S-adenosylmethionine-dependent methyltransferase inhibitor DZNep blocks transcription and translation of SARS-CoV-2 genome with a low tendency to select for drug-resistant viral variants.

We report the in vitro antiviral activity of DZNep (3-Deazaneplanocin A; an inhibitor of S-adenosylmethionine-dependent methyltransferase) against SARS-CoV-2, besides demonstrating its protective efficacy against lethal infection of infectious bronchitis virus (IBV, a member of the Coronaviridae family). DZNep treatment resulted in reduced synthesis of SARS-CoV-2 RNA and proteins without affecting other steps of viral life cycle. We demonstrated that deposition of N6-methyl adenosine (m6A) in SARS-CoV-2 RNA in the infected cells recruits heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), an RNA binding protein which serves as a m6A reader. DZNep inhibited the recruitment of hnRNPA1 at m6A-modified SARS-CoV-2 RNA which eventually suppressed the synthesis of the viral genome. In addition, m6A-marked RNA and hnRNPA1 interaction was also shown to regulate early translation to replication switch of SARS-CoV-2 genome. Furthermore, abrogation of methylation by DZNep also resulted in defective synthesis of the 5' cap of viral RNA, thereby resulting in its failure to interact with eIF4E (a cap-binding protein), eventually leading to a decreased synthesis of viral proteins. Most importantly, DZNep-resistant mutants could not be observed upon long-term sequential passage of SARS-CoV-2 in cell culture. In summary, we report the novel role of methylation in the life cycle of SARS-CoV-2 and propose that targeting the methylome using DZNep could be of significant therapeutic value against SARS-CoV-2 infection.

Polyalthia longifolia leaves methanolic extract targets entry and budding of viruses-an in vitro experimental study against paramyxoviruses.

ETHNOPHARMACOLOGICAL RELEVANCE: Synthetic antiviral drugs have several limitations including high cost. Thus research on antiviral property of medicinal plants is continuously gaining importance. Polyalthia longifolia possesses several medicinal properties and has been used in traditional ayurvedic medicine for treatment of dermatological ailments as kushta, visarpa/herpes virus infection and also to treat pyrexia of unknown origin as mentioned in Visarpa Chikitsa. AIM OF THE STUDY: Keeping in view the cytotoxic, anti-cancer activity and antiviral efficacy of Polyalthia longifolia against herpes, present study was undertaken to evaluate the in vitro antiviral activity of methanolic extract of Polyalthia longifolia leaves, if any, and to unravel the possible target(s)/mechanism of action. MATERIAL AND METHODS: Antiviral activity of Polyalthia longifolia methanolic extract was studied using Vero cell lines against paramyxoviruses, namely-peste des petits ruminants virus (PPRV) and Newcastle disease virus (NDV). Cytotoxicity of the test extract was evaluated employing MTT assay. Virucidal activity, and viral-attachment, virus entry and release assays were determined in Vero cells using standard experimental protocols. The viral RNA in the virus-infected cells was quantified by qRT-PCR. RESULTS: At non-cytotoxic concentration, methanolic extract of Polyalthia longifolia leaves was found to inhibit the replication of PPRV and NDV at viral entry and budding level, whereas other steps of viral life cycle such as attachment and RNA synthesis remained unaffected. CONCLUSIONS: Polyalthia longifolia leaves extract possesses promising antiviral activity against paramyxoviruses and acts by inhibiting the entry and budding of viruses; and this plant extract evidently possesses excellent and promising potential for development of effective herbal antiviral drug.

Sequence and phylogenetic analysis of host-range (E3L, K3L, and C7L) and structural protein (B5R) genes of buffalopox virus isolates from buffalo, cattle, and human in India.

Buffalopox virus (BPXV), a close variant of vaccinia virus (VACV) has emerged as a zoonotic pathogen. The host tropism of poxviruses is governed by host-range genes. Among the host-range genes: E3L, K3L, and C7L are essential for virus replication by preventing interferon resistance, whereas B5R is essential for spread of the virus and evasion from the host's immune response as in VACV. We report sequence analysis of host-range genes: E3L, K3L, C7L, and membrane protein gene (B5R) of BPXVs from buffalo, cattle, and human from recent outbreaks in India-their phylogenetic relationship with reference strain (BP4) and other Orthopoxviruses. BPXVs revealed a sequence homology with VACVs including zoonotic Brazilian VACV-like viruses. The aa sequences of E3L and K3L genes were 100 % similar in buffalo, cattle, and human isolates. However, four significant point mutations (I11K; N12K and S36F in C7L gene and D249G in B5R gene) were observed specific to buffalo isolate only. This signifies that different strains of BPXV were circulated during the outbreak. The mutations in C7L and B5R could play an important role in adaptation of BPXV in human and cattle which needs further functional studies. The strain of BPXV isolated from buffalo may not be adopted in human and cow. Various point mutations were observed in the host-range genes of reference strain (BPXV-BP4) which may be due to several passages of virus in cell culture. The phylogeny constructed based on concatenated gene sequences revealed that BPXVs are not as closely related to vaccine strain (Lister and Lister-derived strain-LC16m8), as hypothesized earlier, rather they are more closely related to reference strain (BPXV-BP4) and other vaccinia and vaccinia-like viruses such as Passatempo and Aracatuba viruses. The availability of information regarding host tropism determinants would allow us to understand molecular mechanism of species tropism of poxviruses which would be useful in unveiling new strategies to control zoonotic poxviral infections.