Latent Virus Research Articles

Gene editing for latent herpes simplex virus infection reduces viral load and shedding in vivo

Anti-HSV therapies are only suppressive because they do not eliminate latent HSV present in ganglionic neurons, the source of recurrent disease. We have developed a potentially curative approach against HSV infection, based on gene editing using HSV-specific meganucleases delivered by adeno-associated virus (AAV) vectors. Gene editing performed with two anti-HSV-1 meganucleases delivered by a combination of AAV9, AAV-Dj/8, and AAV-Rh10 can eliminate 90% or more of latent HSV DNA in mouse models of orofacial infection, and up to 97% of latent HSV DNA in mouse models of genital infection. Using a pharmacological approach to reactivate latent HSV-1, we demonstrate that ganglionic viral load reduction leads to a significant decrease of viral shedding in treated female mice. While therapy is well tolerated, in some instances, we observe hepatotoxicity at high doses and subtle histological evidence of neuronal injury without observable neurological signs or deficits. Simplification of the regimen through use of a single serotype (AAV9) delivering single meganuclease targeting a duplicated region of the HSV genome, dose reduction, and use of a neuron-specific promoter each results in improved tolerability while retaining efficacy. These results reinforce the curative potential of gene editing for HSV disease.

Chronic and Latent Viral Infections and Leukocyte Telomere Length across the Lifespan of Female and Male Individuals Living with or without HIV

Background: Chronic/latent viral infections may accelerate immunological aging, particularly among people living with HIV (PLWH). We characterized chronic/latent virus infections across their lifespan and investigated their associations with leukocyte telomere length (LTL). Methods: Participants enrolled in the CARMA cohort study were randomly selected to include n = 15 for each decade of age between 0 and >60 y, for each sex, and each HIV status. Cytomegalovirus (CMV), Epstein–Barr virus (EBV), human herpesvirus 8 (HHV-8), herpes simplex virus 1 (HSV-1), and HSV-2 infection were determined serologically; HIV, hepatitis C (HCV), and hepatitis B (HBV) were self-reported. LTLs were measured using monochrome multiplex qPCR. Associations between the number of viruses, LTL, and sociodemographic factors were assessed using ordinal logistic and linear regression modeling. Results: The study included 187 PLWH (105 female/82 male) and 190 HIV-negative participants (105 female/84 male), ranging in age from 0.7 to 76.1 years. Living with HIV, being older, and being female were associated with harbouring a greater number of chronic/latent non-HIV viruses. Having more infections was in turn bivariately associated with a shorter LTL. In multivariable analyses, older age, living with HIV, and the female sex remained independently associated with having more infections, while having 3–4 viruses (vs. 0–2) was associated with a shorter LTL. Conclusions: Our results suggest that persistent viral infections are more prevalent in PLWH and females, and that these may contribute to immunological aging. Whether this is associated with comorbidities later in life remains an important question.

Whole genome sequence analysis of shallot virus X from India reveals it to be a natural recombinant with positive selection pressure

BackgroundShallots are infected by various viruses like Onion yellow dwarf virus (OYDV), Leek yellow stripe virus (LYSV), Shallot latent virus (SLV) and Shallot virus X (ShVX). In India, they have been found to be persistently infected by ShVX. ShVX also infects onion and garlic in combination with other carlaviruses and potyviruses. ShVX is a member of genus Allexivirus of family Alphaflexiviridae. ShVX has a monopartite genome, which is represented by positive sense single-stranded RNA. Globally, only six complete and 3 nearly complete genome sequences of ShV X are reported to date. This number is insufficient to measure a taxon’s true molecular diversity. Moreover, the complete genome sequence of ShVX from Asia has not been reported as yet. Therefore, this study was undertaken to generate a complete genome sequence of ShVX from India.ResultsShallot virus X (ShVX) is one of the significant threats to Allium crop production. In this study, we report the first complete genome sequence of the ShVX from India through Next-generation sequencing (NGS). The complete genome of the ShVX (Accession No. OK104171), from this study comprised 8911 nucleotides. In-silico analysis of the sequence revealed variability between this isolate and isolates from other countries. The dissimilarities are spread all over the genome specifically some non-coding intergenic regions. Statistical analysis of individual genes for site-specific selection indicates a positive selection in NABP region. The presence of a recombination event was detected in coat protein region. The sequence similarity percentage and phylogenetic analysis indicate ShVX Indian isolate is a distinctly different isolate. Recombination and site-specific selection may have a function in the evolution of this isolate. This is the first detailed study of the ShVX complete genome sequence from Southeast Asia.ConclusionThis study presents the first report of the entire genome sequence of an Indian isolate of ShVX along with an in-depth exploration of its evolutionary traits. The findings highlight the Indian variant as a naturally occurring recombinant, emphasizing the substantial role of recombination in the evolution of this viral species. This insight into the molecular diversity of strains within a specific geographical region holds immense significance for comprehending and forecasting potential epidemics. Consequently, the insights garnered from this research hold practical value for shaping ShVX management strategies and providing a foundation for forthcoming studies delving into its evolutionary trajectory.

