Rhinovirus C replication is associated with the endoplasmic reticulum and triggers cytopathic effects in an in vitro model of human airway epithelium.

The clinical impact of rhinovirus C (RV-C) is well-documented; yet, the viral life cycle remains poorly defined. Thus, we characterized RV-C15 replication at the single-cell level and its impact on the human airway epithelium (HAE) using a physiologically-relevant in vitro model. RV-C15 replication was restricted to ciliated cells where viral RNA levels peaked at 12 hours post-infection (hpi), correlating with elevated titers in the apical compartment at 24hpi. Notably, infection was associated with a loss of polarized expression of the RV-C receptor, cadherin-related family member 3. Visualization of double-stranded RNA (dsRNA) during RV-C15 replication revealed two distinct replication complex arrangements within the cell, likely corresponding to different time points in infection. To further define RV-C15 replication sites, we analyzed the expression and colocalization of giantin, phosphatidylinositol-4-phosphate, and calnexin with dsRNA. Despite observing Golgi fragmentation by immunofluorescence during RV-C15 infection as previously reported for other RVs, a high ratio of calnexin-dsRNA colocalization implicated the endoplasmic reticulum as the primary site for RV-C15 replication in HAE. RV-C15 infection was also associated with elevated stimulator of interferon genes (STING) expression and the induction of incomplete autophagy, a mechanism used by other RVs to facilitate non-lytic release of progeny virions. Notably, genetic depletion of STING in HAE attenuated RV-C15 and -A16 (but not -B14) replication, corroborating a previously proposed proviral role for STING in some RV infections. Finally, RV-C15 infection resulted in a temporary loss in epithelial barrier integrity and the translocation of tight junction proteins while a reduction in mucociliary clearance indicated cytopathic effects on epithelial function. Together, our findings identify both shared and unique features of RV-C replication compared to related rhinoviruses and define the impact of RV-C on both epithelial cell organization and tissue functionality-aspects of infection that may contribute to pathogenesis in vivo.

Human Antiviral Protein MxA Forms Novel Metastable Membraneless Cytoplasmic Condensates Exhibiting Rapid Reversible Tonicity-Driven Phase Transitions.

Phase-separated biomolecular condensates of proteins and nucleic acids form functional membrane-less organelles (e.g., stress granules and P-bodies) in the mammalian cell cytoplasm and nucleus. In contrast to the long-standing belief that interferon (IFN)-inducible human myxovirus resistance protein A (MxA) associated with the endoplasmic reticulum (ER) and Golgi apparatus, we report that MxA formed membraneless metastable (shape-changing) condensates in the cytoplasm. In our studies, we used the same cell lines and methods as those used by previous investigators but concluded that wild-type MxA formed variably sized spherical or irregular bodies, filaments, and even a reticulum distinct from that of ER/Golgi membranes. Moreover, in Huh7 cells, MxA structures associated with a novel cytoplasmic reticular meshwork of intermediate filaments. In live-cell assays, 1,6-hexanediol treatment led to rapid disassembly of green fluorescent protein (GFP)-MxA structures; FRAP revealed a relative stiffness with a mobile fraction of 0.24 ± 0.02 within condensates, consistent with a higher-order MxA network structure. Remarkably, in intact cells, GFP-MxA condensates reversibly disassembled/reassembled within minutes of sequential decrease/increase, respectively, in tonicity of extracellular medium, even in low-salt buffers adjusted only with sucrose. Condensates formed from IFN-α-induced endogenous MxA also displayed tonicity-driven disassembly/reassembly. In vesicular stomatitis virus (VSV)-infected Huh7 cells, the nucleocapsid (N) protein, which participates in forming phase-separated viral structures, associated with spherical GFP-MxA condensates in cells showing an antiviral effect. These observations prompt comparisons with the extensive literature on interactions between viruses and stress granules/P-bodies. Overall, the new data correct a long-standing misinterpretation in the MxA literature and provide evidence for membraneless MxA biomolecular condensates in the uninfected cell cytoplasm.IMPORTANCE There is a long-standing belief that interferon (IFN)-inducible human myxovirus resistance protein A (MxA), which displays antiviral activity against several RNA and DNA viruses, associates with the endoplasmic reticulum (ER) and Golgi apparatus. We provide data to correct this misinterpretation and further report that MxA forms membraneless metastable (shape-changing) condensates in the cytoplasm consisting of variably sized spherical or irregular bodies, filaments, and even a reticulum. Remarkably, MxA condensates showed the unique property of rapid (within 1 to 3 min) reversible disassembly and reassembly in intact cells exposed sequentially to hypotonic and isotonic conditions. Moreover, GFP-MxA condensates included the VSV nucleocapsid (N) protein, a protein previously shown to form liquid-like condensates. Since intracellular edema and ionic changes are hallmarks of cytopathic effects of a viral infection, the tonicity-driven regulation of MxA condensates may reflect a mechanism for modulation of MxA function during viral infection.

