Triphala: An Indian Ayurvedic herbal formulation for coronavirus (SARS-CoV-2) disease (Covid-19)

This review paper updated the significance and pharmaceutical effects of Triphala as an alternative traditional herbal Indian folk medicine used as a immunity booster during the recent outbreak of coronavirus-2 (SARS-CoV-2) mutants, Delta (B.1.617.2) and Delta Plus (AY.1). The current outbreak of coronavirus-2 mutants, Delta (B.1.617.2) and Delta Plus (AY.1) is wreaking havoc in India. The new epicentre of the highest number of corona viral mutant infection cases and death rate has been recorded in Indonesia. The hallmark of the coronavirus disease is the cytokine storm, a massive cytokine and chemokine release due to an uncontrolled dysregulation of the host immune defence that causes loss of function of multiple organs and leading to death. One of the evidence to support Triphala alone inhibited the RNA viruses including human coronavirus. Triphala herbal formulation can reduce the production of progeny of human coronavirus, HCoV-NL63 particles and have an antiviral effect under in vitro conditions. In India, Triphala herbal formulation with an additional supplementation of pumpkin seeds, coconut water, sugar cane juice, Aloe vera juice, neem (Azardirachta indica) leaf juice, and melatonin rich diet has played an important role in controlling coronavirus disease than Triphala alone. However, clinical trials of Triphala with new additional supplementations is lacking for the scientific validation. On the basis literature survey, there is a ray of hope for the Triphala with additional supplementation as a new therapeutic drug for combating Covid-19 viral infections.

The HSV1 Tail-Anchored Membrane Protein pUL34 Contains a Basic Motif That Supports Active Transport to the Inner Nuclear Membrane Prior to Formation of the Nuclear Egress Complex

Herpes simplex virus type 1 nucleocapsids are released from the host nucleus by a budding process through the nuclear envelope called nuclear egress. Two viral proteins, the integral membrane proteins pUL34 and pUL31, form the nuclear egress complex at the inner nuclear membrane, which is critical for this process. The nuclear import of both proteins ensues separately from each other: pUL31 is actively imported through the central pore channel, while pUL34 is transported along the peripheral pore membrane. With this study, we identified a functional bipartite NLS between residues 178 and 194 of pUL34. pUL34 lacking its NLS is mislocalized to the TGN but retargeted to the ER upon insertion of the authentic NLS or a mimic NLS, independent of the insertion site. If co-expressed with pUL31, either of the pUL34-NLS variants is efficiently, although not completely, targeted to the nuclear rim where co-localization with pUL31 and membrane budding seem to occur, comparable to the wild-type. The viral mutant HSV1(17+)Lox-UL34-NLS mt is modestly attenuated but viable and associated with localization of pUL34-NLS mt to both the nuclear periphery and cytoplasm. We propose that targeting of pUL34 to the INM is facilitated by, but not dependent on, the presence of an NLS, thereby supporting NEC formation and viral replication.

Tradeoffs for a viral mutant with enhanced replication speed

RNA viruses exist as genetically heterogeneous populations due to high mutation rates, and many of these mutations reduce fitness and/or replication speed. However, it is unknown whether mutations can increase replication speed of a virus already well adapted to replication in cultured cells. By sequentially passaging coxsackievirus B3 in cultured cells and collecting the very earliest progeny, we selected for increased replication speed. We found that a single mutation in a viral capsid protein, VP1-F106L, was sufficient for the fast-replication phenotype. Characterization of this mutant revealed quicker genome release during entry compared to wild-type virus, highlighting a previously unappreciated infection barrier. However, this mutation also reduced capsid stability in vitro and reduced replication and pathogenesis in mice. These results reveal a tradeoff between overall replication speed and fitness. Importantly, this approach—selecting for the earliest viral progeny—could be applied to a variety of viral systems and has the potential to reveal unanticipated inefficiencies in viral replication cycles.

Tradeoffs for a viral mutant with enhanced replication speed

RNA viruses exist as genetically heterogeneous populations due to high mutation rates and many of these mutations reduce fitness and/or replication speed. However, it is unknown whether mutations can increase replication speed of a virus already well adapted to replication in cultured cells. By sequentially passaging coxsackievirus B3 in cultured cells and collecting the very earliest progeny, we selected for increased replication speed. We found that a single mutation in a viral capsid protein, VP1-F106L, was sufficient for the fast-replication phenotype. Characterization of this mutant revealed quicker genome release during entry compared to wild-type virus, highlighting a previously unappreciated infection barrier. However, this mutation also reduced capsid stability in vitro and reduced replication and pathogenesis in mice. These results reveal a tradeoff between overall replication speed and fitness. Importantly, this approach— selecting for the earliest viral progeny—could be applied to a variety of viral systems and has the potential to reveal unanticipated inefficiencies in viral replication cycles.SignificanceViruses have characteristic replication speeds within a given cell type. Many factors can slow the rate of viral replication, including attenuating mutations and host antiviral responses. However, it has been unclear whether it would be possible to “speed up” a virus that already replicates efficiently in a specific cell type. Here, we selected for a mutant coxsackievirus with enhanced replication speed by sequentially harvesting the very earliest progeny in multiple rounds of selection. A single mutation conferred the fast replication phenotype. While this mutant virus has enhanced replication in cultured cells due to faster genome uncoating, it was attenuated in mice. These results highlight selective pressures that shape viral populations in different environments.

