3'-ends of the IHNV are identical to other novirhadoviruses. Sequence homology and phylogenetic analysis of the glycoprotein genes show that 220-90 strain is 97% identical to most of the IHNV strains. Comparison of the virulent 220-90 genomic sequences with less virulent WRAC isolate shows more than 300 nucleotides changes in the genome, which doesn't allow one to speculate putative residues involved in the virulence of IHNV.

Conclusion

We have molecularly characterized one of the well studied IHNV isolates, 220-90 of genogroup M, which is virulent for rainbow trout, and compared phylogenetic relationship with North American and other strains. Determination of the complete nucleotide sequence is essential for future studies on pathogenesis of IHNV using a reverse genetics approach and developing efficient control strategies.     

The role of anti-malarial drugs in eliminating malaria

Background

Pregnant women develop protective anti-VSA IgG1 and IgG3 when infected by Plasmodium falciparum. The major target of IgG from serum of infected pregnant women is VAR2CSA.

Methods

In this study, ELISA was used to compare the level of VAR2CSA DBL5ε- specific IgG subclasses at enrolment and at delivery in a cohort of pregnant women in Senegal. All antibody measures were analysed in relation to placental infection according to parity.

Results

The results show an interaction between immune response to placental malaria and parity. A higher level of anti- DBL5ε- IgG3 at enrolment and a higher increase between enrolment and delivery were found in primigravidae who presented with uninfected placenta at delivery in comparison to those who presented with an infection of the placenta. However, high antibody level at delivery was associated with the infection of the placenta in multigravidae.

Conclusion

This high level of IgG3 in uninfected primigravidae suggests a protective role of these antibodies in this susceptible group, highlighting the importance of VAR2CSA in general and of some of its variants still to be defined, in the induction of protective immunity to pregnancy malaria.     

Role of PKCtheta in macrophage-mediated immune response to <Emphasis Type="Italic">Salmonella typhimurium</Emphasis> infection in mice

Background

The serine/threonine protein kinase C (PKC) theta has been firmly implicated in T cell-mediated immunity. Because its role in macrophages has remained undefined, we employed PKCtheta-deficient (PKCtheta ^?/?) mice in order to investigate if PKCtheta plays a role in macrophage-mediated immune responses during bacterial infections.

Results

Our results demonstrate that PKCtheta plays an important role in host defense against the Gram-negative, intracellular bacterium Salmonella typhimurium, as reflected both by markedly decreased survival and a significantly enhanced number of bacteria in spleen and liver of PKCtheta ^?/? mice, when compared to wild-type mice. Of note, albeit macrophages do not express detectable PKCtheta, PKCtheta mRNA expression was found to be profoundly upregulated during the first hours of lipopolysaccharide (LPS)/interferon-gamma (IFNgamma)-, but not IL-4-mediated cell polarization conditions in vitro. Mechanistically, despite expressing normal levels of classically activated macrophage (CAM) markers, PKCtheta-deficient CAMs expressed significantly higher levels of the anti-inflammatory cytokine IL-10 in vivo and in vitro when challenged with S. typhimurium or LPS/IFNgamma. Neutralization of IL-10 recovered immune control to S. typhimurium infection in PKCtheta-deficient macrophages.

Conclusions

Taken together, our data provide genetic evidence that PKCtheta promotes a potent pro-inflammatory CAM phenotype that is instrumental to mounting protective anti-bacterial immunity. Mechanistically, PKCtheta exerts a host-protective role against S. typhimurium infection, and acts as an essential link between TLR4/IFNgammaR signaling and selective suppression of the anti-inflammatory cytokine IL-10 at the onset of CAM differentiation in the course of a bacterial infection.

     

A new example of viral intein in Mimivirus

Background

Poxviruses encode a range of immunomodulatory genes to subvert or evade the challenges posed by the innate and adaptive immune responses. However, the inactivated poxviruses possessed immunostimulating capacity and were used as a prophylactic or metaphylactic application that efficiently reduced susceptibility to infectious diseases in different species. This fact is intensively studied in different genera of poxviruses. However, little is known about the basic mechanisms adopted by sheeppox virus (SPPV). SPPV causes an acute disease of sheep that recently, has been observed to reinfect its host in spite of vaccination.