High-Throughput Screening Identifies Mixed-Lineage Kinase 3 as a Key Host Regulatory Factor in Zika Virus Infection.

The Zika virus (ZIKV) life cycle involves multiple steps and requires interactions with host factors. However, the inability to systematically identify host regulatory factors for ZIKV has hampered antiviral development and our understanding of pathogenicity. Here, using a bioactive compound library with 2,659 small molecules, we applied a high-throughput and imaging-based screen to identify host factors that modulate ZIKV infection. The screen yielded hundreds of hits that markedly inhibited or potentiated ZIKV infection in SNB-19 glioblastoma cells. Among the hits, URMC-099, a mixed-lineage kinase 3 (MLK3) inhibitor, significantly facilitated ZIKV replication in both SNB-19 cells and the neonatal mouse brain. Using gene silencing and overexpression, we further confirmed that MLK3 was a host restriction factor against ZIKV. Mechanistically, MLK3 negatively regulated ZIKV replication through induction of the inflammatory cytokines interleukin-6 (IL-6), IL-8, tumor necrosis factor alpha (TNF-α), and monocyte chemoattractant protein 1 (MCP-1) but did not modulate host interferon-related pathways. Importantly, ZIKV activated the MLK3/MKK7/Jun N-terminal protein kinase (JNK) pathway in both SNB-19 cells and neonatal mouse brain. Together, these findings reveal a critical role for MLK3 in regulating ZIKV infection and facilitate the development of anti-ZIKV therapeutics by providing a number of screening hits.IMPORTANCE Zika fever, an infectious disease caused by the Zika virus (ZIKV), normally results in mild symptoms. Severe infection can cause Guillain-Barré syndrome in adults and birth defects, including microcephaly, in newborns. Although ZIKV was first identified in Uganda in 1947 in rhesus monkeys, a widespread epidemic of ZIKV infection in South and Central America in 2015 and 2016 raised major concerns. To date, there is no vaccine or specific medicine for ZIKV. The significance of our research is the systematic discovery of small molecule candidates that modulate ZIKV infection, which will allow the development of antiviral therapeutics. In addition, we identified MLK3, a key mediator of host signaling pathways that can be activated during ZIKV infection and limits virus replication by inducing multiple inflammatory cytokines. These findings broaden our understanding of ZIKV pathogenesis.

A persistently replicating SARS-CoV-2 variant derived from an asymptomatic individual.

BACKGROUND: Since the first outbreak of SARS-CoV-2, the clinical characteristics of the Coronavirus Disease 2019 (COVID-19) have been progressively changed. Data reporting a viral intra-host and inter-host evolution favouring the appearance of mild SARS-CoV-2 strains are since being accumulating. To better understand the evolution of SARS-CoV-2 pathogenicity and its adaptation to the host, it is therefore crucial to investigate the genetic and phenotypic characteristics of SARS-CoV-2 strains circulating lately in the epidemic. METHODS: Nasopharyngeal swabs have been analyzed for viral load in the early (March 2020) and late (May 2020) phases of epidemic in Brescia, Italy. Isolation of SARS-CoV-2 from 2 high viral load specimens identified on March 9 (AP66) and on May 8 (GZ69) was performed on Vero E6 cells. Amount of virus released was assessed by quantitative PCR. Genotypic characterization of AP66 and GZ69 was performed by next generation sequencing followed by an in-depth in silico analysis of nucleotide mutations. RESULTS: The SARS-CoV-2 GZ69 strain, isolated in May from an asymptomatic healthcare worker, showed an unprecedented capability of replication in Vero E6 cells in the absence of any evident cytopathic effect. Vero E6 subculturing, up to passage 4, showed that SARS-CoV-2 GZ69 infection was as productive as the one sustained by the cytopathic strain AP66. Whole genome sequencing of the persistently replicating SARS-CoV-2 GZ69 has shown that this strain differs from the early AP66 variant in 9 nucleotide positions (C2939T; C3828T; G21784T; T21846C; T24631C; G28881A; G28882A; G28883C; G29810T) which lead to 6 non-synonymous substitutions spanning on ORF1ab (P892S; S1188L), S (K74N; I95T) and N (R203K, G204R) proteins. CONCLUSIONS: Identification of the peculiar SARS-CoV-2 GZ69 strain in the late Italian epidemic highlights the need to better characterize viral variants circulating among asymptomatic or paucisymptomatic individuals. The current approach could unravel the ways for future studies aimed at analyzing the selection process which favours viral mutations in the human host.