Ivermectin for COVID-19 in Peru: 14-fold reduction in nationwide excess deaths, p=.002 for effect by state, then 13-fold increase after ivermectin use restricted

Introduction. On May 8, 2020, Peru’s Ministry of Health approved ivermectin (IVM), a drug of Nobel Prize-honored distinction, for inpatient and outpatient treatment of COVID-19. As IVM treatments proceeded in that nation of 33 million residents, excess deaths decreased 14-fold over four months through December 1, 2020, consistent with clinical benefits of IVM for COVID-19 found in several RCTs. But after IVM use was sharply restricted under a new president, excess deaths then increased 13-fold. Methods. To evaluate possible IVM treatment effects suggested by these aggregate trends, excess deaths were analyzed by state for ages ≥ 60 in Peru’s 25 states. To identify potential confounding factors, Google mobility data, population densities, SARS-CoV-2 genetic variations and seropositivity rates were also examined.Results. The 25 states of Peru were grouped by extent of IVM distributions: maximal (mass IVM distributions through operation MOT, a broadside effort led by the army); medium (locally managed IVM distributions); and minimal (restrictive policies in one state, Lima). The mean reduction in excess deaths 30 days after peak deaths was 74% for the maximal IVM distribution group, 53% for the medium group and 25% for Lima. Reduction of excess deaths is correlated with extent of IVM distribution by state with a p value of 0.002 using the Kendall τb test, well below the confidence threshold of 0.05 for an established clinical effect.Conclusion. Mass treatments with IVM, a drug safely used in 3.7 billion doses worldwide since 1987, most likely caused the 14-fold reductions in excess deaths in Peru, prior to their 13-fold increase under reversed IVM policy. This strongly suggests that IVM treatments can likewise effectively complement immunizations to help eradicate COVID-19. The indicated biological mechanism of IVM, competitive binding with SARS-CoV-2 spike protein, is likely non-epitope specific, possibly yielding full efficacy against emerging viral mutant strains.

The time for COVID-19 vaccination

The composition and dynamics of viral mutant spectra in infected individuals advice that to avoid selection of SARS-CoV-2 escape mutants, vaccination campaigns for COVID-19 should be launched when disease incidence is low.

Evolutionary Genetics of Mycobacterium tuberculosis and HIV-1: “The Tortoise and the Hare”

The already enormous burden caused by Mycobacterium tuberculosis and Human Immunodeficiency Virus type 1 (HIV-1) alone is aggravated by co-infection. Despite obvious differences in the rate of evolution comparing these two human pathogens, genetic diversity plays an important role in the success of both. The extreme evolutionary dynamics of HIV-1 is in the basis of a robust capacity to evade immune responses, to generate drug-resistance and to diversify the population-level reservoir of M group viral subtypes. Compared to HIV-1 and other retroviruses, M. tuberculosis generates minute levels of genetic diversity within the host. However, emerging whole-genome sequencing data show that the M. tuberculosis complex contains at least nine human-adapted phylogenetic lineages. This level of genetic diversity results in differences in M. tuberculosis interactions with the host immune system, virulence and drug resistance propensity. In co-infected individuals, HIV-1 and M. tuberculosis are likely to co-colonize host cells. However, the evolutionary impact of the interaction between the host, the slowly evolving M. tuberculosis bacteria and the HIV-1 viral “mutant cloud” is poorly understood. These evolutionary dynamics, at the cellular niche of monocytes/macrophages, are also discussed and proposed as a relevant future research topic in the context of single-cell sequencing.

Atmospheric PM2.5 before and after Lockdown in relation to COVID-19 Evolution and daily Viral Counts: Could Viral Natural Selection have occurred due to changes in the Airborne Pollutant PM2.5 acting as a Vector for SARS-CoV-2?

