CXCR4 WHIM syndrome is a cancer predisposition condition for virus-induced malignancies.

Warts, hypogammaglobulinaemia, infections and myelokathexis syndrome (WHIMS) is a rare combined primary immunodeficiency caused by the gain of function of the CXCR4 chemokine receptor. We present the prevalence of cancer in WHIMS patients based on data from the French Severe Chronic Neutropenia Registry and an exhaustive literature review. The median follow-up of the 14 WHIMS 'patients was 28.5 years. A central review and viral evaluation of pathological samples were organized, and we conducted a thorough literature review to identify all reports of WHIMS cases. Six French patients were diagnosed with cancer at a median age of 37.6 years. The 40-year risk of malignancy was 39% (95% confidence interval [CI]: 6%-74%). We observed two human papillomavirus (HPV)-induced vulvar carcinomas, three lymphomas (two Epstein-Barr virus [EBV]-related) and one basal cell carcinoma. Among the 155 WHIMS cases from the literature, 22 cancers were reported in 16 patients, with an overall cancer 40-year risk of 23% (95% CI: 13%-39%). Malignancies included EBV-associated lymphoproliferative disorders and HPV-positive genital and anal cancers as in the French cohort. Worldwide, nine cases of malignancy were associated with HPV and four with EBV. Immunocompromised WHIMS patients appear to be particularly susceptible to developing early malignancy, mainly HPV-induced carcinomas, followed by EBV-related lymphomas.

[Monkeypox to Mpox or the re-emergence of an old zoonosis]. / De la variole du singe à la Mpox ou la réémergence d'une ancienne zoonose.

Monkeypox is a zoonotic disease caused by the Monkeypox virus (MPXV) of the Orthopoxvirus genus. The first human cases occurred in Africa in the 1970s and remained confined to the African continent for a long time until 2003, when several dozen cases occurred in the United States, following contamination by prairie dogs. Unprecedented transmission events have led to more than 80,000 reported cases worldwide between May 2022 and February 2023, primarily affecting men who have sex with men. The changing epidemiology of Mpox has raised concerns about its ability to become endemic beyond its traditional geographic areas. Confirmatory diagnosis is based on direct detection by molecular biology. Pre- or post-exposure smallpox vaccination was widely deployed in early summer 2022 to limit the spread of the disease. In case of severe forms, the use of antivirals can be considered, only tecovirimat being recommended in this indication. The current epidemic has had the merit of showing that a disease that was previously confined to regions of initial virus circulation can spread very rapidly in Western countries and of the need to reinforce the implementation of tools for the surveillance and control of communicable diseases.

[The cutaneous virome: from virology to personalized medicine]. / Le virome cutané : de la virologie à la médecine personnalisée.

The virome of the skin, defined as all viruses detected in the skin, represents a significant part of the microbiota. A much more recent discovery than the bacterial flora, the existence of the cutaneous virome has been revealed by recent metagenomic studies. The normal human skin virome is dominated by bacteriophages, Papillomaviridae, whose genomic diversity has proved extraordinary, and Polyomaviridae. Many yet unknown viral genomes within this virome await identification. The composition of the virome of the skin has been shown to be strictly individual and relatively stable over time, resulting from adaptation to everyone's genetics, lifestyle and mechanisms of immunological tolerance finely selected over the course of evolution. Yet little studied, the virome of the skin and all its interactions with other microbiota and the host are attracting growing interest. Indeed, constitutional or acquired alterations in the homeostasis between the commensal virome and the skin, ranging from sub-clinical viral dysbiosis to severe transformation of keratinocytes or adnexal cells, have been observed. These recent observations are stimulating the search for innovative solutions aimed at measuring or even modulating its pathological expression, with a view to personalized medicine.

ELANE neutropenia and solid tumors: Four cases from the French severe chronic neutropenia registry.

Neutropenia related to ELANE gene mutations predisposes patients to infection and leukemia/myelodysplasia, but little is known about the predisposition to cancer. Among a cohort of 147 patients, we identified four with malignant solid tumors (papillary thyroid cancer, anal squamous cell cancer, papillary renal cell carcinoma, and adrenocortical carcinoma), all aged 25-50 years. Three occurred with cyclic neutropenia, and one occurred with severe chronic neutropenia. Previous radiotherapy was identified as a risk factor in one patient. No genetic predisposition was identified in the three other patients.

