Virology: Research and Treatment

An evidence-based approach, such as those developed in clinical and veterinary medicine, was applied to the detection of Plum pox virus (PPV) during the dormant period. A standardized methodology was used for the calculation of parameters of the operational capacity of DASI-ELISA and RT-PCR in wintertime. These methods are routinely handled to test the sanitary status of plants in national or international trading and in those cases concerning export-import of plant materials. Diagnosis often has to be performed during the dormant period, when plant material is commercialized. Some guidelines to interpret diagnostic results of wintertime are provided in an attempt to minimize risks associated with the methods and over-reliance on the binary outcome of a single assay. In order to evaluate if a complementary test increased the confidence of PPV diagnosis when discordant results between DASI-ELISA and RT-PCR are obtained, NASBA-FH also was included. Likelihood ratios of each method were estimated based on the sensitivity and specificity obtained in wintertime. Subsequently, a Bayesian approach was performed to calculate post-test probability of PPV infection in spring. Results of evidence-based approach show that different PPV prevalences require different screening tests. Thus, at very low PPV prevalence levels DASI-ELISA should be used as the election method, whilst at the highest PPV prevalence levels RT-PCR should be performed. NASBA-FH could be used at medium prevalences to clarify discordances between DASI-ELISA and RT-PCR.

Porcine reproductive and respiratory syndrome virus (PRRSV) is an economically significant agent for which there currently are no effective treatments. Development of antiviral agents for PRRSV as well as many other viruses has been limited by toxicity of known antiviral compounds. In contrast, antibiotics for non-virus microbial infections have been widely useful, in part because of their acceptable toxicity in animals. We report here the discovery that the quinolone-containing compound Plasmocin^™, as well as the quinolones nalidixic acid and ciprofloxacin, have potent anti-PRRSV activity in vitro. PRRSV replication was inhibited by these antibiotics in both cultured MARC-145 cells and cultured primary alveolar porcine macrophages (PAMs). Furthermore, sub-optimal concentrations of nalidixic acid synergized with antiviral cytokines (AK-2 or IFN-γ) to quantitatively and qualitatively inhibit PRRSV replication in MARC-145 cells or PAMs. The antiviral activity of Plasmocin and nalidixic acid correlated with reduced actin expression in MARC-145 cells. Replication of the related lactate dehydrogenase-elevating virus (LDV) was also inhibited in primary mouse macrophages by Plasmocin. These results are significant to the development of antiviral strategies with potentially reduced toxicity, and provide a model system to better understand regulation of arterivirus replication.

An intriguing difference between the E2 glycoprotein of CSFV and the other groups of pestiviruses (nonCSFV) is a lack of two cysteine residues on positions cysteine 751 and 798. Other groups of pestivirus are not restricted to one species as swine, whereas CSFV is restricted to swine and wild boar. We constructed chimeric CSFV/BVDV E2 genes based on a 2D model of E2 proposed by van Rijn et al. (van Rijn et al. 1994, J Virol 68, 3934–42) and confirmed their expression by immunostaining of plasmid-transfected SK6 cells. No equivalents for the antigenic units B/C and A were found on E2 of BVDVII. This indicates major structural differences in E2. However, the immunodominant BVDVII domain A, containing epitopes with essential amino acids between position 760–764, showed to be dependent on the presence of the region defined by amino acids 684 to 796. As for the A domain of CSFV, the BVDVII A-like domain seemed to function as a separate unit. These combined domains in E2 proved to be the only combination which was functional in viral background of CSFV C-strain. The fitness of this virus (vflc36^{BVDVII 684–796}) seemed to be reduced compared to vflc9 (with the complete antigenic region of BVDVII).

Since the isolation and discovery of human T-cell leukemia virus type 1 (HTLV-1) over 30 years ago, researchers have utilized animal models to study HTLV-1 transmission, viral persistence, virus-elicited immune responses, and HTLV-1-associated disease development (ATL, HAM/TSP). Non-human primates, rabbits, rats, and mice have all been used to help understand HTLV-1 biology and disease progression. Non-human primates offer a model system that is phylogenetically similar to humans for examining viral persistence. Viral transmission, persistence, and immune responses have been widely studied using New Zealand White rabbits. The advent of molecular clones of HTLV-1 has offered the opportunity to assess the importance of various viral genes in rabbits, non-human primates, and mice. Additionally, over-expression of viral genes using transgenic mice has helped uncover the importance of Tax and Hbz in the induction of lymphoma and other lymphocyte-mediated diseases. HTLV-1 inoculation of certain strains of rats results in histopathological features and clinical symptoms similar to that of humans with HAM/TSP. Transplantation of certain types of ATL cell lines in immunocompromised mice results in lymphoma. Recently, “humanized” mice have been used to model ATL development for the first time. Not all HTLV-1 animal models develop disease and those that do vary in consistency depending on the type of monkey, strain of rat, or even type of ATL cell line used. However, the progress made using animal models cannot be understated as it has led to insights into the mechanisms regulating viral replication, viral persistence, disease development, and, most importantly, model systems to test disease treatments.