A novel mitochondrial MAVS/Caspase-8 platform links RNA virus-induced innate antiviral signaling to Bax/Bak-independent apoptosis.

Semliki Forest virus (SFV) requires RNA replication and Bax/Bak for efficient apoptosis induction. However, cells lacking Bax/Bak continue to die in a caspase-dependent manner. In this study, we show in both mouse and human cells that this Bax/Bak-independent pathway involves dsRNA-induced innate immune signaling via mitochondrial antiviral signaling (MAVS) and caspase-8. Bax/Bak-deficient or Bcl-2- or Bcl-xL-overexpressing cells lacking MAVS or caspase-8 expression are resistant to SFV-induced apoptosis. The signaling pathway triggered by SFV does neither involve death receptors nor the classical MAVS effectors TNFR-associated factor-2, IRF-3/7, or IFN-ß but the physical interaction of MAVS with caspase-8 on mitochondria in a FADD-independent manner. Consistently, caspase-8 and -3 activation are reduced in MAVS-deficient cells. Thus, after RNA virus infection MAVS does not only elicit a type I antiviral response but also recruits caspase-8 to mitochondria to mediate caspase-3 activation and apoptosis in a Bax/Bak-independent manner.

[Herpes simplex virus type 1 latency and reactivation: an update]. / Latence et réactivation du virus de l'herpès simplex de type 1 (HSV-1) - Une mise à jour.

Following primary infections HSV-1 replicates productively in epithelial cells and enters sensory neurons via nerve termini. After retrograde transport the virus genome is delivered into the cell nucleus, where it establishes lifelong latent infections. During latency, the virus genome remains as a chromatinized episome expressing only a set of latency-associated transcripts (LAT) and a group of microRNAs that inhibit expression of key lytic viral functions. Periodically the virus can reactivate to reinitiate lytic, secondary infections at peripheral tissues. The ability to establish both lytic and latent infections relies on the coexistence in the virus genome of two alternative gene expression programs, under the control of epigenetic mechanisms. Latency is an adaptive phenotype that allows the virus to escape immune host responses and to reactivate and disseminate to other hosts upon recognizing danger signals such as stress, neurologic trauma or growth factor deprivation.

Bilateral entry and release of Middle East respiratory syndrome coronavirus induces profound apoptosis of human bronchial epithelial cells.

The newly emerged Middle East respiratory syndrome coronavirus (MERS-CoV) infects human bronchial epithelial Calu-3 cells. Unlike severe acute respiratory syndrome (SARS)-CoV, which exclusively infects and releases through the apical route, this virus can do so through either side of polarized Calu-3 cells. Infection results in profound apoptosis within 24 h irrespective of its production of titers that are lower than those of SARS-CoV. Together, our results provide new insights into the dissemination and pathogenesis of MERS-CoV and may indicate that the virus differs markedly from SARS-CoV.

Evolutionary comparison between viral lysis rate and latent period.

Marine viruses shape the structure of the microbial community. They are, thus, a key determinant of the most important biogeochemical cycles in the planet. Therefore, a correct description of the ecological and evolutionary behavior of these viruses is essential to make reliable predictions about their role in marine ecosystems. The infection cycle, for example, is indistinctly modeled in two very different ways. In one representation, the process is described including explicitly a fixed delay between infection and offspring release. In the other, the offspring are released at exponentially distributed times according to a fixed release rate. By considering obvious quantitative differences pointed out in the past, the latter description is widely used as a simplification of the former. However, it is still unclear how the dichotomy "delay versus rate description" affects long-term predictions of host-virus interaction models. Here, we study the ecological and evolutionary implications of using one or the other approaches, applied to marine microbes. To this end, we use mathematical and eco-evolutionary computational analysis. We show that the rate model exhibits improved competitive abilities from both ecological and evolutionary perspectives in steady environments. However, rate-based descriptions can fail to describe properly long-term microbe-virus interactions. Moreover, additional information about trade-offs between life-history traits is needed in order to choose the most reliable representation for oceanic bacteriophage dynamics. This result affects deeply most of the marine ecosystem models that include viruses, especially when used to answer evolutionary questions.