Initial TK-deficient HSV-1 infection in the lip alters contralateral lip challenge immune dynamics.

Primary infection with herpes simplex type 1 (HSV-1) occurring around the mouth and nose switches rapidly to lifelong latent infection in sensitive trigeminal ganglia (TG) neurons. Sporadic reactivation of these latent reservoirs later in life is the cause of acute infections of the corneal epithelium, which can cause potentially blinding herpes simplex keratitis (HSK). There is no effective vaccine to protect against HSK, and antiviral drugs provide only partial protection against recurrences. We previously engendered an acute disease-free, non-reactivating latent state in mice when challenged with virulent HSV-1 in orofacial mucosa, by priming with non-neurovirulent HSV-1 (TKdel) before the challenge. Herein, we define the local immune infiltration and inflammatory chemokine production changes after virulent HSV-1 challenge, which were elicited by TKdel prime. Heightened immunosurveillance before virulent challenge, and early enhanced lymphocyte-enriched infiltration of the challenged lip were induced, which corresponded to attenuation of inflammation in the TG and enhanced viral control. Furthermore, classical latent-phase T cell persistence around latent HSV-1 reservoirs were severely reduced. These findings identify the immune processes that are likely to be responsible for establishing non-reactivating latent HSV-1 reservoirs. Stopping reactivation is essential for development of efficient vaccine strategies against HSV-1.

The Chemokine System in Oncogenic Pathways Driven by Viruses: Perspectives for Cancer Immunotherapy.

Chemokines interact with glycosaminoglycans of the extracellular matrix and activate heptahelical cellular receptors that mainly consist of G Protein-Coupled Receptors and a few atypical receptors also with decoy activity. They are well-described targets of oncogenic pathways and key players in cancer development, invasiveness, and metastasis acting both at the level of cancer cells and cells of the tumor microenvironment. Hence, they can regulate cancer cell proliferation and survival and promote immune or endothelial cell migration into the tumor microenvironment. Additionally, oncogenic viruses display the potential of jeopardizing the chemokine system by encoding mimics of chemokines and receptors as well as several products such as oncogenic proteins or microRNAs that deregulate their human host transcriptome. Conversely, the chemokine system participates in the host responses that control the virus life cycle, knowing that most oncoviruses establish asymptomatic latent infections. Therefore, the deregulated expression and function of chemokines and receptors as a consequence of acquired or inherited mutations could bias oncovirus infection toward pro-oncogenic pathways. We here review these different processes and discuss the anticancer therapeutic potential of targeting chemokine availability or receptor activation, from signaling to decoy-associated functions, in combination with immunotherapies.

Acyclovir-Resistant Herpes Simplex Virus 1 Keratitis: A Concerning and Emerging Clinical Challenge.

PURPOSE: To describe the clinical and virological profiles of patients with herpes simplex keratitis (HSK) caused by acyclovir-resistant (ACVR) strains of herpes simplex virus 1 (HSV-1). DESIGN: Multicenter retrospective case series. METHODS: HSV-1 resistance to ACV was confirmed using sequencing of genes encoding HSV-1 thymidine kinase (TK) and DNA polymerase (DNA pol). Data were collected on the number of HSK episodes before and after the diagnosis of resistance, ocular findings including the type of HSK, immune status of patients, antiviral treatments, and HSV-1 genotypic resistance profiles. RESULTS: This study evaluated 18 HSK patients (13 male and 5 female, aged 66.8 ± 4.7 years) with ACVR HSV-1-positive ocular samples. Genotypic resistance testing was performed because of frequent recurrences despite adequate antiviral prophylaxis (AVP) (n = 13, 72%), or poor response to suppressive antiviral therapy (n = 5, 28%). Resistance mutations were found in the TK (n = 15, 83%) or in the DNA pol gene (n = 3, 17%). Prior to the diagnosis of resistance, the duration of disease was 29.8 ± 20.4 years, with more than 10 HSK recurrences in 15 patients (83%). The number of recurrences between the first episode and the diagnosis of resistance was significantly lower in immunocompromised patients (n = 6, 33%) than in immunocompetent patients (n = 12; 67%) (11.5 ± 4.9 vs 16.4 ± 1.9, P = .05). CONCLUSION: HSV-1 resistance to ACV must be suspected in HSK patients with recurrences despite AVP and/or in cases that respond poorly to a suppressive antiviral regimen. Immunocompromised patients and/or those with longstanding disease may be particularly at risk for developing resistance.