Hepatitis B and C are serious public health problems worldwide. Thalassemia patients are dependent on blood transfusions throughout their life and are at high risk of viral infections. The aim of this study was to estimate the prevalence of hepatitis B/C infections and different clinical parameters in multitransfused thalassemia population. In this study, 262 multitransfused β-thalassemia patients were enrolled from the capital twin cities of Pakistan. The presence of hepatitis B virus (HBV)/hepatitis C virus (HCV), alanine aminotransferase (ALT) level, serum creatinine, serum ferritin, hepatomegaly, splenomegaly, and splenectomy were analyzed. The overall prevalence of HBV and HCV was 3.08% and 55.73%, respectively, with 100% of patients older than 20 years had HCV infection. The ALT levels among HBV- and HCV-positive thalassemia patients were 92.62 ± 41.57 U/L and 98 ± 63.65 U/L, respectively; creatinine values observed were 0.4 ± 0.35 mg/dL (for HBV) and 0.39 ± 0.24 mg/dL (for HCV), while serum ferritin levels were 6865.87 ± 1649.13 ng/dL (for HBV) and 5445.95 ± 3059.28 ng/dL (for HCV). A total of 74.8% and 82.20% of HBV- and HCV-positive patients had hepatomegaly with an average increase in liver size of 4.17 and 4.33 cm, respectively. Splenomegaly was observed in 64.9% and 67.12% of HBV- and HCV-positive patients with an average increase in spleen size of 4 and 4.46 cm, respectively. Splenectomy was observed among 14.50% and 15.75% of HBV- and HCV-infected thalassemia patients. There is a strong need to properly screen blood before transfusions to reduce the future load of viral hepatitis from Pakistan.

By targeting CD4⁺ effector T cells, HIV has a dramatic impact on the depletion, expansion and function of the different polarized T cell subsets. The maturation of T cell lineages is in part driven by intrinsic transcription factors that potentially influence how efficiently HIV replicates. In this review, we explore whether transcription factors that are required for polarizing T cells influence HIV replication. In particular, we examine provirus transcription as well as the establishment and maintenance of HIV latency. Furthermore, it is suggested these factors may provide novel cell-specific therapeutic strategies for targeting the HIV latent reservoir.

Almost 7 million children under the age 5 die each year, and most of these deaths are attributable to vaccine-preventable infections. Young infants respond poorly to infections and vaccines. In particular, dendritic cells secrete less IL-12 and IL-18, CD8^pos T cells and NK cells have defective cytolysis and cytokine production, and CD4^pos T cell responses tend to bias towards a Th2 phenotype and promotion of regulatory T cells (T_regs). The basis for these differences is not well understood and may be in part explained by epigenetic differences, as well as immaturity of the infant's immune system. Here we present a third possibility, which involves active suppression by immune regulatory cells and place in context the immune suppressive pathways of mesenchymal stromal cells (MSC), myeloid-derived suppressor cells (MDSC), CD5^pos B cells, and T_regs. The immune pathways that these immune regulatory cells inhibit are similar to those that are defective in the infant. Therefore, the immune deficiencies seen in infants could be explained, in part, by active suppressive cells, indicating potential new avenues for intervention.

In 1964, Epstein, Barr, and Achong published a report outlining their discovery of viral particles in lymphoblasts isolated from a patient with Burkitt lymphoma. The Epstein-Barr virus (EBV) was the first human cancer virus to be described, and its discovery paved the way for further investigations into the oncogenic potential of viruses. In the decades following the discovery of EBV, multinational research efforts led to the discovery of further viral causes of various human cancers. Lymphomas are perhaps the cancer type that is most closely associated with oncogenic viruses: infection with EBV, human T-lymphotropic virus 1 (HTLV-1), human immunodeficiency virus (HIV), Kaposi sarcoma-associated herpesvirus/human herpesvirus 8, and hepatitis C virus have all been associated with lymphomagenesis. Lymphomas have also played an important role in the history of oncoviruses, as both the first human oncovirus (EBV) and the first human retrovirus (HTLV-1) were discovered through isolates taken from patients with unique lymphoma syndromes. The history of the discovery of these 2 key oncoviruses is presented here, and their impact on further medical research, using the specific example of HIV research, is briefly discussed.

Dengue disease is an emerging mosquito-borne viral infection transmitted between humans by Aedes spp. that are distributed mainly in the tropical and subtropical region along with chikungunya and zika diseases. The distribution of dengue disease is influenced by local variation, such as geography, rainfall, temperature, and rapid urbanization or migration. The epidemy of mosquito-borne infection significantly led to increased number of cases and hyperendemicity which induce a more severe form of dengue accompanied by cocirculation of chikungunya and zika. The rapid global spreading of dengue disease created public health burdens that are presently unfulfilled by the absence of specific therapy, simple diagnosis tool for the early phase, and effective and efficient vector control system. This review highlights the current situation of dengue distribution, epidemiology, and new strategies for early dengue diagnosis and risk prediction of severity that can be used to improve oversight and alleviate the heavy burden of the disease.

