Clonal deleted latent membrane protein 1 variants of Epstein-Barr virus are predominant in European extranodal NK/T lymphomas and disappear during successful treatment.

Extranodal natural killer/T-cell lymphomas (NK/TL), rare in Europe, are Epstein-Barr virus (EBV) associated lymphomas with poor outcomes. Here, we determined the virus type and analyzed the EBV latent membrane protein-1 (LMP1) gene sequence in NK/TL from French patients. Six clones of viral LMP1 were sequenced by Sanger technology in blood from 13 patients before treatment with an l-asparaginase based regimen and, for 8 of them, throughout the treatment. Blood LMP1 sequences from 21 patients without any known malignancy were tested as controls. EBV Type A was identified for 11/13 patients and for all controls. Before treatment, a clonal LMP1 gene containing a 30 bp deletion (del30) was found in 46.1% of NK/TL and only in 4.8% of controls. Treatment was less effective in these patients who died more rapidly than the others. Patients with a deleted strain evolving toward a wild-type strain during treatment reached complete remission. The LMP1 gene was sequenced by highly sensitive next-generation sequencing technology in five NK/TL nasopharyngeal biopsies, two of them originating from the previous patients. Del30 was present in 100% of the biopsies; two viruses at least coexisted in three biopsies. These results suggest that del30 may be associated with poor prognosis NK/TL and that strain evolution could be used as a potential marker to monitor treatment.

Q fever and pregnancy: experience from the Limoges Regional University Hospital.

INTRODUCTION: Q fever is an ubiquitous zoonosis caused by Coxiella burnetii. Its tropism for the uterus is a potential source of obstetric complications. MATERIALS AND METHODS: We describe the obstetric consequences of Q fever diagnosed during pregnancy from a series of cases. When an antenatal diagnosis was made, antibiotic therapy with roxithromycin (Rulid(®)) was started until delivery. RESULTS: Between 2007 and 2012, 30 patients were treated for Q fever diagnosed during pregnancy, i.e. 1.9 cases per 1000 people. The most common reasons for performing serology was intrauterine growth retardation, preterm labor and oligoamnios. Q fever was diagnosed as acute and chronic in 26 and 4 cases, respectively. Progression to chronic disease occurred in 8 % of acute forms of the diseases. The prevalence of obstetric complications was 66 %, including 10 % foetal deaths, 31 % preterm delivery and 27 % low birthweight <10th percentile. The obstetric complication rate amongst the 22 patients treated with ante partum macrolides was 60, 30 % of which involved prematurity and 33 % involved low growth. No cases of foetal death were found on treatment and no congenital malformation and placental or neonatal injury was found. No case of disease reactivation was diagnosed in the eight patients who became pregnant again. CONCLUSION: Q fever during pregnancy is responsible for severe obstetric complications. It must be diagnosed early and its clinical forms known in order to start appropriate antibiotic therapy.

HIV-1 subtype B-infected MSM may have driven the spread of transmitted resistant strains in France in 2007-12: impact on susceptibility to first-line strategies.

BACKGROUND: Our study describes the prevalence of transmitted drug resistance (TDR) among 1318 French patients diagnosed at the time of primary HIV-1 infection (PHI) in 2007-12. METHODS: HIV-1 resistance-associated mutations (RAMs) were characterized using both the 2009 WHO list of mutations and the French ANRS algorithm. A genotypic susceptibility score was estimated for each first-line recommended ART combination. RESULTS: Patients were mainly MSM (72.6%). Non-B variants were identified in 33.7% of patients. The proportion of TDR was estimated as 11.7% (95% CI 10.0-13.5). The prevalences of PI-, NRTI-, first-generation NNRTI and etravirine/rilpivirine-associated RAMs were 2.5%, 5.2%, 3.9% and 3.2%, respectively. Single, dual and triple class resistance was found in 9.6%, 1.0% and 1.1% of cases, respectively. Additionally, 5/331 strains isolated in 2010-12 had integrase inhibitor (II)-related RAMs (isolated E157Q mutation in all cases). TDR was more common among MSM than in other groups (12.9% versus 8.6%, P = 0.034), and in case of B versus non-B subtype infections (13.6% versus 7.9%, P = 0.002). The proportions of fully active combinations were ≥99.2%, ≥97.3% and ≥95.3% in cases of PI-, II- and NNRTI-based regimens, respectively. In 2010-12, the proportion of fully active efavirenz-based ART was lower in cases of subtype B versus non-B infection (P = 0.021). CONCLUSIONS: Compared with our previous studies, the proportion of NRTI- and first-generation NNRTI-related TDR has continued to decline in French seroconverters. However, subtype B-infected MSM could drive the spread of resistant HIV strains. Finally, we suggest preferring PI- or II- to NNRTI-based combinations to treat PHI patients.