A novel and efficient Apple latent spherical virus-based gene silencing method for functional genomic studies in Chenopodium quinoa

Quinoa (Chenopodium quinoa Willd.), with its resilience in harsh environments and excellent nutritional value, has become crucial for global food security. Despite recent progress in genomic research, the inability to perform functional studies in quinoa due to the absence of transformation techniques remains a significant obstacle. In this work, we present the development of a novel Apple Latent Spherical Virus (ALSV)-mediated virus-induced gene silencing (VIGS) protocol that will allow to perform functional genomics studies in quinoa in a fast and simple way. The method was fine-tuned using ALSV plasmids that carry partial gene sequences of phytoene desaturase (PDS) from Nicotiana benthamiana and quinoa. The developed technique involves an initial inoculation in Nicotiana plants through agroinfiltration with Agrobacterium tumefaciens cultures carrying the different viral constructs. Viral extracts were prepared using local or systemic leaves, which were then used as inoculum to infect quinoa leaves through mechanical damage. The method was successfully tested in two contrasting quinoa varieties, although some differences were observed in infection phenotype and viral susceptibility. The effect of insertion sequence size in the viral vectors was also analyzed, resulting in differences in bleaching or chlorosis phenotype and impact on plant growth. The presence of the virus in inoculated plants was confirmed, and the reduction in PDS gene expression in silenced plants was verified. Because quinoa lacks stable transformation protocols, limiting heterologous expression assays, our ALSV-based VIGS protocol is very attractive for loss-of-function gene studies.

Epigenetic reprograming in myalgic encephalomyelitis/chronic fatigue syndrome: A narrative of latent viruses.

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a chronic disease presenting with severe fatigue, post-exertional malaise, and cognitive disturbances-among a spectrum of symptoms-that collectively render the patient housebound or bedbound. Epigenetic studies in ME/CFS collectively confirm alterations and/or malfunctions in cellular and organismal physiology associated with immune responses, cellular metabolism, cell death and proliferation, and neuronal and endothelial cell function. The sudden onset of ME/CFS follows a major stress factor that, in approximately 70% of cases, involves viral infection, and ME/CFS symptoms overlap with those of long COVID. Viruses primarily linked to ME/CFS pathology are the symbiotic herpesviruses, which follow a bivalent latent-lytic lifecycle. The complex interaction between viruses and hosts involves strategies from both sides: immune evasion and persistence by the viruses, and immune activation and viral clearance by the host. This dynamic interaction is imperative for herpesviruses that facilitate their persistence through epigenetic regulation of their own and the host genome. In the current article, we provide an overview of the epigenetic signatures demonstrated in ME/CFS and focus on the potential strategies that latent viruses-particularly Epstein-Barr virus-may employ in long-term epigenetic reprograming in ME/CFS. Epigenetic studies could aid in elucidating relevant biological pathways impacted in ME/CFS and reflect the physiological variations among the patients that stem from environmental triggers, including exogenous viruses and/or altered viral activity.