[Pathogenic effect of pandemic influenza virus H1N1 under replication in cultures of human cells].

The propagation of the pandemic influenza virus H1N1 in cultures of bronchial (Calu-3) and intestinal (Caco-2) differentiated epithelial cells of human origin was studied. The canine epithelial cell lines, MDCK-H and MDCK-2, were comparatively tested. The two human cell lines were found to be highly sensitive to the influenza pandemic strains A/Hamburg/05/09 and A/Moscow/501/2011 and maintained their replication without addition of trypsin to culture medium. Virus strains of seasonal influenza H1N1, such as A/Moscow/450/2003, A/Memphis/14/96, and laboratory strain A/PR/8/34, multiplied in these human cells in similar manner. The intracellular cleavage HA0-->HA1+HA2 by the host virus-activating protease (IAP) occurred in both human cell lines under infection with each influenza virus H1N1 including pandemic ones. Comparatively, this cleavage of all influenza H1N1 virus strains appeared to be either undetectable or low-detectible in MDCK-H and MDCK-2, respectively, thereby implying low levels of active IAP in these cells. Multiplication of pandemic and seasonal influenza H1N1 viruses in Calu-3 and Caco-2 cells caused cytopathic effect, which was accompanied with low autophagy and apoptosis events. These data allow recommending human cell lines, Calu-3 and Caco-2, for optimized isolation and passaging of clinical strains of Influenza pandemic viruses H1N1.

[Biochemical markers of virus cytopathogenicity in macrophages].

The results of macrophage metabolism studies at their infection by viruses differing in the level of virulence are presented. With the purpose of optimizing the estimation of viral cytopathogenic effects on macrophages, an index of cell reactions, which allows one to reveal the degree of virus influence in standard units, is offered. Generally, the application of high-sensitivity methods for functional activity determination and identification of the correlative communication between its changes and morphological features of cells can be prescribed to objective identification methods of not only viral reproduction, but also differentiation of types and the degree of their cytopathogenic effects.

Interplay of acute and persistent infections caused by Venezuelan equine encephalitis virus encoding mutated capsid protein.

Venezuelan equine encephalitis virus (VEEV) is a significant human and animal pathogen. The highlight of VEEV replication in vitro, in cells of vertebrate origin, is the rapid development of cytopathic effect (CPE), which is strongly dependent upon the expression of viral capsid protein. Besides being an integral part of virions, the latter protein is capable of (i) binding both the nuclear import and nuclear export receptors, (ii) accumulating in the nuclear pore complexes, (iii) inhibiting nucleocytoplasmic trafficking, and (iv) inhibiting transcription of cellular ribosomal and messenger RNAs. Using our knowledge of the mechanism of VEEV capsid protein function in these processes, we designed VEEV variants containing combinations of mutations in the capsid-coding sequences. These mutations made VEEV dramatically less cytopathic but had no effect on infectious virus production. In cell lines that have defects in type I interferon (IFN) signaling, the capsid mutants demonstrated very efficient persistent replication. In other cells, which have no defects in IFN production or signaling, the same mutants were capable of inducing a long-term antiviral state, downregulating virus replication to an almost undetectable level. However, ultimately, these cells also developed a persistent infection, characterized by continuous virus replication and beta IFN (IFN-beta) release. The results of this study demonstrate that the long-term cellular antiviral state is determined by the synergistic effects of type I IFN signaling and the antiviral reaction induced by replicating viral RNA and/or the expression of VEEV-specific proteins. The designed mutants represent an important model for studying the mechanisms of cell interference with VEEV replication and development of persistent infection.

Direct cytopathic effects of particular hepatitis B virus genotypes in severe combined immunodeficiency transgenic with urokinase-type plasminogen activator mouse with human hepatocytes.