Genotypic prediction of HIV-1 CRF01-AE tropism.

HIV-1 subtype CRF01-AE predominates in south Asia and has spread throughout the world. The virus tropism must be determined before using CCR5 antagonists. Genotypic methods could be used, but the prediction algorithms may be inaccurate for non-B subtypes like CRF01-AE and the correlation with the phenotypic approach has not been assessed. We analyzed 61 CRF01-AE V3 clonal sequences of known phenotype from the GenBank database. The sensitivity of the Geno2pheno10 genotypic algorithm was 91%, but its specificity was poor (54%). In contrast, the combined 11/25 and net charge rule was highly specific (98%) but rather insensitive (64%). We thus identified subtype CRF01-AE determinants in the V3 region that are associated with CXCR4 use and developed a new simple rule for optimizing the genotypic prediction of CRF01-AE tropism. The concordance between the predicted CRF01-AE genotype and the phenotype was 95% for the clonal data set. We then validated this algorithm by analyzing the data from 44 patients infected with subtype CRF01-AE, whose tropism was determined using a recombinant phenotypic entry assay and V3-loop bulk sequencing. The CRF01-AE genotypic tool was 70% sensitive and 96% specific for predicting CXCR4 use, and the concordance between genotype and phenotype was 84%, approaching the concordance obtained for predicting the tropism of HIV-1 subtype B. Genotypic predictions that use a subtype CRF01-AE-specific algorithm appear to be preferable for characterizing coreceptor usage both in pathophysiological studies and for ensuring the appropriate use of CCR5 antagonists.

Efficacy of L-asparaginase with methotrexate and dexamethasone (AspaMetDex regimen) in patients with refractory or relapsing extranodal NK/T-cell lymphoma, a phase 2 study.

Extranodal NK/T-cell lymphoma, nasal type, is a rare and highly aggressive disease with a grim prognosis. No therapeutic strategy is currently identified in relapsing patients. We report the results of a French prospective phase II trial of an L-asparaginase-containing regimen in 19 patients with relapsed or refractory disease treated in 13 centers. Eleven patients were in relapse and 8 patients were refractory to their first line of treatment. L-Asparaginase-based treatment yielded objective responses in 14 of the 18 evaluable patients after 3 cycles. Eleven patients entered complete remission (61%), and only 4 of them relapsed. The median overall survival time was 1 year, with a median response duration of 12 months. The main adverse events were hepatitis, cytopenia, and allergy. The absence of antiasparaginase antibodies and the disappearance of Epstein-Barr virus serum DNA were significantly associated with a better outcome. These data confirm the excellent activity of L-asparaginase-containing regimens in extranodal NK/T-cell lymphoma. L-Asparaginase-based treatment should thus be considered for salvage therapy, especially in patients with disseminated disease. First-line L-asparaginase combination therapy for extranodal NK/T-cell lymphoma warrants evaluation in prospective trials. This trial is registered at www.clinicaltrials.gov as #NCT00283985.

Chromosomally integrated human herpesvirus 6: questions and answers.

Chromosomally integrated human herpesvirus 6 (ciHHV-6) is a condition in which the complete HHV-6 genome is integrated into the host germ line genome and is vertically transmitted in a Mendelian manner. The condition is found in less than 1% of controls in the USA and UK, but has been found at a somewhat higher prevalence in transplant recipients and other patient populations in several small studies. HHV-6 levels in whole blood that exceed 5.5 log10 copies/ml are strongly suggestive of ciHHV-6. Monitoring DNA load in plasma and serum is unreliable, both for identifying and for monitoring subjects with ciHHV-6 due to cell lysis and release of cellular DNA. High HHV-6 DNA loads associated with ciHHV-6 can lead to erroneous diagnosis of active infection. Transplant recipients with ciHHV-6 may be at increased risk for bacterial infection and graft rejection. ciHHV-6 can be induced to a state of active viral replication in vitro. It is not known whether ciHHV-6 individuals are put at clinical risk by the use of drugs that have been associated with HHV-6 reactivation in vivo or in vitro. Nonetheless, we urge careful observation when use of such drugs is indicated in individuals known to have ciHHV-6. Little is known about whether individuals with ciHHV-6 develop immune tolerance for viral proteins. Further research is needed to determine the role of ciHHV-6 in disease.

