Presence Of Virus-specific Antibodies Research Articles

#41: Mechanistic Immune Correlates of Protection Following Vaccination Against Rift Valley Fever Virus

Abstract Background Rift Valley fever virus (RVFV) is a zoonotic arbovirus endemic to large areas of Africa and the Arabian peninsula that causes a significant annual burden of disease in ruminant animals, and is associated with periodic outbreaks in human populations. RVFV causes a wide spectrum of disease in infected patients ranging from a self-limiting febrile illness to hepatitis, encephalitis, hemorrhagic fever, and death. Despite the significant economic and public health impact of RVFV, there are no currently available vaccines or therapeutics available for RVFV. Methods Previous work has established that vaccination with an attenuated strain of RVFV, RVFV-ΔNSs, confers protection against subsequent lethal challenge in a murine model of infection. To characterize mechanistic immune correlates of protection, we vaccinated C57BL/6 mice with RVFV-ΔNSs then depleted vaccinated mice of various memory lymphocyte subsets, including CD4+, CD8+, CD4+/CD8+ T-cells before challenging mice with a lethal dose of WT RVFV. To assess for the role of antibodies in providing protection, we depleted mice of their B cells prior to vaccination with RVFV-ΔNSs, confirmed the lack of RVFV specific antibodies, and then challenged them with lethal WT RVFV. To determine the minimal immunologic factors required for protection, we vaccinated a series of donor mice, transferred either immunoglobulins or splenocytes to naïve recipient mice, then challenged them with WT RVFV. Results Vaccination with RVFV-ΔNSs protected mice from subsequent challenge, and this protection was not abrogated by depletion of CD4+, CD8+, or combined CD4+ and CD8+ lymphocytes, suggesting that RVFV-specific T-cells are dispensable for protection in the presence of virus specific antibodies. Vaccinated mice depleted of CD20+ B-cells were also protected against WT challenge, suggesting that B-cells/antibodies are also dispensable for protection in the presence of an intact T-cell response. Passive transfer of antibodies from vaccinated mice to recipient mice protected them against subsequent WT challenge, demonstrating that antibodies are sufficient for protection. Adoptive transfer of total splenocytes protected mice against subsequent WT challenge, indicating that lymphocytes are also sufficient to mediate protection. Conclusions In this study we defined the mechanistic immune correlates of protection following vaccination with attenuated RVFV-ΔNSs against WT RVFV in a murine model. Here, we have shown that both humoral and cellular immunity are sufficient to mediate protection in recipient animals, and can compensate for each other when one is missing. These results further our understanding of the protective immunologic response to infection by RVFV, and will help inform further vaccine development efforts against this important emerging pathogen.

Persistent Roseoloviruses Infection in Adult Patients with Epilepsy.

Background: Human herpesviruses (HHV)-6A, HHV-6B and HHV-7 are considered to be involved in the pathogenesis of epilepsy, a common neurological disorder. The objective of this study was to determine the association of roseoloviruses infection with epilepsy. Methods: 53 epilepsy patients and 104 ordinary blood donors were analyzed to determine presence of virus-specific antibodies by enzyme-linked immunosorbent assay (ELISA) and immunofluorescence assay (IFA), genomic sequences, viral load and gene expression by polymerase chain reactions (PCRs) and restriction analysis, HHV-6 protein expression by IFA and level of cytokines by ELISA. Results: Roseoloviruses genomic sequences in DNA samples from whole blood were found in 86.8% of patients versus 54.8% of controls and active infection was revealed only in patients with epilepsy (19.6% of roseolovirus-positive patients). Significantly higher viral load and more frequent gene expression was detected in patients compared to the controls. HHV-6-encoded protein expression was demonstrated in 53.3% of patients with previously detected HHV-6 DNA. Changes in level of cytokines were determined in patients with elevated viral load compared to the patients without elevated viral loads and to the controls. Conclusions: Results on frequent active HHV-6 and HHV-7 infection in epilepsy patient’ peripheral blood indicate on possible involvement of these viruses in the disease development.

