Protection against yellow fever in monkeys by immunization with yellow fever virus nonstructural protein NS1.

Immunization of monkeys with yellow fever virus-specified nonstructural protein NS1 resulted in protection against fatal hepatitis as well as marked reduction in the magnitude of viremia after subcutaneous challenge with yellow fever virus. The results may be relevant to the design of possible subunit or recombinant flavivirus vaccines.     

Yellow fever vaccine protects mice against Zika virus infection

Zika virus (ZIKV) emerged as an important infectious disease agent in Brazil in 2016. Infection usually leads to mild symptoms, but severe congenital neurological disorders and Guillain-Barré syndrome have been reported following ZIKV exposure. Creating an effective vaccine against ZIKV is a public health priority. We describe the protective effect of an already licensed attenuated yellow fever vaccine (YFV, 17DD) in type-I interferon receptor knockout mice (A129) and immunocompetent BALB/c and SV-129 (A129 background) mice infected with ZIKV. YFV vaccination provided protection against ZIKV, with decreased mortality in A129 mice, a reduction in the cerebral viral load in all mice, and weight loss prevention in BALB/c mice. The A129 mice that were challenged two and three weeks after the first dose of the vaccine were fully protected, whereas partial protection was observed five weeks after vaccination. In all cases, the YFV vaccine provoked a substantial decrease in the cerebral viral load. YFV immunization also prevented hippocampal synapse loss and microgliosis in ZIKV-infected mice. Our vaccine model is T cell-dependent, with AG129 mice being unable to tolerate immunization (vaccination is lethal in this mouse model), indicating the importance of IFN-γ in immunogenicity. To confirm the role of T cells, we immunized nude mice that we demonstrated to be very susceptible to infection. Immunization with YFV and challenge 7 days after booster did not protect nude mice in terms of weight loss and showed partial protection in the survival curve. When we evaluated the humoral response, the vaccine elicited significant antibody titers against ZIKV; however, it showed no neutralizing activity in vitro and in vivo. The data indicate that a cell-mediated response promotes protection against cerebral infection, which is crucial to vaccine protection, and it appears to not necessarily require a humoral response. This protective effect can also be attributed to innate factors, but more studies are needed to strengthen this hypothesis. Our findings open the way to using an available and inexpensive vaccine for large-scale immunization in the event of a ZIKV outbreak.     

铕螯合剂DTTAEuNa标记流行性出血热单克隆抗体的实验研究

用Ｎ′（对异硫氰基苄基）二乙三胺Ｎ１,Ｎ２,Ｎ３四乙酸铕钠（ＤＴＴＡＥｕＮａ）作为螯合剂,标记不同蛋白浓度的流行性出血热单克隆抗体（ＥＨＦＭｃＡｂ）,经ＳｅｐｈａｋｅｘＧ５０凝胶柱分离标记的单抗分子,通过对层析液的紫外吸收峰处的蛋白光密度,时间分辨荧光强度以及免疫活性测定表明：二批Ｅｕ标记单抗的比活性分别为７８和１４７个Ｅｕ３＋／ＥＨＦＭｃＡｂ,标记回收率分别为４１％和４２％,标记物的免疫活性良好,可用于流行性出血热的抗原及抗体检测     

Virus-induced CRISPR-Cas9 system improved resistance against tomato yellow leaf curl virus

Molecular Biology Reports - Plant viruses are the most significant factors associated with massive economical losses in agricultural industries worldwide. Accordingly, many studies are dedicated to...     

Ultrastructural changes caused by yellow fever virus at the level of the kidneys in newborn mice

The author is studying the ultrastructural modifications provoked by the yellow fever virus in the kidneys of baby mice. As a result of the study it has been found that minor changes start appearing as early as the first day and these lead finally to necrosis. The process consists of 5 phases which are the development of endoplasmatic reticulum, the envelopment of the mitochondria by the folds of endoplasmatic reticulum, mitochondrial autophagocytosis, the development of microvilli at the cell surface and the total necrosis of the renal cell.     