BACKGROUND & AIMS: Little is known about the direct cytopathic effect of hepatitis B virus (HBV) and its association with particular viral genotypes or genetic mutations. We investigate HBV genotype-related differences in viral replication, antigen expression, and histopathology in severe combined immunodeficiency transgenic with urokinase-type plasminogen activator mice harboring human hepatocytes. METHODS: Mice were inoculated with wild-type of different genotype strains (3 for each HBV/A2, B1, and C2) recovered from preinfected-mice sera or patient sera. RESULTS: Histologic analysis of mice infected with HBV/C2 for 22-25 weeks showed abundant ground-glass appearance of the hepatocytes and fibrosis in the humanized part of the murine liver owing to the activation of hepatic stellate cells mediated by oxidative stress through transforming growth factor-beta1 signaling, whereas neither was observed with HBV/A2 and B1. The HBV-DNA level in sera was the highest in mice infected with HBV/C2 compared with those with HBV/A2 and HBV/B1 (10(9), 10(7), and 10(4) log copies/mL, respectively, P < .05) during 6-8 weeks postinoculation. HB core-related antigen excretion had a similar trend among the genotypes, whereas secretion of HB surface antigen was more pronounced for HBV/A2 followed by HBV/C2 and much less for HBV/B1. Introduction of precore stop-codon mutation in the HBV/B1 caused a significant increase in viral replication, antigen expression, and a histopathologic picture similar to HBV/C2. CONCLUSIONS: By using a humanized in vivo model, we show that different HBV genotypes and even particular mutations resulted in different virologic and histopathologic outcomes of infection, indicating that particular genetic variants of HBV may be directly cytopathic in immunosuppressive conditions.

To kill or be killed: how viruses interact with the cell death machinery.

A virus (from the Latin virus meaning toxin or poison) is a small infectious agent that can only replicate inside the cells of another organism. Viruses are found wherever there is life and have probably existed since living cells first evolved. Viruses do not have their own metabolism and require a host cell to make new products. The range of structural and biochemical (i.e., cytopathic) effects that viruses have on the host cell is extensive. Most viral infections eventually result in the death of the host cell. The causes of death include cell lysis, alterations to the cell's surface membrane and various modes of programmed cell death. Some viruses cause no apparent changes to the infected cell. Cells in which the virus is latent and inactive show few signs of infection and often function normally. This causes persistent infection and the virus is often dormant for many months or years. Some viruses can cause cells to proliferate without causing malignancy, whereas others are established causes of cancer. Human organisms use a genetically controlled cell death programme that prevents the spreading of viral infection and kills the virus. Between 19 and 21 November 2009, with sponsorship from the Journal of Internal Medicine, the Swedish Research Foundation and the Swedish Cancer Society hosted a conference in Stockholm entitled: 'To kill or to be killed. Viral evasion strategies and interference with cell death machinery'. Four comprehensive reviews from this conference are presented in this issue of the Journal of Internal Medicine. These reviews include descriptions of: the modulation of host innate and adaptive immune defenses by cytomegalovirus; the impact of gamma-chain family cytokines on T cell homoeostasis in HIV-1 infection and the therapeutic implications; approaches to killing tumours by depriving them of the mechanisms for detoxification; and viral strategies for the evasion of immunogenic cell death.

In vitro and in vivo effects of West Nile virus propagated in cells of different passage.

Tissue cultures are used to propagate viruses for use in mosquito infection studies and to detect live virus in field-collected specimens. Microscopic evaluation of cytopathic effects is used to visualize virus presence. In this study, both low-passage (LP; n = 35) and high-passage (HP; n = 218) lines of African green monkey kidney (Vero) cells were infected with West Nile virus, and virus growth kinetics were quantitated over different incubation periods (IPs) (2, 6, 10, 24, 48, 72, and 96 h postinfection). Virus titers were significantly (P < 0.05) higher in the HP compared with LP line 24, 48, and 96 h postinfection. No differences were observed in plaque morphology between the LP and HP lines. Culex pipiens quinquefasciatus were fed infectious blood meals created using supernatant from the 2 lines and maintained at 28 degrees C for a 12-day IP. Although the virus dose was higher (P < 0.05) in the HP compared with the LP line, there were no significant differences in mosquito infection or dissemination rates at the end of the IP. The significance of these observations for assessing virus presence and pathogenicity is discussed.

[Phage display selection on MRC-5 cells yield peptides specific for HCMV-binding].