Flaviviruses at the territory of Serbia – present situation and challenges

The aim of this study is to summarize the data on the occurrence, presence and prevalence of some zoonotic flaviviruses, which have been actively circulating in the territory of the Republic of Serbia during past decade. The genus Flavivirus, family Flaviviridae,encompasses vector-transmitted viruses with zoonotic potential. The genus is comprised of more than 70 viruses transmitted to humans by mosquitoes and ticks. Some of those are associated only with human diseases (such as yellow fever and dengue fever), whereas others can cause diseases in both humans and animals. Animals are major reservoirs and primary hosts for the latter group. The virus transmission to other animal species or humans occurs via diverse vectors – mostly mosquitoes and ticks. A range of these virus species is widely distributed worldwide, especially in tropical and sub-tropical climatic zones. Some representatives of these species have only recently been identified at the territory of Republic of Serbia. This paper gives an evidence on the presence and distribution of West-Nile virus, tick-borne encephalitis (TBE) virus and Usutu virus that were conducted within the study in Serbia. The research pointed out the presence of recently detected West-Nile virus and Usutu virus as well as the current circulation of tick-borne encephalitis virus, for which only limited serological evidence on the presence of virus-specific antibodies in humans was available so far. In the aspect of public health, the full understanding of the importance of identification and prevalence of different flaviviruses still require further comprehensive entomologicalacarological, seroepidemiological, clinical and virological research.

Clinical Outcomes Associated with Expansion of Fcr-Gamma-Deficient Natural Killer Cells in Allogeneic Stem Cell Transplant Patients

Background Natural killer (NK) cells are a part of the innate immune system and play a major role in immunity against viral infections and malignancies. Recently a new subset of NK cells has been discovered that lacks FcRg transmembrane signaling adaptor. These FcRg-deficient NK cells have been termed FcR gamma-negative NK or g-NK cells. The g-NK cells display a robust reaction against CMV and other viral infections in the presence of virus-specific antibodies. Study Objectives A longitudinal, prospective study is being conducted to observe the generation of g-NK cells in allogeneic hematopoietic transplant (AHSCT) patients after reactivation of CMV. The potential effect of g-NK cells on immunity against CMV and other viral infections, graft versus host disease (GVHD) and disease-free survival is being studied. Methods Blood samples from healthy donors and AHSCT patients were obtained pre- and post-transplantation. Flow cytometry was used to identify g-NK cells. Clinical data regarding CMV viremia, viral infections, GVHD and tumor free survival was collected and analyzed for 55 AHSCT patients. Results Of the 55 AHSCT patients, 49 were exposed to CMV and 13/49 (26%) generated g-NK cells. Subjects who were never exposed to CMV did not generate g-NK cells. Among patients with g-NK cell expansion, a higher incidence of Epstein-Barr virus (30.77% vs 19.05%) and adenovirus infections (7.69% vs 0%) was noted. Conversely, a trend towards decreased incidence of BK-virus infection (0% vs 11.9%) and GVHD (61.54% vs 71.43%) was observed. It is premature to comment on survival rate at 180 and 360 days. Conclusions G-NK cells appear in AHSCT recipients in response to CMV exposure. The response to some other common viral infections is variable in subjects with g-NK cell expansion. A slight decrease in incidence of GVHD has been observed in this population.

Development of the Abbott RealTime ZIKA assay for the qualitative detection of Zika virus RNA from serum, plasma, urine, and whole blood specimens using the m2000 system

Zika virus is an arthropod-borne flavivirus that has rapidly developed into a world-wide concern. Discovered in 1947, the virus was relatively obscure until an outbreak occurred in 2007 in the Yap islands and spread eventually to the Americas in 2015. Only 20% of patients infected with Zika virus develop symptoms. However, there can be serious consequences of infection including birth defects in developing fetuses and links to Guillain-Barré syndrome. The swift rise in infections has necessitated the development of diagnostic tests for both the detection of viral RNA and the presence of virus-specific antibodies. Abbott has developed a dual target RT-PCR assay for the detection of Zika virus RNA within serum, plasma, whole blood, and urine using the automated m2000 system for sample extraction to result reporting. The Abbott RealTime ZIKA assay has a limit of detection of 30 copies per mL in serum, 40 copies per mL in plasma and urine, and 120 copies per mL in whole blood and demonstrates high specificity against challenges from closely related infectious agents.