Immune checkpoint inhibitors for progressive multifocal leukoencephalopathy: a new gold standard?

OBJECTIVES: Progressive multifocal leukoencephalopathy (PML) is a very rare and opportunistic encephalitis caused by JC polyomavirus that is linked to profound immunosuppression and is usually fatal unless immune function can be restored. Immune checkpoint inhibitors (ICI) are monoclonal antibodies (mAbs) that block either CTLA-4 or PD-1 inhibitor receptors, thus enhancing antiviral T-cell activity. Successful treatment of PML by ICI has recently generated some enthusiasm in case reports/small series of patients. However, the initial enthusiasm was mitigated by some individual case reports that did not show any benefit. More data are thus warranted about efficacy of immune checkpoint inhibitors in the specific context of PML. METHODS AND RESULTS: We report here the outcomes of six PML patients treated by ICI between 2017 and 2019. Underlying causes of immunosuppression consisted in hematologic malignancies (n = 4), primary immune deficiency (n = 1) and use of immunosuppressive therapies for myasthenia gravis (n = 1). Three patients were alive with a mean follow-up of 21 months (14-33) after first ICI infusion, including one patient with frank clinical response, one with stabilization, and one with initial worsening and further stabilization of PML. The three other patients rapidly died from PML. CONCLUSIONS: Our data suggest that ICI may be effective for PML treatment but were less impressive than the ones previously reported. Larger studies are thus warranted to confirm this efficacy and to identify the predictive factors of response.

Clinical Features and Diagnosis of Anterior Segment Inflammation Related to Cytomegalovirus in Immunocompetent African, Asian, and Caucasian Patients.

Objective: To report the clinical features and treatment outcomes in immunocompetent patients with anterior segment inflammation (ASI) related to human cytomegalovirus (HCMV) depending on their ethnic origin.Material and Methods: Multicenter retrospective study of 38 patients with at least one test, either HCMV-positive PCR or GWc.Results: Features of Posner-Schlossman syndrome were observed in 50% of the eyes, Fuchs heterochromic iridocyclitis in 13% of the eyes, chronic nonspecific anterior uveitis in 21% of the eyes, and corneal endotheliitis in 18% of the eyes. PCR and GWc were positive for HCMV in 50% and 96.2% of the eyes, respectively. Glaucoma was diagnosed in 50% of eyes. Treatment was oral valganciclovir in about half of the patients. Other treatments were intravenous ganciclovir and/or ganciclovir topical ointment and/or intravitreal ganciclovir.Conclusions: No obvious association of specific clinical features with individual ethnicity could be identified. We found a high rate of glaucoma in all ethnic groups. There was a delay in diagnosis and specific treatment of HCMV in most patients.

Investigation of Inherited Chromosomally Integrated Human Herpesvirus-6A+ and -6B+ in a Patient with Ulipristal Acetate-Induced Fulminant Hepatic Failure.

Inherited chromosomally integrated (ici) human herpes virus 6 (HHV-6) is estimated to occur in 0.6-2.7% of people worldwide. HHV-6 comprises two distinct species: HHV-6A and HHV-6B. Both HHV-6A and HHV-6B integration have been reported. Several drugs are capable of activating iciHHV-6 in tissues, the consequences of which are poorly understood. We report herein a case of a woman with iciHHV-6A+ and iciHHV-6B+, who developed ulipristal acetate (a selective progesterone receptor modulator)-induced fulminant hepatic failure that required liver transplantation. We confirmed the presence of ~one copy per cell of both HHV-6A and HHV-6B DNA in her hair follicles using multiplex HHV-6A/B real-time PCR and demonstrated the Mendelian inheritance of both iciHHV-6A and iciHHV-6B in her family members over three generations. Because of the rarity of this presentation, we discuss herein the possible links between reactivated HHV-6 from iciHHV-6A and/or iciHHV-6B and adverse drug reactions, suggesting that iciHHV-6 could be screened before the introduction of any hepatotoxic drugs to exclude HHV-6 active disease or combined idiosyncratic drug-induced liver injury in these patients.

Herpes Simplex Virus 1 Replication, Ocular Disease, and Reactivations from Latency Are Restricted Unilaterally after Inoculation of Virus into the Lip.