Results

By injecting inactivated or attenuated sheeppox virus SPPV vaccine in adult male Swiss mice, SPPV was found to reduce macrophages' functions in a local event that occurs at the site of application 12 h after vaccine administration as indicated by increased level of IL-10 and decreased level of SOD from cultured peritoneal macrophages. In contrast increased levels of IL-12, and SOD activity from cultured splenic macrophages, lymphocyte response to PHA-P, and in-vivo response to T-dependant Ag were detected. These effects were observed in both attenuated and inactivated SPPV, but more prominent in attenuated one.

Conclusion

The results of this study help to elucidate, the phenomenon of existence natural SPPV infections in sheep instead of vaccination and the basic mechanisms responsible for the immunostimulating capacity of sheeppox virus. Locally, SPPV shows evidence for an immune escape mechanism that alleviates the host's immune response. Later and systemically, the virus protects the host from any fatal consequences of the immune system suppression.     

A SYBR Green RT-PCR assay in single tube to detect human and bovine noroviruses and control for inhibition

Background

A subset of the virus-specific CD8+ cytotoxic T lymphocytes (CTL) isolated from the lungs of mice infected with human respiratory syncytial virus (RSV) is impaired in the ability to secrete interferon γ (IFNγ), a measure of functionality. It was suggested that the impairment specifically suppressed the host cellular immune response, a finding that could help explain the ability of RSV to re-infect throughout life.

Results

To determine whether this effect is dependent on the virus, the route of infection, or the type of infection (respiratory, disseminated, or localized dermal), we compared the CTL responses in mice following intranasal (IN) infection with RSV or influenza virus or IN or intradermal (ID) infection with vaccinia virus expressing an RSV CTL antigen. The impairment was observed in the lungs after IN infection with RSV, influenza or vaccinia virus, and after a localized ID infection with vaccinia virus. In contrast, we observed a much higher percentage of IFNγ secreting CD8+ lymphocytes in the spleens of infected mice in every case.

Conclusion

The decreased functionality of CD8+ CTL is specific to the lungs and is not dependent on the specific virus, viral antigen, or route of infection.     

Comparative analysis of apoptotic changes in peripheral immune organs and lungs following experimental infection of piglets with highly pathogenic and classical porcine reproductive and respiratory syndrome virus

Background

Our previous studies have demonstrated that piglets infected with highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) may develop significant thymus atrophy, which related to thymocytes apoptosis. However, apart from that detected in the thymus, there are no reports describing cell apoptosis induced by HP-PRRSV infection. In this study, we analyzed comparatively the pathological changes, cell apoptosis and viral load in peripheral immune organs including tonsil, inguinal lymph nodes (ILNs) and spleen and lungs following experimental infection of piglets with HP-PRRSV HuN4 and classical PRRSV CH-1a.

Findings

HP-PRRSV HuN4 exhibited much stronger cell tropism than CH-1a in immune organs and lungs of piglets. HuN4 infection led to the serious injuries in tonsils, ILNs, spleens and lungs, especially apoptosis in these organs was significant.

Conclusions

HuN4 infection induced severe lesions (gross pathology, histopathology and cell apoptosis) in the peripheral immune organs and lungs of infected piglets. Large numbers of apoptotic cells in immune organs and lung induced by HuN4 may play a role in the pathogenesis of the HP-PRRS and the distinct injuries caused by HuN4 infection may be associated with the high mortality rate of HP-PRRS in pigs.     

Malaria prevention in north-eastern Tanzania: patterns of expenditure and determinants of demand at the household level

Background

In sub-Saharan Africa, preschool children represent the population most vulnerable to malaria and malnutrition. It is widely recognized that malnutrition compromises the immune function, resulting in higher risk of infection. However, very few studies have investigated the relationship between malaria, malnutrition and specific immunity. In the present study, the anti- Plasmodium falciparum IgG antibody (Ab) response was evaluated in children according to the type of malnutrition.

