Molecular detection and genomic characterization of Samak Micromys paramyxovirus-1 and –2 in Micromys minutus, Republic of Korea

Paramyxoviruses are well-known as pathogen with wide range host and implied in critical public health threat. The hosts are small and large mammals, including marine species. To date, in Republic of Korea, the natural virus diversity of M. minutus remains unclear. The study is aimed to screen RNA viruses targeted with specific mammal species may help to prepare for future pandemics. A total of 145 M. minutus were trapped and necropsied for various tissues. The species of M. mimutus were classified by field morphological characteristic and sequencing of mtDNA cyt b gene. Molecular detection of paramyxovirus screening was performed using nested PCR reaction, continued by genomic characterization and analysis for N-glycosylation potential at glycoprotein gene, phylogenetic and cophylogenetic analysis. In results, 39.3% of M. minutus isolates tested positive for paramyxovirus with the highest percentages in late-adult male animals. One sample Samak Mycromys paramyxovirus (SMPV1) was 19.911 nt with a GC content of 40.8%. The genome structure of SMPV-1 consisted of eight genes in the order 3’-N-P-M-F-SH-TM-G-L-5’. The other three samples were SMPV-2 with full genome length 18,199 nt with a GC content of 40.18%, 40.15% and 40.19%. The genome structure of SMPV-2 consisted of seven genes in the order 3ʹ-N-P-M-F-TM-G-L-5ʹ. The inclusion of SMPV-1 and − 2 sequences in paramyxoviruses is crucial for the surveillance of emerging infectious diseases and provides insights into the genetic diversity of paramyxoviruses in small wild mammals.   
(INV)

Fully human single-domain antibody targeting a highly conserved cryptic epitope on the Nipah virus G protein

Nipah virus infection, one of the top priority diseases recognized by the World Health Organization, can cause severe respiratory illness and fatal encephalitis in humans. The mortality rate is high (50-95%). A key element in combating NiV infections is the development of NiV-neutralizing antibodies. In this research, the authors utilize the platform to discover a unique NiV-neutralizing fully human single-domain antibodies (UdAb) that specifically targets a conserved cryptic epitope located at the dimeric interface of the NiV G glycoprotein. By targeting a conserved epitope on the head domain of the NiV G glycoprotein, that it may disrupt G protein tetramerization, thereby hindering F protein activation and membrane fusion. Compared to the full-length monoclonal antibody m102.4, this single-domain antibody displays significantly greater potency in inhibiting viral membrane fusion and more efficient penetration into the murine brain. As a result, this single-domain antibody has potential applications in vaccine development and shows promise as a treatment option to stop Nipah virus infections.
(DKW)

Promotion of virus assembly and Organization by the measles virus matrix protein

There are currently no approved antivirals to treat measles virus (Mev) or other paramyxoviruses, despite the fact that MeV is still a serious human infection. Here, the researchers employed cryo-electron tomography to clarify the rules guiding paramyxovirus assembly in human cells infected with MeV. During the phases of virus assembly and budding as well as virion release, the three-dimensional (3D) configuration of the MeV structural proteins, which include the Matrix protein (M), the ribonucleoprotein complex (RNP) and surface glycoproteins (F and H) was described. The M protein is seen as a well-organized membrane associated two dimensional (2D) paracrystalline array. The discovery of a two layered F-M lattice raises the possibility that interactions between the F and M could coordinate crucial MeV assembly processes. The RNP complex continues to be connected to and nearer to the M lattice in this model. In conclusion, the M lattice makes it easier for the RNP and surface glycoproteins to be incorporated and concentrated in a well-ordered manner at virus assembly locations.
(SN)

A novel factor I activity in Nipah virus inhibits human complement pathways through cleavage of C3b

Nipah virus (NiV) belongs to the family Paramyxoviridae, genus Henipavirus, and its host is the megabat. Nipah virus is an emerging zoonotic viral pathogen that can cause serious respiratory and neurological diseases. The complement cascade initiated by three main pathways: the classical pathway, the lectin pathway, or the alternative pathway. These three pathways converge on the C3, which is activated by cleavage to C3a and C3b. C3b covalently bind to virus components and facilitate opsonin action and phagocytosis. Factor I cleaves and inactivates C3b. A novel mechanism has been identified that NiV can cleave and inactivate C3b via factor I-like protease activity to avoid killing from complement. Inhibiting this ability of NiV could be the basis for the development of safer vaccine and more effective therapies.
(TT)

Feline morbillivirus infection associated with fatal encephalitis in a Bengal cat

The genus Morbillivirus of the family Paramyxoviridae belongs to viruses that are highly contagious to humans and animals, such as canine distemper virus and measles virus. Feline morbillivirus (FeMV) is a novel morbillivirus discovered in 2012 in stray cats in Hong Kong and China, and FeMV has been detected in domestic cats all over the world. The authors investigated the possible neurotoxicity of FeMV to cats and reported a case of a cat with fatal encephalitis associated with FeMV infection. The brain was removed from a 2-month-old Bengal cat that showed neurological symptoms, and inclusion bodies were observed within the neurons on histopathological examination. They extracted total RNA from brain samples and performed sequencing analysis and the sequence of FeMV Locarno/CH2011 was determined. In addition, in situ hybridization (ISH) was used to detect FeMV RNA in the somatic cells of neurons. These results contribute to the increased recognition of FeMV as a neurotropic pathogen in cats.
(SM)

Establishment of serological neutralizing tests using pseudotyped viruses for comprehensive detection of antibodies against all 18 lyssaviruses