Role of the transmembrane domains of prM and E proteins in the formation of yellow fever virus envelope

Flavivirus envelope proteins have been shown to play a major role in virus assembly. These proteins are anchored into cellular and viral membranes by their C-terminal domain. These domains are composed of two hydrophobic stretches separated by a short hydrophilic segment containing at least one charged residue. We investigated the role of the transmembrane domains of prM and E in the envelope formation of the flavivirus yellow fever virus (YFV). Alanine scanning insertion mutagenesis has been used to examine the role of the transmembrane domains of prM and E in YFV subviral particle formation. Most of the insertions had a dramatic effect on the release of YFV subviral particles. Some of these mutations were introduced into the viral genome. The ability of these mutant viruses to produce infectious particles was severely reduced. The alanine insertions did not affect prM-E heterodimerization. In addition, replacement of the charged residues present in the middle of the transmembrane domains had no effect on subviral particle release. Taken together, these data indicate that the transmembrane domains of prM and E play a crucial role in the biogenesis of YFV envelope. In addition, these data indicate some differences between the transmembrane domains of the hepaciviruses and the flaviviruses.     

Yellow fever virus/dengue-2 virus and yellow fever virus/dengue-4 virus chimeras: biological characterization,immunogenicity, and protection against dengue encephalitis in the mouse model

Two yellow fever virus (YFV)/dengue virus chimeras which encode the prM and E proteins of either dengue virus serotype 2 (dengue-2 virus) or dengue-4 virus within the genome of the YFV 17D strain (YF5.2iv infectious clone) were constructed and characterized for their properties in cell culture and as experimental vaccines in mice. The prM and E proteins appeared to be properly processed and glycosylated, and in plaque reduction neutralization tests and other assays of antigenic specificity, the E proteins exhibited profiles which resembled those of the homologous dengue virus serotypes. Both chimeric viruses replicated in cell lines of vertebrate and mosquito origin to levels comparable to those of homologous dengue viruses but less efficiently than the YF5.2iv parent. YFV/dengue-4 virus, but not YFV/dengue-2 virus, was neurovirulent for 3-week-old mice by intracerebral inoculation; however, both viruses were attenuated when administered by the intraperitoneal route in mice of that age. Single-dose inoculation of either chimeric virus at a dose of 10(5) PFU by the intraperitoneal route induced detectable levels of neutralizing antibodies against the homologous dengue virus strains. Mice which had been immunized in this manner were fully protected from challenge with homologous neurovirulent dengue viruses by intracerebral inoculation compared to unimmunized mice. Protection was associated with significant increases in geometric mean titers of neutralizing antibody compared to those for unimmunized mice. These data indicate that YFV/dengue virus chimeras elicit antibodies which represent protective memory responses in the mouse model of dengue encephalitis. The levels of neurovirulence and immunogenicity of the chimeric viruses in mice correlate with the degree of adaptation of the dengue virus strain to mice. This study supports ongoing investigations concerning the use of this technology for development of a live attenuated viral vaccine against dengue viruses.     

黄热病毒的基因组及其蛋白研究进展

黄热病毒(Yellowfever virus,YFV)是属于黄病毒科(Flavivirdae)黄病毒属(Flavivirus)的典型代表,为RNA病毒其不仅是第一个被发现的导致人类疾病的"滤过性颗粒",也是第一个被证实通过蚊蜱传播的病毒。黄热病是一种区域性疾病,在南美     

Genetic diversity in exotic oat germplasm & resistance against barley yellow dwarf virus

Oat (Avena sativa L.) is an important fodder crop of Pakistan, though with low productivity. The present study was conducted to evaluate the performance and genetic diversity of exotic oat germplasm, with emphasis on cereal yellow dwarf virus resistance. A total of 16 exotic line (introduced from Aarhus University Denmark) and 1 local line (provided by The University of Agriculture Peshawar), were grown during the season 2017–18 in Completely Randomized Block Design with three replications across two locations of Khyber Pakhtunkhwa i.e., Peshawar and Kohat. Field testing enabled to collect the data on BYDV incidence, BYDV severity, aphid infestation, plant height, leaf area, panicle length, panicle weight, spikelets per panicle, 1000 grain weight (g), grain yield (g), biological yield (g) and harvest index (%). Prevalence of BYDV was variable across location and over time. Six weeks data showed high disease pressure at Peshawar (85%), with SA-O-01 genotype having AUDPC value of 95%. Almost all the varieties showed less tolerance towards the Aphids attack. Line SA-O-15 showed the maximum 1000 grain weight (42.6 g) at Kohat, while SA-O-4 showed the maximum 1000 grain weight (60.7 g) at Peshawar. Line SA-O-05 (3634 g per (0.9 m²) plot) gave the maximum biological yield at Kohat station, while Line SA-O-01 gave the maximum biological yield (2517 g) at Peshawar. Mean grain yield for Kohat was recorded 0.155 g per (0.9 m²) plot while for Peshawar it was 0.231 g per (0.9 m²) plot. At Kohat line SA-O-10 produced the maximum grain yield (0.229 g), while line SA-O-12 produced the maximum grain yield at Peshawar (0.288 g). Molecular genotyping with a set of 4 RAPD primers revealed substantial diversity among17 oat lines. A total of 23 loci were amplified showing a high level of variations and polymorphism among the proposed lines. The maximum number of loci was recorded for GLA-04 (8), while the minimum number of loci was recorded for GLD-18 (4). Among the tested RAPD primers the maximum gene diversity (0.529) was recorded for loci GLA-03B230, GLA-04B130, GLA-04B300, GLB-05B150 and GLA-18B100 while the minimum (0.118) genetic diversity was recorded for loci GLA-03B600, GLB-05B330 and GLA-18B500. A clear divergence was found between most of the exotic oat lines. The observed genetic diversity in exotic oat germplasm and its resistance towards Barley Yellow Dwarf virus could be useful for oat genetic improvement and broadening the genetic background of cultivated oat germplasm.     