Herpes Simplex Keratitis after PRK

Background: Herpes simplex virus (HSV) is a double stranded DNA virus that infects much of the world’s population. The World Health Organization estimates 67% of people under the age of 50 have HSV-1 and 11% of people under the age of 50 have HSV-2. While most systemic infections remain inactive and do not cause symptoms, the activation and appearance of ocular HSV can be a destructive and vision-threatening issue. HSV keratitis can be epithelial, stromal or a combination. Patients who have recently undergone photorefractive keratectomy (PRK) have a higher risk of HSV keratitis. Because of this, HSV keratitis should be considered in any PRK post-op patient that presents with unusual signs and symptoms such as decreased vision, increased IOPs, ocular pain, photophobia, iritis and corneal haze. When diagnosed in a timely manner, the prognosis is generally favorable. Current treatments for HSV keratitis include antiviral coverage as well as immunosuppression with topical steroids. Case Report: This case involves a patient that developed HSV keratitis while on topical steroid therapy following PRK. The weeks following the initial presentation were spent battling corneal haze, iritis, pain, increased IOPs and decreased visual acuity. After close monitoring and treatment changes based on each clinical presentation, the patient recovered with good vision and minimal corneal scarring. The findings and recommended treatment options are discussed. Conclusion: HSV affects much of the world’s population. Reactivation of the latent virus can lead to vision threatening corneal disease. HSV is capable of causing inflammation and pathology throughout most of the anterior segment of the eye, and therefore should be considered when managing any new or recalcitrant anterior pathology. Although the diagnosis is usually clinical, laboratory testing can be used to confirm a diagnosis. Treatment aims to decrease the viral load and to limit stromal scarring.

Germinal center cytokines driven epigenetic control of Epstein-Barr virus latency gene expression.

Epstein-Barr virus (EBV) persistently infects 95% of adults worldwide and is associated with multiple human lymphomas that express characteristic EBV latency programs used by the virus to navigate the B-cell compartment. Upon primary infection, the EBV latency III program, comprised of six Epstein-Barr Nuclear Antigens (EBNA) and two Latent Membrane Protein (LMP) antigens, drives infected B-cells into germinal center (GC). By incompletely understood mechanisms, GC microenvironmental cues trigger the EBV genome to switch to the latency II program, comprised of EBNA1, LMP1 and LMP2A and observed in GC-derived Hodgkin lymphoma. To gain insights into pathways and epigenetic mechanisms that control EBV latency reprogramming as EBV-infected B-cells encounter microenvironmental cues, we characterized GC cytokine effects on EBV latency protein expression and on the EBV epigenome. We confirmed and extended prior studies highlighting GC cytokine effects in support of the latency II transition. The T-follicular helper cytokine interleukin 21 (IL-21), which is a major regulator of GC responses, and to a lesser extent IL-4 and IL-10, hyper-induced LMP1 expression, while repressing EBNA expression. However, follicular dendritic cell cytokines including IL-15 and IL-27 downmodulate EBNA but not LMP1 expression. CRISPR editing highlighted that STAT3 and STAT5 were necessary for cytokine mediated EBNA silencing via epigenetic effects at the EBV genomic C promoter. By contrast, STAT3 was instead necessary for LMP1 promoter epigenetic remodeling, including gain of activating histone chromatin marks and loss of repressive polycomb repressive complex silencing marks. Thus, EBV has evolved to coopt STAT signaling to oppositely regulate the epigenetic status of key viral genomic promoters in response to GC cytokine cues.

Direct and indirect effects of CYTOR lncRNA regulate HIV gene expression.

The implementation of antiretroviral therapy (ART) has effectively restricted the transmission of Human Immunodeficiency Virus (HIV) and improved overall clinical outcomes. However, a complete cure for HIV remains out of reach, as the virus persists in a stable pool of infected cell reservoir that is resistant to therapy and thus a main barrier towards complete elimination of viral infection. While the mechanisms by which host proteins govern viral gene expression and latency are well-studied, the emerging regulatory functions of non-coding RNAs (ncRNA) in the context of T cell activation, HIV gene expression and viral latency have not yet been thoroughly explored. Here, we report the identification of the Cytoskeleton Regulator (CYTOR) long non-coding RNA (lncRNA) as an activator of HIV gene expression that is upregulated following T cell stimulation. Functional studies show that CYTOR suppresses viral latency by directly binding to the HIV promoter and associating with the cellular positive transcription elongation factor (P-TEFb) to activate viral gene expression. CYTOR also plays a global role in regulating cellular gene expression, including those involved in controlling actin dynamics. Depletion of CYTOR expression reduces cytoplasmic actin polymerization in response to T cell activation. In addition, treating HIV-infected cells with pharmacological inhibitors of actin polymerization reduces HIV gene expression. We conclude that both direct and indirect effects of CYTOR regulate HIV gene expression.