Ocular herpes simplex keratitis (HSK) is a consequence of viral reactivations from trigeminal ganglia (TG) and occurs almost exclusively in the same eye in humans. In our murine oro-ocular (OO) model, herpes simplex virus 1 (HSV-1) inoculation in one side of the lip propagates virus to infect the ipsilateral TG. Replication here allows infection of the brainstem and infection of the contralateral TG. Interestingly, HSK was observed in our OO model only from the eye ipsilateral to the site of lip infection. Thus, unilateral restriction of HSV-1 may be due to differential kinetics of virus arrival in the ipsilateral versus contralateral TG. We inoculated mice with HSV-1 reporter viruses and then superinfected them to monitor changes in acute- and latent-phase gene expression in TG after superinfection compared to the control (single inoculation). Delaying superinfection by 4 days after initial right lip inoculation elicited failed superinfecting-virus gene expression and eliminated clinical signs of disease. Initial inoculation with thymidine kinase-deficient HSV-1 (TKdel) completely abolished reactivation of wild-type (WT) superinfecting virus from TG during the latent stage. In light of these seemingly failed infections, viral genome was detected in both TG. Our data demonstrate that inoculation of HSV-1 in the lip propagates virus to both TG, but with delay in reaching the TG contralateral to the side of lip infection. This delay is responsible for restricting viral replication to the ipsilateral TG, which abrogates ocular disease and viral reactivations from the contralateral side. These observations may help to understand why HSK is observed unilaterally in humans, and they provide insight into vaccine strategies to protect against HSK.IMPORTANCE Herpetic keratitis (HK) is the leading cause of blindness by an infectious agent in the developed world. This disease can occur after reactivation of herpes simplex virus 1 in the trigeminal ganglia, leading to dissemination of virus to, and infection of, the cornea. A clinical paradox is evidenced by the bilateral presence of latent viral genomes in both trigeminal ganglia, while for any given patient the disease is unilateral with recurrences in a single eye. Our study links the kinetics of early infection to unilateral disease phenomenon and demonstrates protection against viral reactivation when kinetics are exploited. Our results have direct implications in the understanding of human disease pathogenesis and immunotherapeutic strategies for the treatment of HK and viral reactivations.

Identification by high-throughput sequencing of HPV variants and quasispecies that are untypeable by linear reverse blotting assay in cervical specimens.

The linear reverse blotting assays are valid methods for accurate human papillomavirus (HPV) typing required to manage women at risk of developing cervical cancer. However, some samples showed a positive signal in HPV lines but failed to display a positive signal in subsequent typing lines (designated as HPV-X), which indicate that certain types were not available on the respective typing blots. The aim of this study is to elucidate the types or variants of HPV through the high-throughput sequencing (HTS) of 54 ASCUS cervical samples in which the viruses remained untypeable with INNO LiPA HPV® assays. Low-risk (LR)-HPV types (HPV6, 30, 42, 62, 67, 72, 74, 81, 83, 84, 87, 89, 90 and 114), high-risk (HR)-HPV35 and possibly (p)HR-HPV73 were detected among HPV-X. Individual multiple infections (two to seven types) were detected in 40.7% of samples. Twenty-two specimens contained variants characterised by 2-10 changes. HPV30 reached the maximal number of 17 variants with relative abundance inferior or equal to 2.7%. The presence of L1 quasispecies explains why linear reverse blotting assays fail when variants compete or do not match the specific probes. Further studies are needed to measure the LR-HPV quasispecies dynamics and its role during persistent infection.

Antiviral effects of Cacicol®, a heparan sulfate biomimetic for corneal regeneration therapy, for herpes simplex virus type-1 and varicella zoster virus infection.