OBJECTIVE: To screen the special binding peptides to the MRC-5 cells infected by human cytomegalovirus (HCMV) so as to study the progress how HCMV recognizes susceptive host cells. METHODS: The special binding peptides to MRC-5 cells infected by HCMV rather than normal MRC-5 cells were screened by phage display system, and then identified by ELISA, the sequence of single strand DNA were determined and analyzed. In addition, the amino acid sequence of target protein was compared in protein sequence database from BLAST in GenBank, and then the physico-chemical property was also analyzed by Vector NTI software. RESULTS: The high affinity phage-ligands to MRC-5 infected by HCMV were found, and then sequences of peptides are EVNMSDS, NVSVFET and GQQPTTV respectively. These three peptides sequences were found in HCMV gB and gH protein. CONCLUSION: The peptides that can special bind HCMV-infected MRC-5 cells has been screened by phage display system. This new finding should bring new ideas for studying the functional domain through which HCMV can recognize and bind host cells.

Autophagy is induced and supports virus replication in Enterovirus A71-infected human primary neuronal cells.

Enterovirus A71 (EV-A71), which belongs to the family Picornaviridae, can invade the central nervous system (CNS) and cause severe CNS complications or death. The EV-A71 antigen has been detected in the neurons in the brains of humans who died from EV-A71 infection. However, the effect of EV-A71 infection on human neuronal cells remains poorly understood. Human neural stem cells (NSCs) and IMR-32 neuroblastoma cells were differentiated into neuronal cells for this study. Although the neuronal cells were permissive to EV-A71 infection, EV-A71 infection did not induce an obvious cytopathic effect on the neuronal cells. EV-A71 infection did not induce apoptosis in neuronal cells. However, autophagy and autophagic flux were induced in EV-A71-infected neuronal cells. The production of autophagosomes was shown to be important for EV-A71 viral RNA (vRNA) replication in neuronal cells.

Genetic characterization of a novel recombinant PRRSV2 from lineage 8, 1 and 3 in China with significant variation in replication efficiency and cytopathic effects.

There are four major porcine reproductive and respiratory syndrome virus 2 (PRRSV2) lineages circulating in China based on classification system, including lineages 1 (NADC30-like), 3 (QYYZ-like), 5.1 (VR2332-like) and 8 (JXA1-like/CH-1a-Like), which leads to the potential recombination. In the present study, a novel variant of PRRSV2 strain named JS18-3 was isolated from piglets suffering severe breathing difficulties in Jiangsu Province of China in 2018. Full-length genome analysis indicated that JS18-3 shared 86.5%, 87.9%, 84.2%, 82.2% and 86.4% nucleotide similarity with PRRSVs CH-1a, JXA1, VR2332, QYYZ and NADC30, respectively. 4871-6635 of JS18-3 shared the highest identity of 99.3% in nucleotide sequence with HP-PRRSV representative strain JXA1 indicating ongoing evolution to HP-PRRSV. JS18-3 was classified into classical lineage 8 of PRRSV2 based on phylogenetic analysis of complete genome and ORF5. Genomic break points in structural (ORF3) and non-structural (NSP2, NSP3) regions of genomes were detected in recombination analysis. JS18-3 is a recombinant isolate from lineages 8, 1 and 3. Replication enhancement and severe cytopathic effects caused by JS18-3 were observed in Marc-145 cells and porcine alveolar macrophages (PAMs) as compared to JX07, a typical strain of lineage 8. Pathogenicity results indicated that piglets inoculated with JS18-3 presented persistent fever, dyspnoea, serious microscopic lung lesions and lymph node congestion. The study suggests that lineage 8 of PRRSV2 is involved in continuing evolution by genetic recombination and mutation leading to outbreaks in vaccinated pigs in China.

Potent antitumor activity of double-regulated oncolytic adenovirus-mediated ST13 for colorectal cancer.

Following targeted gene virotherapy, the suppression of tumorigenicity 13 (ST13) gene was inserted into the double-regulated oncolytic adenovirus SG500 to ensure more safety and potent antitumor activity against colorectal cancer in vitro and in vivo. We generated the ST13-expressing oncolytic adenovirus SG500-ST13, with which colorectal carcinoma cell lines SW620 and HCT116, and the lung fibroblast cell line WI38, were infected. Crystal violet staining was carried out to detect the cytopathic effect in cells, and the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method was used to assay cell viability. The effect of apoptosis induced by SG500-ST13 was confirmed by Hoechst staining and the TdT-mediated dUTP-biotin nick-end labeling method. To further identify the antitumor effects of SG500-ST13 on HCT116 xenografts in Balb/c nude mice, the induction of cell death was assessed by hematoxylin-eosin staining. Immunohistochemical study was also carried out.