Post-Transcriptional HIV-1 Latency: A Promising Target for Therapy?

Human Immunodeficiency Virus type 1 (HIV-1) latency represents a significant hurdle in finding a cure for HIV-1 infections, despite tireless research efforts. This challenge is partly attributed to the intricate nature of HIV-1 latency, wherein various host and viral factors participate in multiple physiological processes. While substantial progress has been made in discovering therapeutic targets for HIV-1 transcription, targets for the post-transcriptional regulation of HIV-1 infections have received less attention. However, cumulative evidence now suggests the pivotal contribution of post-transcriptional regulation to the viral latency in both in vitro models and infected individuals. In this review, we explore recent insights on post-transcriptional latency in HIV-1 and discuss the potential of its therapeutic targets, illustrating some host factors that restrict HIV-1 at the post-transcriptional level.

Latent Epstein-Barr virus infection collaborates with Myc over-expression in normal human B cells to induce Burkitt-like Lymphomas in mice.

Epstein-Barr virus (EBV) is an important cause of human lymphomas, including Burkitt lymphoma (BL). EBV+ BLs are driven by Myc translocation and have stringent forms of viral latency that do not express either of the two major EBV oncoproteins, EBNA2 (which mimics Notch signaling) and LMP1 (which activates NF-κB signaling). Suppression of Myc-induced apoptosis, often through mutation of the TP53 (p53) gene or inhibition of pro-apoptotic BCL2L11 (BIM) gene expression, is required for development of Myc-driven BLs. EBV+ BLs contain fewer cellular mutations in apoptotic pathways compared to EBV-negative BLs, suggesting that latent EBV infection inhibits Myc-induced apoptosis. Here we use an EBNA2-deleted EBV virus (ΔEBNA2 EBV) to create the first in vivo model for EBV+ BL-like lymphomas derived from primary human B cells. We show that cord blood B cells infected with both ΔEBNA2 EBV and a Myc-expressing vector proliferate indefinitely on a CD40L/IL21 expressing feeder layer in vitro and cause rapid onset EBV+ BL-like tumors in NSG mice. These LMP1/EBNA2-negative Myc-driven lymphomas have wild type p53 and very low BIM, and express numerous germinal center B cell proteins (including TCF3, BACH2, Myb, CD10, CCDN3, and GCSAM) in the absence of BCL6 expression. Myc-induced activation of Myb mediates expression of many of these BL-associated proteins. We demonstrate that Myc blocks LMP1 expression both by inhibiting expression of cellular factors (STAT3 and Src) that activate LMP1 transcription and by increasing expression of proteins (DNMT3B and UHRF1) known to enhance DNA methylation of the LMP1 promoters in human BLs. These results show that latent EBV infection collaborates with Myc over-expression to induce BL-like human B-cell lymphomas in mice. As NF-κB signaling retards the growth of EBV-negative BLs, Myc-mediated repression of LMP1 may be essential for latent EBV infection and Myc translocation to collaboratively induce human BLs.

First Report of Sweet Potato Virus E(SPVE) Infecting Sweet Potato in China.