THU-264 - Discovery of a Novel Human Pegivirus in Blood Associated with Hepatitis C Virus Co-Infection

Hepatitis C virus (HCV) and human pegivirus (HPgV), formerly GBV-C, are the only known human viruses in the Hepacivirus and Pegivirus genera, respectively, of the family Flaviviridae. We present the discovery of a second pegivirus, provisionally designated human pegivirus 2 (HPgV-2), by next-generation sequencing of plasma from an HCV-infected patient with multiple bloodborne exposures who died from sepsis of unknown etiology. HPgV-2 is highly divergent, situated on a deep phylogenetic branch in a clade that includes rodent and bat pegiviruses, with which it shares <32% amino acid identity. Molecular and serological tools were developed and validated for high-throughput screening of plasma samples, and a panel of 3 independent serological markers strongly correlated antibody responses with viral RNA positivity (99.9% negative predictive value). Discovery of 11 additional RNA-positive samples from a total of 2440 screened (0.45%) revealed 93–94% nucleotide identity between HPgV-2 strains. All 12 HPgV-2 RNA-positive cases were identified in individuals also testing positive for HCV RNA (12 of 983; 1.22%), including 2 samples co-infected with HIV, but HPgV-2 RNA was not detected in non-HCV-infected individuals (p<0.0001), including those singly infected by HIV (p = 0.0075) or HBV (p = 0.0077), nor in volunteer blood donors (p = 0.0082). Nine of the 12 (75%) HPgV-2 RNA positive samples were reactive for antibodies to viral serologic markers, whereas only 28 of 2,429 (1.15%) HPgV-2 RNA negative samples were seropositive. Longitudinal sampling in two individuals revealed that active HPgV-2 infection can persist in blood for at least 7 weeks, despite the presence of virus-specific antibodies. One individual harboring both HPgV-2 and HCV RNA was found to be seronegative for both viruses, suggesting a high likelihood of simultaneous acquisition of HCV and HPgV-2 infection from an acute co-transmission event. Taken together, our results indicate that HPgV-2 is a novel bloodborne infectious virus of humans and likely transmitted via the parenteral route.

Discovery of a Novel Human Pegivirus in Blood Associated with Hepatitis C Virus Co-Infection.

Hepatitis C virus (HCV) and human pegivirus (HPgV), formerly GBV-C, are the only known human viruses in the Hepacivirus and Pegivirus genera, respectively, of the family Flaviviridae. We present the discovery of a second pegivirus, provisionally designated human pegivirus 2 (HPgV-2), by next-generation sequencing of plasma from an HCV-infected patient with multiple bloodborne exposures who died from sepsis of unknown etiology. HPgV-2 is highly divergent, situated on a deep phylogenetic branch in a clade that includes rodent and bat pegiviruses, with which it shares <32% amino acid identity. Molecular and serological tools were developed and validated for high-throughput screening of plasma samples, and a panel of 3 independent serological markers strongly correlated antibody responses with viral RNA positivity (99.9% negative predictive value). Discovery of 11 additional RNA-positive samples from a total of 2440 screened (0.45%) revealed 93–94% nucleotide identity between HPgV-2 strains. All 12 HPgV-2 RNA-positive cases were identified in individuals also testing positive for HCV RNA (12 of 983; 1.22%), including 2 samples co-infected with HIV, but HPgV-2 RNA was not detected in non-HCV-infected individuals (p<0.0001), including those singly infected by HIV (p = 0.0075) or HBV (p = 0.0077), nor in volunteer blood donors (p = 0.0082). Nine of the 12 (75%) HPgV-2 RNA positive samples were reactive for antibodies to viral serologic markers, whereas only 28 of 2,429 (1.15%) HPgV-2 RNA negative samples were seropositive. Longitudinal sampling in two individuals revealed that active HPgV-2 infection can persist in blood for at least 7 weeks, despite the presence of virus-specific antibodies. One individual harboring both HPgV-2 and HCV RNA was found to be seronegative for both viruses, suggesting a high likelihood of simultaneous acquisition of HCV and HPgV-2 infection from an acute co-transmission event. Taken together, our results indicate that HPgV-2 is a novel bloodborne infectious virus of humans and likely transmitted via the parenteral route.