BACKGROUND: Cacicol®, a topical eye biopolymer containing a poly-carboxymethylglucose sulfate solution that is a regenerating matrix therapy agent, intended for wound healing of persistent corneal epithelial defects. Based on the chemical composition, we hypothesized that Cacicol® may compete with natural heparan sulfate (HS) which initiates cell surface attachment of herpes simplex virus type-1 (HSV-1), varicella zoster virus (VZV) and human adenovirus (HAdV), three viruses associated with corneal infections. METHODS: Cacicol® was compared to vehicle in the following viral strains: HSV-1 SC16 strain and HSV-1 PSLR, a clinical isolate highly resistant to acyclovir and foscarnet; VZV ATH and VZV FLO, two VZV clinical isolates; and HAdV-D37 strain. Viruses in Cacicol® or vehicle were added to cells for 1 h during adsorption then viral replication was assessed by plaque reduction assays on Vero cells for HSV-1 and MeWo cells for VZV and by immunostaining assay on Hep-2 cells for HAdV-D37. RESULTS: The vehicle had no effect, dose-dependent effects were demonstrated when HSV-1 SC16, HSV-1 PSLR, VZV ATH and VZV FLO were inoculated in the presence of Cacicol®, inhibiting viral replication by 98.4%, 98.9%, 90.1% and 89.0%, respectively. Cacicol® had no antiviral effect against HAdV-D37. CONCLUSIONS: Cacicol® has a significant antiviral activity on HSV-1 and VZV, but not on HAdV-D37. The lack of effect on HAdV is probably because it is less dependent on HS interactions for cell entry. Clinical studies are necessary to determine Cacicol® for an adjunct or alternative therapy of corneal HSV-1 or VZV infection, particularly for the management of antiviral resistant HSV-1.

Recurrent herpetic keratitis despite antiviral prophylaxis: A virological and pharmacological study.

Recurrent herpes simplex keratitis (HSK) is a leading infectious cause of blindness in industrialized countries. Antiviral prophylaxis (AVP) may fail to prevent recurrence of HSK due to viral resistance, inadequate dosing, or poor patient compliance. In this prospective multicenter study, we enrolled immunocompetent patients with recurrent HSK despite AVP. Ocular samples were tested by PCR for herpes simplex virus 1 (HSV-1). HSV-1 drug resistance was assessed with a genotypic assay based on UL23 and UL30 gene sequencing. After curative full dose valacyclovir (VACV) treatment was started, peak and trough acyclovir (ACV) plasma concentrations were measured, and patient compliance to AVP was assessed with a questionnaire. The study sample was comprised of 43 patients. Six (14%) patients were positive for HSV-1 using PCR, of whom 5 (83%) harbored genotypically ACV-resistant (ACVR) virus, due to mutations in UL23 (n = 4) or UL30 (n = 1). Disease duration was statistically significantly longer in patients with viral resistance compared to other HSK patients [35.5 ± 23.4 years (range, 6.8-68.4 years) versus 11.1 ± 12.3 years (range, 0.8-56.3 year) respectively; Mann-Whitney p = 0.01)]. While patients were treated with full dose VACV, trough ACV plasma concentrations were below the threshold for ACV sensitivity in 9.5% of cases, and compliance was poor in 5.3% of cases. To summarize, HSV-1 resistance to ACV seems to be a significant cause of failure of prophylaxis in patients with HSK and is associated with longer disease duration. Most PCR-positive samples contained genotypically ACVR virus and identification may aid in adapting treatment. Incomplete 24-h drug coverage may also explain some cases of failure of prophylaxis.

Mutation in the L3 Ribosomal Protein Could Be Associated with Risk of Selection of High-Level Linezolid-Resistant Staphylococcus epidermidis Strains.

Linezolid (LZD) has arisen as an alternative treatment in diabetic foot osteitis due to staphylococci. LZD resistance selection is difficult and involved various molecular mechanisms. As a better knowledge of those mechanisms could be beneficial for pathogenic strains' screening, we simulated in vitro the spontaneous mutagenesis process that leads to LZD-resistant strains from two Staphylococcus epidermidis strains responsible for monomicrobial diabetic foot osteitis. LZD high resistance was selected for both strains, with the same timeline of mutation appearance. Mutation in L3 protein (G152D) occurred first and quickly, but did not cause phenotypically detectable resistance or fitness cost. It was later followed by different 23S rRNA mutations (G2505A, G2447T), leading this time to detectable resistance (minimum inhibitory concentration [MIC] ≥8 mg/L). This phenomenon underlies the difficulty of resistance selection in coagulase-negative staphylococci (CoNS). This study is the first description of G2505A mutation in CoNS. Various phenotypical impacts were observed depending on strain and mutation: (i) fitness cost of G2505A and G2447T mutations; (ii) loss of erythromycin resistance concomitantly with L3 mutation selection; (iii) correlation between number of mutated rrl copies and LZD resistance level for G2447T. In conclusion, the risk of selection of high-level LZD-resistant S. epidermidis strains is weak, but does exist. It could probably appear in case of long-term treatment and be favored in the case of a pre-existing mutation in L3 ribosomal protein. Thus, broad screening conditions for pathogenic strains should probably be considered.