Methods

Anthropometric assessment and blood sample collection were carried out during a cross-sectional survey including rural Senegalese preschool children. This cross-sectional survey was conducted in July 2003 at the onset of the rainy season. Malnutrition was defined as stunting (height-for-age <-2 z-scores) or wasting (weight-for-height <-2 z-scores). The analysis was performed on all malnourished children in July (n = 161, either stunted, n = 142 or wasted, n = 19), pair-matched to well-nourished controls. The IgG Ab response to P. falciparum whole extracts (schizont antigens) was assessed by ELISA in sera of the included children.

Results

Both the prevalence of anti-malarial immune responders and specific IgG Ab levels were significantly lower in malnourished children than in controls. Depending on the type of malnutrition, wasted children and stunted children presented a lower specific IgG Ab response than their respective controls, but this difference was significant only in stunted children (P = 0.026). This down-regulation of the specific Ab response seemed to be explained by severely stunted children (HAZ ≤ -2.5) compared to their controls (P = 0.03), while no significant difference was observed in mildly stunted children (-2.5 < HAZ <-2.0). The influence of child malnutrition on the specific anti- P. falciparum Ab response appeared to be independent of the intensity of infection.

Conclusion

Child malnutrition, and particularly stunting, may down-regulate the anti- P. falciparum Ab response, both in terms of prevalence of immune responders and specific IgG Ab levels. This study provides further evidence for the influence of malnutrition on the specific anti-malarial immune response and points to the importance of taking into account child malnutrition in malaria epidemiological studies and vaccine trials.     

Functional consequences of DECTIN-1 early stop codon polymorphism Y238X in rheumatoid arthritis

Introduction

We have previously demonstrated that ex vivo inhibition of costimulatory molecules on antigen-pulsed dendritic cells (DCs) can be useful for induction of antigen-specific immune deviation and suppression of autoimmune arthritis in the collagen induced arthritis (CIA) model. The current study evaluated a practical method of immune modulation through temporary systemic inhibition of the costimulatory molecule CD40.

Methods

Mice with collagen II (CII)-induced arthritis (CIA) were administered siRNA targeting the CD40 molecule. Therapeutic effects were evaluated by clinical symptoms, histopathology, Ag-specific T cell and B cell immune responses.

Results

Systemic administration of CD40-targeting siRNA can inhibit antigen-specific T cell response to collagen II, as well as prevent pathogenesis of disease in both a pre- and post-immunization manner in the CIA model. Disease amelioration was associated with suppression of Th1 cytokines, attenuation of antibody production, and upregulation of T regulatory cells.

Conclusions

These studies support the feasibility of transient gene silencing at a systemic level as a mechanism of resetting autoreactive immunity.     

Second site escape of a T20-dependent HIV-1 variant by a single amino acid change in the CD4 binding region of the envelope glycoprotein

Background

Many novel studies and therapies are possible with the use of human embryonic stem cells (hES cells) and their differentiated cell progeny. The hES cell derived CD34 hematopoietic stem cells can be potentially used for many gene therapy applications. Here we evaluated the capacity of hES cell derived CD34 cells to give rise to normal macrophages as a first step towards using these cells in viral infection studies and in developing novel stem cell based gene therapy strategies for AIDS.

Results

Undifferentiated normal and lentiviral vector transduced hES cells were cultured on S17 mouse bone marrow stromal cell layers to derive CD34 hematopoietic progenitor cells. The differentiated CD34 cells isolated from cystic bodies were further cultured in cytokine media to derive macrophages. Phenotypic and functional analyses were carried out to compare these with that of fetal liver CD34 cell derived macrophages. As assessed by FACS analysis, the hES-CD34 cell derived macrophages displayed characteristic cell surface markers CD14, CD4, CCR5, CXCR4, and HLA-DR suggesting a normal phenotype. Tests evaluating phagocytosis, upregulation of the costimulatory molecule B7.1, and cytokine secretion in response to LPS stimulation showed that these macrophages are also functionally normal. When infected with HIV-1, the differentiated macrophages supported productive viral infection. Lentiviral vector transduced hES cells expressing the transgene GFP were evaluated similarly like above. The transgenic hES cells also gave rise to macrophages with normal phenotypic and functional characteristics indicating no vector mediated adverse effects during differentiation.