Improvement of a DNA vaccine against BoHV-1 using chemical and molecular adjuvants

Improvement of a DNA vaccine against BoHV-1 using chemical and molecular adjuvants

Human Polyomaviruses in Skin Diseases

Polyomaviruses are a family of small, nonenveloped viruses with a circular double-stranded DNA genome of ∼5,000 base pairs protected by an icosahedral protein structure. So far, members of this family have been identified in birds and mammals. Until 2006, BK virus (BKV), JC virus (JCV), and simian virus 40 (SV40) were the only polyomaviruses known to circulate in the human population. Their occurrence in individuals was mainly confirmed by PCR and the presence of virus-specific antibodies. Using the same methods, lymphotropic polyomavirus, originally isolated in monkeys, was recently shown to be present in healthy individuals although with much lower incidence than BKV, JCV, and SV40. The use of advanced high-throughput sequencing and improved rolling circle amplification techniques have identified the novel human polyomaviruses KI, WU, Merkel cell polyomavirus, HPyV6, HPyV7, trichodysplasia spinulosa-associated polyomavirus, and HPyV9. The skin tropism of human polyomaviruses and their dermatopathologic potentials are the focus of this paper.

Diagnostic Strategy Used To Establish Etiologies of Encephalitis in a Prospective Cohort of Patients in England

The laboratory diagnostic strategy used to determine the etiology of encephalitis in 203 patients is reported. An etiological diagnosis was made by first-line laboratory testing for 111 (55%) patients. Subsequent testing, based on individual case reviews, resulted in 17 (8%) further diagnoses, of which 12 (71%) were immune-mediated and 5 (29%) were due to infection. Seventy-five cases were of unknown etiology. Sixteen (8%) of 203 samples were found to be associated with either N-methyl-d-aspartate receptor or voltage-gated potassium channel complex antibodies. The most common viral causes identified were herpes simplex virus (HSV) (19%) and varicella-zoster virus (5%), while the most important bacterial cause was Mycobacterium tuberculosis (5%). The diagnostic value of testing cerebrospinal fluid (CSF) for antibody was assessed using 139 samples from 99 patients, and antibody was detected in 46 samples from 37 patients. Samples collected at 14 to 28 days were more likely to be positive than samples taken 0 to 6 days postadmission. Three PCR-negative HSV cases were diagnosed by the presence of virus-specific antibody in the central nervous system (CNS). It was not possible to make an etiological diagnosis for one-third of the cases; these were therefore considered to be due to unknown causes. Delayed sampling did not contribute to these cases. Twenty percent of the patients with infections with an unknown etiology showed evidence of localized immune activation within the CNS, but no novel viral DNA or RNA sequences were found. We conclude that a good standard of clinical investigation and thorough first-line laboratory testing allows the diagnosis of most cases of infectious encephalitis; testing for CSF antibodies allows further cases to be diagnosed. It is important that testing for immune-mediated causes also be included in a diagnostic algorithm.

Complement Protein C1q Reduces the Stoichiometric Threshold for Antibody-Mediated Neutralization of West Nile Virus

Virus neutralization is governed by the number of antibodies that bind a virion during the cellular entry process. Cellular and serum factors that interact with antibodies have the potential to modulate neutralization potency. Although the addition of serum complement can increase the neutralizing activity of antiviral antibodies in vitro, the mechanism and significance of this augmented potency in vivo remain uncertain. Herein, we show that the complement component C1q increases the potency of antibodies against West Nile virus by modulating the stoichiometric requirements for neutralization. The addition of C1q does not result in virolysis but instead reduces the number of antibodies that must bind the virion to neutralize infectivity. For IgG subclasses that bind C1q avidly, this reduced stoichiometric threshold falls below the minimal number of antibodies required for antibody-dependent enhancement (ADE) of infection of cells expressing Fc-gamma receptors (CD32) and explains how C1q restricts the ADE of flavivirus infection.