Sweet potato (Ipomoea batatas [L.] Lam.) is a versatile crop, cultivated in the subtropical and tropical areas, as food, fodder, and industrial raw material crop. In China, sweet potato has been used as a health-care food in recent years, as it contains a wide range of nutrients and xenobiotic phytochemicals. However, viral diseases are major constraint for the sweet potato yield and quality, especially the seed production and quality. Over 30 species of viruses infect sweet potato worldwide (Clark et al. 2012). More recently, a few new viruses infected sweet potato were identified, such as sweet potato virus E (SPVE), which was reported in Korea(Jo et al. 2020). In May 2022, a sweet potato sample (JSXZ1) with virus-like symptom, such as mosaic and vein clearing were collected from sweet potato germplasm Xuzhou resource nursery, Jiangsu Province, China (N34˚16', E117˚18') (Fig. S1A). To investigate the virus disease, the sample JSXZ1 showing the typical symptoms of disease was prepared for Small-RNA (sRNA) deep-sequencing. The sRNA library was constructed using TruSeq™ Small RNA Sample Prep Kits (Illumina, San Diego, USA) and sequenced using the Illumine Hiseq 2500 platform by LC-Bop Technologies (Hangzhou) CO., LTD. The sample was sequenced to obtain 26, 358, 439 raw reads and 22, 969, 139 clean reads after quality control trimming and analysis. The Velvet 1.0.5 software was used to de novo assemble the clean reads (18 to 28 nt) into larger contigs, which were then compared with the nucleotide sequences in the National Center for Biotechnology Information (NCBI) database using the BLASTn algorithm. Viruses found in the sample were sweet potato latent virus (SPLV), sweet potato feathery mottle virus (SPFMV), sweet potato chlorotic stunt virus (SPCSV), sweet potato badnavirus A (SPBV-A) and sweet potato badnavirus B (SPBV-B). Surprisingly, besides the viruses listed above, 28 contigs matched sequences of SPVE isolate GS (MH388502). To verify the result, total RNA was extracted from the sample JSXZ1 and from other leave samples (JSXZ2-JSXZ5) that contained SPFMV, SPVC, SPLV, SPVG respectively stored in lab using FastPure Universal Plant Total RNA Isolation Kit (Vazyme Biotech Co., LTD, Nanjing, China). cDNA was synthesized using random primer (hexadeoxyribonucleotide mixture; pd(N)6). The cDNA serves as template in PCR using a newly designed primer pairs based on SPVE p1 gene (SPVE-F: 5'- TCACCAAAAAGAATGCTACAAC-3'/SPVE-R: 5'-GAAATCCTCCCACTCTCCATA-3'). An expected ~500-bp PCR fragment was obtained in JSXZ1, while none of the fragment was obtained from JSXZ2-JSXZ5 (Fig. S1B). The PCR fragment was cloned into pMD18-T vector (Takara Bio Inc., Beijing, China) and plasmid DNA from transformed Escherichia coli DH5α cell (n=3) were commercially sequenced by Sangon Biotech (Shanghai) Co., Ltd. The sequences of the three fragment clones we obtained were 100% identical when compared. A BLASTN analysis of the sequences revealed that they are specific to SPVE and shared 98.62% nucleotide identity to SPVE GS isolate (MH388502) and one sequence was submitted to GenBank (Accession number OQ948331). To determine the occurrence of SPVE in infected sweet potato plants, a total of 37 leaves samples with viral symptom collected from Shandong Province (n=6) and Jiangsu Province (n=31) were indexed by RT-PCR as described before. Only 9 (24.3%) out of 37 from Shandong (n=1) and Jiangsu (n=8) were positive to SPVE respectively. In addition, five additional viruses (SPFMV, SPVC, SPVG, SPLV, SPCSV) were detected among these 37 samples and always in a mixed infection of two or more viruses. To our knowledge, this is the first report of SPVE infecting sweet potato in China. Sweet potato is an important crop in China and other countries (Zhang et al. 2023). China is the largest sweet potato producer all over the world. In addition, as sweet potato is produced through the vegetative propagation mode, thus, more attention should be paid to detection and monitoring of occurrence of SPVE in China.

Long COVID and post-acute sequelae of SARS-CoV-2 pathogenesis and treatment: A Keystone Symposia report.