Performance of the SureScreen Diagnostics COVID-19 antibody rapid test in comparison with three automated immunoassays.

Lateral flow immunoassays (LFIA) for detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies are used for population surveillance and potentially individual risk assessment. The performance of the SureScreen Diagnostics LFIA targeting the spike protein was evaluated in comparison with 3 automated assays (Abbott Alinity-i SARS-CoV-2 IgG, DiaSorin Liaison® SARS-CoV-2 S1/S2 IgG, Wantai SARS-CoV-2 Ab ELISA). We assessed sensitivity using 110 serum samples from PCR confirmed COVID-19 infected patients. Specificity was evaluated using 120 prepandemic samples, including potential cross-reactive antibodies samples. Sensitivity ranged between 93.3% and 98.7% on samples collected >14 days postsymptom onset. All assays achieved a specificity >98%. Moreover, its performance seems not to be affected by Alpha, Beta or Delta variants over a wide range of antibody titers. The latter showed a very good agreement with the Wantai and the Abbott assays and a substantial agreement with the DiaSorin assay. Our data demonstrate the good clinical performance of the SureScreen Diagnostics LFIA for SARS-CoV-2 seroprevalence screening.

One year of SARS-CoV-2 circulation in the Nouvelle-Aquitaine region, February 2021-2022, France.

Background: Since 2021, 3 variants of concern (VOC) have spread to France, causing successive epidemic waves. Objectives: To describe the features of Alpha, Delta and Omicron VOC circulation in the Nouvelle-Aquitaine region, France, between February 2021 and February 2022. Study design: Data from the three university hospitals (UH) of Nouvelle-Aquitaine were used to describe regional SARS-CoV-2 circulation (RT-PCR positive rates and identified VOC) as well as its consequences (total number of hospitalizations and admissions in intensive care unit). They were analyzed according to the predominant variant and compared with national data. Results: A total of 611,106 SARS-CoV-2 RT-PCR tests were performed in the 3 Nouvelle-Aquitaine UH during the study period. The 37,750 positive samples were analyzed by variant-specific RT-PCR or whole-genome sequencing. In 2021, Alpha VOC was detected from week 5 until week 35. Delta became the most prevalent variant (77.3%) in week 26, reaching 100% in week 35. It was replaced by Omicron, which was initially detected week 48, represented 77% of positive samples in week 52 and was still predominant in February 2022. The RT-PCR positive rates were 4.3, 4.2, and 21.9% during the Alpha, Delta and Omicron waves, respectively. The ratio between intensive care unit admissions and total hospitalizations was lower during the Omicron wave than during the two previous waves due to the Alpha and Delta variants. Conclusion: This study highlighted the need for strong regional cooperation to achieve effective SARS-CoV-2 epidemiological surveillance, in close association with the public health authorities.

Epstein-Barr Virus (EBV) Is Mostly Latent and Clonal in Angioimmunoblastic T Cell Lymphoma (AITL).

The Epstein-Barr virus (EBV) is associated with angioimmunoblastic T cell lymphoma (AITL), a peripheral T lymphoma of poor prognosis in at least 90% of cases. The role of EBV in this pathology is unknown. Using next-generation sequencing, we sequenced the entire EBV genome in biopsies from 18 patients with AITL, 16 patients with another EBV-associated lymphoma, and 2 controls. We chose an EBV target capture method, given the high specificity of this technique, followed by a second capture to increase sensitivity. We identified two main viral strains in AITL, one of them associated with the mutations BNRF1 S542N and BZLF1 A206S and with mutations in the EBNA-3 and LMP-2 genes. This strain was characterized in patients with short post-diagnosis survival. The main mutations found during AITL on the most mutated latency or tegument genes were identified and discussed. We showed that the virus was clonal in all the AITL samples, suggesting that it may be involved in this pathology. Additionally, EBV was latent in all the AITL samples; for one sample only, the virus was found to be latent and probably replicative, depending on the cells. These various elements support the role of EBV in AITL.