Conclusion

Phenotypically normal and functionally competent macrophages could be derived from hES-CD34 cells. Since these cells are susceptible to HIV-1 infection, they provide a uniform source of macrophages for viral infection studies. Based on these results, it is also now feasible to transduce hES-CD34 cells with anti-HIV genes such as inhibitory siRNAs and test their antiviral efficacy in down stream differentiated cells such as macrophages which are among the primary cells that need to be protected against HIV-1 infection. Thus, the potential utility of hES derived CD34 hematopoietic cells for HIV-1 gene therapy can be evaluated.     

Low-molecular-weight heparin reduces hyperoxia-augmented ventilator-induced lung injury via serine/threonine kinase-protein kinase B

Introduction

Factors explaining the greater susceptibility of preterm infants to severe lower respiratory infections with respiratory syncytial virus (RSV) remain poorly understood. Fetal/newborn lambs are increasingly appreciated as a model to study key elements of RSV infection in newborn infants due to similarities in lung alveolar development, immune response, and susceptibility to RSV. Previously, our laboratory demonstrated that preterm lambs had elevated viral antigen and developed more severe lesions compared to full-term lambs at seven days post-infection. Here, we compared the pathogenesis and immunological response to RSV infection in lungs of preterm and full-term lambs.

Methods

Lambs were delivered preterm by Caesarian section or full-term by natural birth, then inoculated with bovine RSV (bRSV) via the intratracheal route. Seven days post-infection, lungs were collected for evaluation of cytokine production, histopathology and cellular infiltration.

Results

Compared to full-term lambs, lungs of preterm lambs had a heightened pro-inflammatory response after infection, with significantly increased MCP-1, MIP-1α, IFN-γ, TNF-α and PD-L1 mRNA. RSV infection in the preterm lung was characterized by increased epithelial thickening and periodic acid-Schiff staining, indicative of glycogen retention. Nitric oxide levels were decreased in lungs of infected preterm lambs compared to full-term lambs, indicating alternative macrophage activation. Although infection induced significant neutrophil recruitment into the lungs of preterm lambs, neutrophils produced less myeloperoxidase than those of full-term lambs, suggesting decreased functional activation.

Conclusions

Taken together, our data suggest that increased RSV load and inadequate immune response may contribute to the enhanced disease severity observed in the lungs of preterm lambs.     

Bacterial lipopolysaccharide induces apoptosis in the trout ovary

Background  

In mammals it is well known that infections can lead to alterations in reproductive function. As part of the innate immune response, a number of cytokines and other immune factors is produced during bacterial infection or after treatment with lipopolysaccharide (LPS) and acts on the reproductive system. In fish, LPS can also induce an innate immune response but little is known about the activation of the immune system by LPS on reproduction in fish. Therefore, we conducted studies to examine the in vivo and in vitro effects of lipopolysaccharide (LPS) on the reproductive function of sexually mature female trout.     

Sex differences in the inflammatory response of primary astrocytes to lipopolysaccharide

Background

Numerous neurological and psychiatric disorders show sex differences in incidence, age of onset, symptomatology or outcome. Astrocytes, one of the glial cell types of the brain, show sex differences in number, differentiation and function. Since astrocytes are involved in the response of neural tissue to injury and inflammation, these cells may participate in the generation of sex differences in the response of the brain to pathological insults. To explore this hypothesis, we have examined whether male and female astrocytes show a different response to an inflammatory challenge and whether perinatal testosterone influences this response.