The CXCL12/CXCR4 Signaling Pathway: A New Susceptibility Factor in Human Papillomavirus Pathogenesis.

The productive human papillomavirus (HPV) life cycle is tightly linked to the differentiation and cycling of keratinocytes. Deregulation of these processes and stimulation of cell proliferation by the action of viral oncoproteins and host cell factors underlies HPV-mediated carcinogenesis. Severe HPV infections characterize the wart, hypogammaglobulinemia, infection, and myelokathexis (WHIM) immunodeficiency syndrome, which is caused by gain-of-function mutations in the CXCR4 receptor for the CXCL12 chemokine, one of which is CXCR41013. We investigated whether CXCR41013 interferes in the HPV18 life cycle in epithelial organotypic cultures. Expression of CXCR41013 promoted stabilization of HPV oncoproteins, thus disturbing cell cycle progression and proliferation at the expense of the ordered expression of the viral genes required for virus production. Conversely, blocking CXCR41013 function restored virus production and limited HPV-induced carcinogenesis. Thus, CXCR4 and its potential activation by genetic alterations in the course of the carcinogenic process can be considered as an important host factor for HPV carcinogenesis.

Epidemiological and clinical characteristics of patients infected with enterovirus D68, France, July to December 2014.

In 2014, the United States (US) experienced a nationwide outbreak of enterovirus D68 (EV-D68) infection with 1,152 cases reported mainly in hospitalised children with severe asthma or bronchiolitis. Following the US alert, 11 laboratories of the French enterovirus (EV) surveillance network participated in an EV-D68 survey. A total of 6,229 respiratory samples, collected from 1 July to 31 December 2014, were screened for EV-D68 resulting in 212 EV-D68-positive samples. These 212 samples corresponded to 200 EV-D68 cases. The overall EV-D68 positivity rates among respiratory samples were of 5% (184/3,645) and 1.1% (28/2,584) in hospitalised children and adults respectively. The maximum weekly EV-D68 positivity rates were of 16.1% for children (n = 24/149; week 43) and 2.6% for adults (n = 3/115; week 42). Of 173 children with EV-D68 infection alone, the main symptoms were asthma (n = 83; 48.0%) and bronchiolitis (n = 37; 21.4%). One child developed acute flaccid paralysis (AFP) following EV-D68-associated pneumonia. Although there was no significant increase in severe respiratory tract infections reported to the French public health authorities, 10.7% (19/177) of the EV-D68 infected children and 14.3% (3/21) of the EV-D68 infected adults were hospitalised in intensive care units. Phylogenetic analysis of the viral protein 1 (VP1) sequences of 179 EV-D68 cases, revealed that 117 sequences (65.4%), including that of the case of AFP, belonged to the B2 variant of clade B viruses. Continuous surveillance of EV-D68 infections is warranted and could benefit from existing influenza-like illness and EV surveillance networks.

Complementary assays for monitoring susceptibility of varicella-zoster virus resistance to antivirals.

The emergence of varicella-zoster virus (VZV) resistance to current antivirals as acyclovir (ACV) constitutes a hindrance to antiviral treatment effectiveness of VZV infections, especially in immunocompromised patients. The molecular mechanisms of VZV resistance reported so far rely on the presence of mutations within thymidine kinase (TK, ORF36) and DNA polymerase (ORF28) viral genes. The aim of this work was to develop reliable and complementary diagnostic methods to detect VZV antiviral resistance: (i) a genotypic assay based on TK and DNA polymerase genes sequencing, (ii) a plaque reduction assay to determine antiviral 50% effective concentrations, and (iii) a functional assay to evaluate in vitro phosphorylation activity of recombinant TKs. As a whole, this study included the analysis of 21 VZV clinical isolates and 62 biological samples from patients experiencing VZV infection. Genetic analysis revealed 3 and 9 new amino acid changes that have not been previously described within the highly conserved TK and DNA polymerase, respectively. Then, VZV isolates bearing newly identified mutations considered as natural polymorphisms were characterized as susceptible to ACV using plaque-reduction assay in MeWo cells. In parallel, the impact of TK changes on ACV phosphorylation activity was examined using a nonradioactive in vitro enzymatic assay.