Rabies is a zoonotic and can cause neurological disease caused by rabies lyssavirus (RABV) and other lyssaviruses. Once clinical symptoms of rabies appear, the disease is almost invariably fatal. At least seven lyssaviruses, RABV, ABLV, DUVV, EBLV-1, EBLV-2, IRKV, and MOKV, have been responsible for fatal infections in humans. Historically, for the rabies vaccine, the research has primarily focused on the cross-reactivity of RABV vaccine immune sera against other lyssaviruses. As these studies have shown, RABV vaccinations do not protect for other phylogroup lyssaviruses. Therefore, it is now important to find vaccine antigens that are effective against novel lyssavirus. The study introduced a new serological neutralizing test (NT) system using vesicular stomatitis virus (VSV) pseudotypes with the glycoproteins of all 18 lyssavirus species. In this study, researchers can assess cross-reactivity of current rabies vaccines and lyssavirus glycoprotein specific antibodies against the entire range of lyssaviruses. The identification of glycoproteins that elicit cross-protective antibodies may serve as the basis for a next-generation vaccine that provides protection against a variety of lyssaviruses, not just RABV. This system will be useful to detect antibodies against lyssaviruses and evaluate their cross-reactivities for developing a future broad-protective vaccine.
(DKW)

Leveraging Synthetic Virology for the Rapid Engineering of Vesicular Stomatitis Virus

Vesicular stomatitis virus, a negative-sense, non-segmented RNA virus of Rhabdoviridae family is a prototype virus that is widely studied for advancing molecular virology. Replicative cycle of VSV is relatively quick, within 4-6 h and favorable for research in vaccines developments. However, there are several limitations among previous traditional technologies for viruses engineering (homologous recombination in plasmids) is taken long time periods and need significant research expertise, also restriction enzyme-based cloning methods are limited to the location of restriction sites. Thus, the study aimed to introduce a new methodology in a framework that resulted a quick design-construct and test the synthetic viruses. A full-length Indiana strain of VSV plasmid was used in this study to rescue the wildtype virus. In brief, the fragment sequence was designed (by rearrange the P and M gene order) to be inserted in plasmid vector for DNA synthesis and gene assembly. Then, it was transformed into selective medium and screened for successful cloning and plasmid construction. The successful cloning of synthesized VSV was transfected into the wild type of VSV and passaged. Last, the rescued products were quantified for gene copy number (by qPCR) and the cytopathic effect was observed under microscope.  As a result, 1.1 kilobases synthetic VSV showed no significant differences compared to the natural viruses on phenotypic analysis after rescue and titration. In conclusion, rapid engineering on synthetic VSV construction provides a novel development on vaccines and drug therapies that lead to improve human health.
(INV)

A single dose of an ALVAC vector based RABV virus-like particle candidate vaccine induces a potent immune response in mice, cats, and dogs

In this article, the authors developed a potential rabies vaccine, ALVAC-RABV-VLP, using CRISPR/Cas9 gene editing technology to reduce vaccine costs and provide a new and safer method of combating the highly fatal zoonotic disease. The vaccine, which uses a canarypox virus vector, was found to be highly potent in mice, cats, and dogs, eliciting a durable and effective humoral immune response compared to commercial ones. The ALVAC-RABBV-VLP effectively activated dendritic cells, follicular T helper cells, and germinal center / plasma cells. The survival rate was 100% in mice challenged with a lethal dose of RABV.

(SN)

Single Amino Acid Substitution in the Matrix Protein of Rabies Virus Is Associated with Neurovirulence in Mice

Rabies is a fatal zoonosis caused by the rabies virus (RABV), with 59,000 cases reported annually worldwide. The viral genome of RABV encodes N, P, M, G, and L structural proteins. The authors created a recombinant HEP (rHEP) based on the sequence of the rabies vaccine strain, HEP-Flury, and compared the growth of the two strains in MNA cells. They found a single amino acid substitution in the M protein (D80N) between the two strains and created rHEP-M(D80N). Mice inoculated with rHEP-M (D80N) showed significant weight loss compared to mice inoculated with rHEP. Furthermore, antigen levels in mouse brain were significantly higher in mice inoculated with rHEP-M(D80N). These results indicate that a single amino acid substitution in the M protein is important in the neuropathogenesis of RABV.
(SM)

Alternative pathway of complement activation has a beneficial role against Chandipura virus infection

Chandipura virus (CHPV) belongs to family Rhabdoviridae, genus Vesiculovirus and is one of the major causes of acute encephalitis in Indian pediatric population. The complement pathway is the first line of defense against pathogen including virus. The complement cascade can be initiated via three pathways; classical, lectin, or alternative pathway depending on virus recognition. These three pathways converge to activate C3, the central molecule of the complement cascade. C3-deficient human serum did not affect virus infectivity, indicating C3 is strongly dependent on CHPV neutralization. Furthermore, C5 is an initiator of MAC (membrane attack complex; lyses virus particles and infected cell) formation and is also found to be essential for CHPV neutralization. Since there is no specific antiviral treatment or vaccine for Chandipura, this study elucidating the role of the human complement system in CHPV neutralization may help in the design of effective treatment.
(TT)

Molecular mechanism of de novo replication of Ebolavirus Polymerase

Ebola virus, rabies virus, human respiratory syncytial virus, and pneumoviruses are of significant zoonotic and economic threats. They cause respiratory infections, hemorrhagic fever, and encephalitis. Large(L) Polymerase, a potential target for antiviral drug development, controls de novo replication of EBOV's L-polymerase. Researchers found that a minimum of 3 bases can drive RNA synthesis elongation, independent of the specific RNA sequence. This binds to the template entry channel with a distinctive tortuous stable bend conformation, enabling de novo replication activity.
(SN)