In 2023, the Keystone Symposia held the first international scientific conference convening research leaders investigating the pathology of post-acute sequelae of COVID-19 (PASC) or Long COVID, a growing and urgent public health priority. In this report, we present insights from the talks and workshops presented during this meeting and highlight key themes regarding what researchers have discovered regarding the underlying biology of PASC and directions toward future treatment. Several themes have emerged in the biology, with inflammation and other immune alterations being the most common focus, potentially related to viral persistence, latent virus reactivation, and/or tissue damage and dysfunction, especially of the endothelium, nervous system, and mitochondria. In order to develop safe and effective treatments for people with PASC, critical next steps should focus on the replication of major findings regarding potential mechanisms, disentangling pathogenic mechanisms from downstream effects, development of cellular and animal models, mechanism-focused randomized, placebo-controlled trials, and closer collaboration between people with lived experience, scientists, and other stakeholders. Ultimately, by learning from other post-infectious syndromes, the knowledge gained may help not only those with PASC/Long COVID, but also those with other post-infectious syndromes.

A macrophage-cell model of HIV latency reveals the unusual importance of the bromodomain axis

BackgroundAlthough macrophages are now recognized as an essential part of the HIV latent reservoir, whether and how viral latency is established and reactivated in these cell types is poorly understood. To understand the fundamental mechanisms of viral latency in macrophages, there is an urgent need to develop latency models amenable to genetic manipulations and screening for appropriate latency-reversing agents (LRAs). Given that differentiated THP-1 cells resemble monocyte-derived macrophages in HIV replication mechanisms, we set out to establish a macrophage cell model for HIV latency using THP-1 cells.MethodsWe created single-cell clones of THP-1 cells infected with a single copy of the dual-labeled HIVGKO in which a codon switched eGFP (csGFP) is under the control of the HIV-1 5’ LTR promoter, and a monomeric Kusabira orange 2 (mKO2) under the control of cellular elongation factor one alpha promoter (EF1α). Latently infected cells are csGFP−, mKO2+, while cells with actively replicating HIV (or reactivated virus) are csGFP+,mKO2+. After sorting for latently infected cells, each of the THP-1 clones with unique integration sites for HIV was differentiated into macrophage-like cells with phorbol 12-myristate 13-acetate (PMA) and treated with established LRAs to stimulate HIV reactivation. Monocyte-derived macrophages (MDMs) harboring single copies of HIVGKO were used to confirm our findings.ResultsWe obtained clones of THP-1 cells with latently infected HIV with unique integration sites. When the differentiated THP-1 or primary MDMs cells were treated with various LRAs, the bromodomain inhibitors JQ1 and I-BET151 were the most potent compounds. Knockdown of BRD4, the target of JQ1, resulted in increased reactivation, thus confirming the pharmacological effect. The DYRK1A inhibitor Harmine and lipopolysaccharide (LPS) also showed significant reactivation across all three MDM donors. Remarkably, LRAs like PMA/ionomycin, bryostatin-1, and histone deacetylase inhibitors known to potently reactivate latent HIV in CD4 + T cells showed little activity in macrophages.ConclusionsOur results indicate that this model could be used to screen for appropriate LRAs for macrophages and show that HIV latency and reactivation mechanisms in macrophages may be distinct from those of CD4 + T cells.

Lytic/Lysogenic Transition as a Life-History Switch

Abstract The transition between lytic and lysogenic life cycles is the most important feature of the life history of temperate viruses. To explain this transition, an optimal life-history model is offered based a discrete-time formulation of phage/bacteria population dynamics that features infection of bacteria by Poisson sampling of virions from the environment. The time step is the viral latency period. In this model density-dependent viral absorption onto the bacterial surface produces virus/bacteria coexistence and density-dependence in bacterial growth is not needed. The formula for the transition between lytic and lysogenic phases is termed the “fitness switch”. According to the model, the virus switches from lytic to lysogenic when its population grows faster as prophage than as virions produced by lysis of the infected cells, and conversely for the switch from lysogenic to lytic. A prophage that benefits the bacterium it infects automatically incurs lower fitness upon exiting the bacterial genome, resulting in its becoming locked into the bacterial genome in what is termed here as a “prophage lock”. The fitness switch qualitatively predicts the ecogeographic rule that environmental enrichment leads to microbialization with a concomitant increase in lysogeny, fluctuating environmental conditions promote virus-mediated horizontal gene transfer, and prophage-containing bacteria can integrate into the microbiome of a eukaryotic host forming a functionally integrated tripartite holobiont. These predictions accord more with the “Piggyback-the-Winner” hypothesis than with the “Kill-the-Winner” hypothesis in virus ecology.