Methods

Cortical astrocyte cultures were prepared from postnatal day 1 (one day after birth) male or female CD1 mice pups. In addition, cortical astrocyte cultures were also prepared from female pups that were injected at birth with 100 μg of testosterone propionate or vehicle. Cultures were treated for 5 hours with medium containing lipopolysaccharide (LPS) or with control medium. The mRNA levels of IL6, interferon-inducible protein 10 (IP10), TNFα, IL1β, Toll-like receptor 4 (TLR4), steroidogenic acute regulatory protein and translocator protein were assessed by quantitative real-time polymerase chain reaction. Statistical significance was assessed by unpaired t-test or by one-way analysis of variance followed by the Tukey post hoc test.

Results

The mRNA levels of IL6, TNFα and IL1β after LPS treatment were significantly higher in astrocytes derived from male or androgenized females compared to astrocytes derived from control or vehicle-injected females. In contrast, IP10 mRNA levels after LPS treatment were higher in astrocytes derived from control or vehicle-injected females than in those obtained from males or androgenized females. The different response of male and female astrocytes to LPS was due neither to differences in the basal expression of the inflammatory molecules nor to differences in the expression of the LPS receptor TLR4. In contrast, the different inflammatory response was associated with increased mRNA levels of translocator protein, a key steroidogenic regulator, in female astrocytes that were treated with LPS.

Conclusions

Male and female cortical astrocytes respond differentially to an inflammatory challenge and this may be predetermined by perinatal testosterone exposure.

     

Infection control and the significance of sputum and other respiratory secretions from adult patients with cystic fibrosis

Background

Although various hematologic abnormalities are seen in tuberculosis, immune thrombocytopenic purpura is a rare event.

Case Presentation

We report a case of a 29 year-old male who was presented with immune thrombocytopenia-induced hemoptysis, macroscopic hematuria and generalized petechiae. The patient was found to have clinical, microbiological and radiological evidence of active pulmonary tuberculosis. The immune thrombocytopenic purpura was successfully treated with anti-tuberculous drugs combined with corticosteroids and high dose immune globulin therapy.

Conclusion

Immune thrombocytopenic purpura can be one of the hematological manifestations of tuberculosis which has a global prevalence with increasing incidence secondary to HIV infection.     

Japanese encephalitis virus infection induces changes of mRNA profile of mouse spleen and brain

Background

Rotaviruses are the single most important cause of severe diarrhea in young children worldwide. The current study was conducted to assess whether colostrum containing rotavirus-specific antibodies (Gastrogard-R^®) could protect against rotavirus infection. In addition, this illness model was used to study modulatory effects of intervention on several immune parameters after re-infection.

Methods

BALB/c mice were treated by gavage once daily with Gastrogard-R^® from the age of 4 to 10 days, and were inoculated with rhesus rotavirus (RRV) at 7 days of age. A secondary inoculation with epizootic-diarrhea infant-mouse (EDIM) virus was administered at 17 days of age. Disease symptoms were scored daily and viral shedding was measured in fecal samples during the post-inoculation periods. Rotavirus-specific IgM, IgG and IgG subclasses in serum, T cell proliferation and rotavirus-specific delayed-type hypersensitivity (DTH) responses were also measured.

Results

Primary inoculation with RRV induced a mild but consistent level of diarrhea during 3-4 days post-inoculation. All mice receiving Gastrogard-R^® were 100% protected against rotavirus-induced diarrhea. Mice receiving both RRV and EDIM inoculation had a lower faecal-viral load following EDIM inoculation then mice receiving EDIM alone or Gastrogard-R^®. Mice receiving Gastrogard-R^® however displayed an enhanced rotavirus-specific T-cell proliferation whereas rotavirus-specific antibody subtypes were not affected.

Conclusions

Preventing RRV-induced diarrhea by Gastrogard-R^® early in life showed a diminished protection against EDIM re-infection, but a rotavirus-specific immune response was developed including both B cell and T cell responses. In general, this intervention model can be used for studying clinical symptoms as well as the immune responses required for protection against viral re-infection.