Antibody-Enhanced Binding of Dengue-2 Virus to Human Platelets

The mechanisms underlying severe thrombocytopenia in dengue hemorrhagic fever/dengue shock syndrome (DHF/DSS) are not completely understood. We present here the first evidence that dengue type 2 virus binds to human platelets only in the presence of virus-specific antibody, supporting a role for immune-mediated clearance of platelets in the pathogenesis of thrombocytopenia in DHF/DSS. Antibody-enhanced binding of virus of platelets was also demonstrated with a panel of eight murine monoclonal antibodies specific for the dengue E protein. The degree of binding was dependent on the antibody used but not on the antibody IgG subclass, indicating that factors other than the platelet Fc receptor are involved in binding of virus-antibody complexes to the platelet surface. Confirmation that antibody-dependent virus binding to platelets is not primarily mediated by the platelet Fc receptor was obtained by demonstrating good binding even when platelets were pretreated with the FcγRII-specific antibody IV.3.

Detection of autoantibodies to neural antigens in the CSF of Japanese encephalitis patients and correlation of findings with the outcome

This study reports the detection of autoantibodies to myelin basic protein (MBP) and neurofilament proteins (NFP) in serum and cerebrospinal fluid (CSF) of Japanese encephalitis patients. The diagnosis of Japanese encephalitis was confirmed in 72 patients by the presence of virus specific antibodies to JEV in the CSF ( 28 72 ), viral antigen in the CSF ( 19 72 ) and simultaneous presence of both antigen and JEV antibodies in the CSF in 25 72 patients. Autoantibodies to either purified NFP (10) or MBP (8) or both (17) were detected in the CSF of 35 patients by ELISA in contrast with the control CSF samples. Amongst them 20 had similar antibodies in the serum as well. Correlation of immunological findings with the clinical outcome revealed that the presence of autoantibodies in the CSF especially to NFP was associated with a fatal outcome ( P < 0.05).

A review of feline infectious peritonitis virus: molecular biology, immunopathogenesis, clinical aspects, and vaccination

Feline infectious peritontis (FIP) has been an elusive and frustrating problem for veterinary practitioners and cat breeders for many years. Over the last several years, reports have begun to elucidate aspects of the molecular biology of the causal virus (FIPV). These papers complement a rapidly growing base of knowledge concerning the molecular organization and replication of coronaviruses in general. The fascinating immunopathogenesis of FIPV infection and the virus' interaction with macrophages has also been the subject of several recent papers. It is now clear that FIPV may be of interest to scientists other than veterinary virologists since its pathogenesis may provide a useful model system for other viruses whose infectivity is enhanced in the presence of virus-specific antibody. With these advances and the recent release of the first commercially-available FIPV vaccine, it is appropriate to review what is known about the organization and replication of coronaviruses and the pathogenesis of FIPV infection.

Murine coronavirus-induced encephalomyelitides in rats: Analysis of immunoglobulins and virus-specific antibodies in serum and cerebrospinal fluid

The humoral intrathecal immune response in coronavirus-induced demyelinating encephalomyelitis in rats associated with an autoimmune reaction to brain antigen, was analysed. The CSF of these animals revealed immune reactions which were directed against coronavirus and other, unknown, antigens. In general, no direct correlation between the disease, the state of the blood-brain barrier (BBB), intrathecal synthesis of Ig and the presence of virus-specific antibodies was detectable, suggesting that the humoral, in contrast to the cellular, immune response does not play a significant pathogenetic role in this CNS disease.

Cell-Mediated Cytotoxicity: Comparison of Primary and Secondary Immune Reactions after Parainfluenza Type 1 Virus Inoculation

H-2 antigen compatibility of effector spleen cells and target cells was necessary for cell-mediated cytotoxicity (CMC) after both 1 and 2 intraperitoneal inoculation of 6/94 virus into mice. T cells were responsible for both primary and secondary CMS reactions. Neither reaction was influenced by the presence of virus-specific antibody added to the test system. Secondary CMC peaked earlier than the primary response and declined more rapidly. In vitro stimulation of primed spleen cells could also be used to detect the secondary CMC response.

Incidence of virus-specific antibodies and autoantibodies in sera of patients with warts.

Summary Sera from fifty-two patients with warts and control subjects, the majority of whom were age and sex matched, were tested for antiviral antibody, anticellular antibody against wart cells, fibrillar anticellular antibody and smooth muscle antibody (SMA). There was a higher prevalence of the autoantibodies in patients with warts as compared to control subjects. There was no correlation between the presence of virus-specific antibodies and the autoantibodies.