Identification of Garlic Viruses Associated with Seed Bulbs and Consumption Bulbs from Several Locations in Indonesia

Virus infection is one of the major constraints in garlic production since the viruses are readily accumulated on vegetative propagation material (bulbs). This research aimed to detect garlic common latent virus (GCLV), shallot latent virus (SLV), onion yellow dwarf virus (OYDV), and leek yellow stripe virus (LYSV) infecting local garlic as seed bulb and imported garlic as consumption bulb. Seed bulb samples were obtained from seed breeders in several garlic growing centers in Indonesia. In contrast, consumption bulb samples were obtained from plant quarantine warehouses and three local markets in Bogor. Some bulb samples were used for morphological observations, and some were germinated in the laboratory until the leaves emerged. Leaves were collected for virus detection by RT-PCR using specific primers for GCLV, SLV, OYDV, and LYSV. Seed and consumption bulbs have differences in their morphological characteristics, especially in the type of neck hardness and the size of the bulb diameter. OYDV and LYSV infections were successfully detected in seed and consumption bulbs, while SLV was only found in consumption bulbs. Nucleotide sequence analysis showed that SLV from consumption bulbs formed one group, GCLV from seed bulbs formed one group, while OYDV and LYSV from seed and consumption bulbs were in different groups, indicating that the viruses came from different strains. Further research through high-throughput detection methods and providing virus-free planting material are needed to anticipate the spread of new strains of garlic viruses in Indonesia.

High throughput RNA sequencing discovers symptomatic and latent viruses: an example from ornamental hibiscus

High throughput RNA sequencing discovers symptomatic and latent viruses: an example from ornamental hibiscus

First report of Verbena latent virus infecting giant goldenrod in Belgium

First report of <i>Verbena latent virus</i> infecting giant goldenrod in Belgium

Ecosystemic Alienation from the Perspective of Paraecology

Individual alienation, which began in the previous century with the industrialization revolution, has now progressed to the level of ecosystem alienation. Catastrophic destruction that occurs with the disruption of natural ecosystem functions proceeds insidiously. The main objective of this study is to make paraecological approaches more understandable, and aid efforts to make nature conservation and environmental ethics a way of life in the solution of environmental problems caused by ecosystemic alienation. With the magic of hedonism, an alienated person becomes lonely and robotic. Today, modernity is the main activator of alienation. Weak living things, which constitute the basic paradigm of modernity, must constantly feed this system. However, maintaining modernity tends to destroy the system by exploiting it. Ecological destruction, such as climate change, drought, and desertification have reached a global threat level. Living things are unaware that they are preparing to perish under the threat of alienation, along with their systems. Ecosystemic alienation, a latent virus that has existed for over a century, is the highest level of alienation. Selling or bartering these functions by calculating the financial value of the services and functions of natural ecosystems is another indicator of alienation. The solution is not to destroy the alienated humans (aliens) responsible for the degradation of ecosystems, but to push them to the limits and neutralize them. Efforts to create virtuous people who will solve ecological problems and adopt living as a part of nature cannot be realized with utopian principles. Adoption of global nature conservation ethics is possible with lifelong education for all ages.

Differential carbonic anhydrase activities control EBV-induced B-cell transformation and lytic cycle reactivation.