Functional requirements of the yellow fever virus capsid protein

Although it is known that the flavivirus capsid protein is essential for genome packaging and formation of infectious particles, the minimal requirements of the dimeric capsid protein for virus assembly/disassembly have not been characterized. By use of a trans-packaging system that involved packaging a yellow fever virus (YFV) replicon into pseudo-infectious particles by supplying the YFV structural proteins using a Sindbis virus helper construct, the functional elements within the YFV capsid protein (YFC) were characterized. Various N- and C-terminal truncations, internal deletions, and point mutations of YFC were analyzed for their ability to package the YFV replicon. Consistent with previous reports on the tick-borne encephalitis virus capsid protein, YFC demonstrates remarkable functional flexibility. Nearly 40 residues of YFC could be removed from the N terminus while the ability to package replicon RNA was retained. Additionally, YFC containing a deletion of approximately 27 residues of the C terminus, including a complete deletion of C-terminal helix 4, was functional. Internal deletions encompassing the internal hydrophobic sequence in YFC were, in general, tolerated to a lesser extent. Site-directed mutagenesis of helix 4 residues predicted to be involved in intermonomeric interactions were also analyzed, and although single mutations did not affect packaging, a YFC with the double mutation of leucine 81 and valine 88 was nonfunctional. The effects of mutations in YFC on the viability of YFV infection were also analyzed, and these results were similar to those obtained using the replicon packaging system, thus underscoring the flexibility of YFC with respect to the requirements for its functioning.     

Neuroadapted yellow fever virus 17D: determinants in the envelope protein govern neuroinvasiveness for SCID mice

A molecular clone of mouse-neuroadapted yellow fever 17D virus (SPYF-MN) was used to identify critical determinants of viral neuroinvasiveness in a SCID mouse model. Virus derived from this clone differs from nonneuroinvasive YF5.2iv virus at 29 nucleotide positions, encoding 13 predicted amino acid substitutions and 2 substitutions in the 3' untranslated region (UTR). The virulence determinants of SPYF-MN for SCID mice were identified by constructing and characterizing intratypic viruses in which the E protein of SPYF-MN was expressed in the YF5.2iv background (SPYF-E) or the E protein of YF5.2iv was expressed in the SPYF-MN background (YF5.2-E). SPYF-E caused lethal encephalitis in young adult SCID mice after intraperitoneal inoculation, with average survival times and tissue virus burdens resembling those of mice inoculated with the parental SPYF-MN virus. To define which domains of the E protein are involved in neuroinvasiveness, two viruses were tested in which the amino acid substitutions in domains I-II and III were segregated. This revealed that substitutions in domain III (residues 305, 326, and 380) were critical for the neuroinvasive phenotype, based on average survival times and tissue burdens of infectious virus. Comparison of growth properties of the various intratypic viruses in cell culture indicated that no inherent defects in replication efficiency were likely to account for the biological differences observed in these experiments. These findings demonstrate that the E protein is a critical factor for yellow fever virus neuropathogenesis in the SCID mouse model and that the neuroinvasive properties depend principally on functions contributed by domain III of this protein. To assess whether critical determinants for neuroinvasion of normal ICR mice by SPYF virus were also in the E protein, sequences of viruses recovered from brains of ICR mice succumbing to encephalitis with the parental SPYF virus were derived. No differences were found in the E protein; however, two substitutions were present in the 3' UTR compared to that of SPYF-MN, one of which is predicted to alter RNA secondary structure in this region. These findings suggest that the 3' UTR may also affect neuroinvasiveness of SPYF virus in the mouse model.