JC virus-specific CD8+ cytotoxic T lymphocytes are associated with a favorable outcome in patients with progressive multifocal leukoencephalopathy. However, very few JC virus T cell epitopes restricted to MHC class I have been defined. Of the two HLA-A^*0201-restricted JCV epitopes, VP1_p36 and VP1_p100, studies have shown that they are conserved T cell epitopes of polyomaviruses. The cross-recognition associated to these epitopes has complicated the efforts of understanding the dynamics of immune response to JC virus. Based on the previously identified HLA-A^*0201 binding T cell epitope of Simian virus 40 T antigen P_281–289 (KCDDVLLLL) and BK virus T antigen P_558–566 (SLQNSEFLL), T cell epitopes of JC Virus T antigen P_282–290 (KCEDVFLLM) and P_557–565 (SLSCSEYLL) were identified. In this report, we demonstrated that JC Virus P_282–290 and P_557–565 were able to stimulate T cell responses in healthy donors’ PBMCs and CD8+ cytotoxic T lymphocytes raised with both peptides could recognize and lyse their targets. Most importantly, there were no T cell cross-recognitions between JC Virus, BK Virus and SV40 virus. Therefore, JCV T-ag epitopes P_282–290 and P_557–565 could be better antigen epitopes compared to VP1_p36 and VP1_p100 to study the dynamics of cellular immune response to JCV in PML patients. In addition, as a HLA-A^*0201 binding T cell epitope, both peptides could be a valuable component of immunotherapies aiming at increasing the cellular immune response against JCV for the treatment of progressive multifocal leukoencephalopathy.

Upon transmission, human immunodeficiency virus type 1 (HIV-1) establishes infection of the lymphatic reservoir, leading to profound depletion of the memory CD4⁺ T cell population despite the induction of the adaptive immune response. The rapid evolution and association of viral variants having distinct characteristics during different stages of infection, the level of viral burden, and rate of disease progression suggest a role for viral variants in this process. Here, we review the literature on HIV-1 variants and disease and discuss the importance of viral fitness for transmission and disease.

Kaposi's sarcoma associated herpesvirus (KSHV or human herpesvirus 8 [HHV-8]) is a gammaherpesvirus highly associated with KS, primary effusion lymphoma (PEL), and multicentric Castleman's disease, an aggressive lymphoproliferative disorder. KSHV, like other gammaherpesvirus latently infects predominantly B-cells and endothelial cells. Infected cells retain the virus from one generation to the next existing as a multicopy circular episomal DNA in the nucleus, expressing a limited subset of viral genes. Of these latently expressed genes, LANA1, the latency associated nuclear antigen is highly expressed in all forms of KS-associated malignancies. Various studies so far show that LANA1 tethers the viral episomes to host chromosomes and binds to specific sites within and close to the TR elements contributing to the stable maintenance of the viral episomes in successive daughter cells. Anti-LANA1 intrabody strategies might represent a new therapeutic approach to treatment of KSHV infections, since LANA1 is regained for KSHV latency. In addition, the use of intrabodies can help drug development by mapping LANA1 inhibiting regions. We report development of several LANA1 specific single chain antibodies from immunized rabbits that can be expressed intracellularly, bind to LANA1 epitopes and can be used for functional KSHV studies on viral latency.

A cross-sectional study in Nigeria was undertaken to determine the epidemiology, seroprevalence, and associated risk factors, of hepatitis E virus (HEV). A total of 462 subjects were used for the study, categorized into four groups: apparently healthy persons, pregnant women, HIV positive subjects, and animal handlers. Information was obtained from subjects using interviewer-administered questionnaire. Blood samples were collected and analyzed for HEV antibodies (IgG and IgM) using enzyme-linked immunosorbent assay (ELISA) technique. Results obtained were analyzed using Statistical Package for Social Sciences (SPSS) version 17.0 statistical software. The overall seroprevalence of IgG and IgM was 42.7 and 0.9%, respectively. Animal handlers had the highest seroprevalence (66.7%). The associated risk factors for IgM seroprevalence were rural dwelling (P = 0.039, odds ratio (OR) 3.3, 95% confidence interval (CI) 0.7–15.4), blood transfusion (P < 0.001, OR 9.6, 95% CI 2.6–35.6), attending to animals (P = 0.032, OR 4.9, 95% CI 0.9–26.6), and waste disposal (P < 0.001). Factors associated with IgG were age (P = 0.044), location (P < 0.001), marital status (P < 0.001), formal education (P < 0.001), farming as occupation (P < 0.001), rural dwelling (P = 0.001), waste disposal (P < 0.001), alcohol consumption (P = 0.001, OR 2.4, 95% CI 1.4–4.0), open defecation (P < 0.001, OR 2.9, 95% CI 1.4–5.7), attending to animals (P < 0.001, OR 2.3, 95% CI 1.6–3.4), consuming unwashed fruits/vegetables (P < 0.001, OR 4.2, 95% CI 0.3–54.1), and stream/river as a source of drinking water (P < 0.001, OR 3.6, 95% CI 1.6–7.8). Preventive public health measures should be reinforced among all communities, particularly domestic animal handlers and pregnant women. Potable water should be provided for all communities. Data suggest that HEV remains an under-recognized and significant public health problem, warranting further attention and research.