Epstein-Barr virus (EBV) contributes to ~1% of all human cancers including several B-cell neoplasms. A characteristic feature of EBV life cycle is its ability to transform metabolically quiescent B-lymphocytes into hyperproliferating B-cell blasts with the establishment of viral latency, while intermittent lytic cycle induction is necessary for the production of progeny virus. Our RNA-Seq analyses of both latently infected naïve B-lymphocytes and transformed B-lymphocytes upon lytic cycle replication indicate a contrasting expression pattern of a membrane-associated carbonic anhydrase isoform CA9, an essential component for maintaining cell acid-base homeostasis. We show that while CA9 expression is transcriptionally activated during latent infection model, lytic cycle replication restrains its expression. Pharmacological inhibition of CA-activity using specific inhibitors retards EBV induced B-cell transformation, inhibits B-cells outgrowth and colony formation ability of transformed B-lymphocytes through lowering the intracellular pH, induction of cell apoptosis and facilitating degradation of CA9 transcripts. Reanalyses of ChIP-Seq data along with utilization of EBNA2 knockout virus, ectopic expression of EBNA2 and sh-RNA mediated knockdown of CA9 expression we further demonstrate that EBNA2 mediated CA9 transcriptional activation is essential for EBV latently infected B-cell survival. In contrast, during lytic cycle reactivation CA9 expression is transcriptionally suppressed by the key EBV lytic cycle transactivator, BZLF1 through its transactivation domain. Overall, our study highlights the dynamic alterations of CA9 expression and its activity in regulating pH homeostasis act as one of the major drivers for EBV induced B-cell transformation and subsequent B-cell lymphomagenesis.

Abstract 3965: A cytolytic effector CD8+ T cell response is associated with remission in adult T-cell leukemia/lymphoma

Abstract Introduction: Adult T-cell leukemia/lymphoma (ATLL) is a rare blood cancer that develops in 3-5% of human T-lymphotropic virus-1 (HTLV1) carriers when the combination of viral oncoproteins and somatic mutations lead to malignant transformation of HTLV1 infected CD4+ T cells. ATLL has the worst overall survival among peripheral T cell lymphomas and remains a fatal disease despite efforts to improve outcomes over the last 35 years. First-line combination chemotherapy rarely achieves a durable response and two immune-modulating treatments (α-CCR4 and α-PD-1 monoclonal antibodies) failed in clinical trials. These findings highlight an urgent need to improve our understanding of the functional state of host immunity in ATLL patients. A host immune response against HTLV1 is pivotal in preventing ATLL development. Malignant ATLL cells can escape immune surveillance by preventing antigen presentation and engaging immune checkpoints. The prospect of an ATLL clone inducing systemic suppression was suggested in the literature but has not been directly examined. Furthermore, HTLV1-specific CD8+ cytotoxic T lymphocytes (CTLs) play an important role during viral latency, yet it remains unknown if they play a clinically relevant role during ATLL treatment. Methodology: In this work, we hypothesized that malignant ATLL cells will exhibit a suppressive immune phenotype and that effective therapeutic interventions will augment anti-ATLL CTLs to promote durable remission. We implemented an immune profiling approach to analyze PBMC samples from ATLL patients. Specifically, we developed a powerful 30-marker spectral flow panel to discriminate malignant ATLL cells from non-transformed lymphocytes (T, B, and NK cells) and monitor the expression of key transcription factors and markers associated with proliferation, cytolysis, and exhaustion. We analyzed diagnostic samples from the Montefiore-Einstein ATLL Biobank and samples collected during a phase 2 clinical trial aimed to assess the efficacy of a promising Belinostat/Zidovudine/Interferon-α combination therapy against ATLL (NCT02737046). Results: Our findings reveal that malignant ATLL cells exhibit an immune phenotype associated with robust immunosuppression and yet distinct from other suppressive immune cells (i.e., regulatory T cells). Additionally, we discovered that ATLL patient samples contained a unique population of phenotypically exhausted CD8+ T cells, the proportion of which was drastically reduced in patients under remission. Remarkably, a decrease in this exhausted subset correlated with the expansion of highly cytolytic CD8+ CTLs. Conclusion: Altogether, our findings suggest that ATLL cells may drive CD8+ CTLs into exhaustion to achieve immune evasion and that a robust anti-ATLL CTL response actively contributes to a remission state. Citation Format: Erik Guillen, Eric Liu, Yanhua Wang, Aditi Shastri, Alejandro Sica, Amit Verma, Xingxing Zhang, Juan Carlos Ramos, Hilda Ye, Gregoire Lauvau. A cytolytic effector CD8+ T cell response is associated with remission in adult T-cell leukemia/lymphoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3965.