BackgroundGenes coding for SARS-CoV-2 have been detected on the microscopic airborne pollutant particulate matter, which has been suggested as a vector for COVID-19 transmission. Lockdown in China has been shown to be associated with significant reduction in pollution including the particulate matter component which coincided with the appearance of a viral mutant (Clade G) which steadily displaced the original Clade D after lockdown. The reason why Clade G developed a fitness advantage is as yet unknown. This paper examines the possible role of airborne particulate matter PM2.5 as selective pressure determining viral Clade predominance and further shedding light on the mode of SARS-CoV-2 transmission.MethodsThe average levels of PM2.5 of a number of cities were obtained from the Air Quality Index (AQI), a real-time assessment of atmospheric pollution. The daily average PM2.5 levels were assessed between January 23rd and April 29th 2020 determined by the timeline when viral counts in Beijing and other cities were available. Daily viral counts of Clades D and G were available starting from the 12th February as determined by the scientific literature published in August 2020. The cities chosen were Beijing, Sheffield, Nottingham, Sydney and Cambridge because of their substantially elevated viral counts compared to other cities. Cities as opposed to vaster areas/nations were chosen as PM2.5 levels vary across regions and countries.ResultsFor the time period assessed, the Beijing PM2.5 pattern initiated with highly elevated mean PM2.5 levels of 155.8µg/m3 (SD+/-73.6) during high viral counts, followed by 82.1µg/m3 (SD+/-44.9) (p<0.04) when the viral counts decreased. In all the other cities assessed, the pattern differed whereby the PM2.5 levels increased significantly over the preceding baseline contemporaneously with the viral count rise. The changes in these cities’ PM2.5 levels were on average 31.5µg/m3 before viral counts rose and 56.35µg/m3 contemporaneous with viral count rise. The average levels of PM2.5 in these cities started to decrease one week after lockdown to 46µg/m3 when measured over 2 weeks post-lockdown.As regards the viral counts from data retrieved from Beijing, the latter part of the bell-shaped curve and a subsequent smaller curve of the viral count was available for evaluation. The average viral count for Clade D in Beijing was 11.1(SD+/-13.5) followed by a mean viral count for Clade G was 13.8(SD+/-9.2). Conversely in all the other cities besides Beijing, the viral counts averaged 45.8 for Clade D and 161 for Clade G. The variation in viral counts between cities suggests the strong possibility of variation in the availability of sampling between cities.The newer variant, Clade G demonstrated viral counts initially appearing in mid-February in Beijing to later displace Clade D as the dominant viral Clade. The appearance of Clade G coincided with the decreasing gradient of PM2.5 levels. A number of significant correlations were obtained between PM2.5 levels and the viral count in all the cities reviewed.ConclusionCOVID-19 viral counts appear to increase concomitant with increasing PM2.5 levels. Viral counts of both Clades correlated differentially with PM2.5 levels in all the cities assessed. The significantly highly elevated PM2.5 levels in Beijing resulted in correlating mainly with Clade D, however Clade G began to appear with decreasing PM2.5 levels, suggesting the beginnings for the initial SARS-CoV-2 Clade evolution. Clade G, the newer variant was able to flourish at lower levels of PM2.5 than Clade D. Clade G may possibly have utilized other sources of particulate matter as a viral vector, such as that derived from tobacco smoking, whereby 66% of Chinese males are smokers and 70% of the Chinese non-smoking population are exposed to 2nd hand smoking.

Development of a cricket paralysis virus-based system for inducing RNA interference-mediated gene silencing in Diaphorina citri

Diaphorina citri, the Asian citrus psyllid, is the insect vector of the phloem-limited bacterium ‘Candidatus Liberibacter asiaticus’, which causes the most devastating citrus disease worldwide: Huanglongbing (HLB). An efficient cure for HLB is still not available and the management of the disease is restricted to the use of pesticides, antibiotics and eradication of infected plants. Plant- and insect-infecting viruses have attracted increasing attention for their potential to manipulate traits in insects, especially insect vectors of plant pathogens. However, so far there are no insect virus-based vectors available for use in D. citri. Cricket paralysis virus (CrPV) is a well-studied insect-infecting dicistrovirus with a wide host range and has been used as a model in previous translational studies. In this work, we demonstrate for the first time that CrPV is infectious and pathogenic to D. citri. We show that specific amino acid mutations in the CrPV primary cleavage DvExNPGP motif resulted in a viral mutant that was attenuated compared to wild-type CrPV during infection of either Drosophila cells line or adult D. citri insects. This attenuated CrPV mutant was then used as the backbone for engineering a recombinant CrPV-based vector to specifically alter D. citri gene expression via the RNA interference (RNAi) pathway, a technology called Virus Induced Gene Silencing (VIGS). As proof-of-concept, we engineered recombinant CrPV-based vectors carrying nucleotide sequences derived from a previously reported D. citri target gene: the inhibitor of apoptosis gene (IA). RT-qPCR analysis of insects either microinjected or fed with the recombinant CrPV mutants showed decreased IA gene expression as soon as viral replication was detected, indicating that the engineered CrPV-based VIGS system enables functional gene silencing in D. citri. This novel insect virus-based tool is easily amenable to genomic modification and represents a technical advance for understanding interactions between insect virus-based VIGS systems and D. citri.