Comprehensive Epstein-Barr Virus Transcriptome by RNA-Sequencing in Angioimmunoblastic T Cell Lymphoma (AITL) and Other Lymphomas.

The Epstein-Barr virus (EBV) is associated with angioimmunoblastic T cell lymphoma (AITL) in more than 80% of cases. Few studies have focused on this association and it is not clear now what role the virus plays in this pathology. We used next-generation sequencing (NGS) to study EBV transcriptome in 14 AITLs compared to 21 other lymphoma samples and 11 cell lines including 4 lymphoblastoid cell lines (LCLs). Viral transcripts were recovered using capture probes and sequencing was performed on Illumina. Bam-HI A rightward transcripts (BARTs) were the most latency transcripts expressed in AITLs, suggesting they may play a role in this pathology. Thus, BARTs, already described as highly expressed in carcinoma cells, are also very present in AITLs and other lymphomas. They were poorly expressed in cell lines other than LCLs. AITLs showed a latency IIc, with BNLF2a gene expression. For most AITLs, BCRF1, which encodes a homologous protein of human interleukin 10, vIL-10, was in addition expressed. This co-expression can contribute to immune escape and survival of infected cells. Considering these results, it can be assumed that EBV plays a pathogenic role in AITLs.

Factors associated with virological response to etravirine in nonnucleoside reverse transcriptase inhibitor-experienced HIV-1-infected patients.

To identify factors associated with virological response (VR) to an etravirine (ETR)-based regimen, 243 patients previously treated with nonnucleoside reverse transcriptase inhibitors (NNRTIs) were studied. The impact of baseline HIV-1 RNA, CD4 cell count, past NNRTIs used, 57 NNRTI resistance mutations, genotypic sensitivity score (GSS) for nucleoside reverse transcriptase inhibitors (NRTIs) and protease inhibitors (PIs), and the number of new drugs used with ETR for the first time on the VR to an ETR regimen were investigated. Among the 243 patients, the median baseline HIV-1 RNA level was 4.4 log(10) copies/ml (interquartile range [IQR], 3.7 to 4.9) and the median CD4 count was 175 cells/mm(3) (IQR, 69 to 312). Patients had been previously exposed to a median of 6 NRTIs, 1, NNRTI, and 5 PIs. Overall, 82% of patients achieved a VR at month 2, as defined by a decrease of at least 1.5 log(10) copies/ml and/or HIV-1 RNA level of <50 copies/ml. No difference in VR was observed between patients receiving or not a boosted PI in combination with ETR. Factors independently associated with a better VR to ETR were the number of drugs (among enfuvirtide, darunavir, or raltegravir) used for the first time in combination with ETR and the presence of the K103N mutation at baseline. Mutations Y181V and E138A were independently associated with poor VR, whereas no effect of the Y181C on VR was observed. In conclusion, ETR was associated with high response rates in NNRTI-experienced patients in combination with other active drugs regardless of the therapeutic class used.

Amoxicillin-induced flare in patients with DRESS (Drug Reaction with Eosinophilia and Systemic Symptoms): report of seven cases and demonstration of a direct effect of amoxicillin on Human Herpesvirus 6 replication in vitro.

Amoxicillin is known to induce exanthema in patients with EBV-induced infectious mononucleosis. It is widely recognized that the reactivation of herpesviruses, including HHV-6 (Human Herpesvirus 6) and EBV (Epstein Barr virus) is associated with DRESS (Drug Reaction with Eosinophilia and Systemic Symptoms). We report 7 cases of amoxicillin-induced flare in patients with DRESS induced by other drugs and investigate whether amoxicillin may have a direct effect on HHV-6 replication in vitro. 7 cases of DRESS with amoxicillin-induced flare were retrospectively analysed. The influence of amoxicillin on HHV-6 HST strain replication was studied in vitro in a human T lymphoblastoid MT4 cell line. The viral replication was quantified by immunofluorescence assay and by real-time polymerase chain reaction (PCR). Comparisons were performed using the Student's t test. Amoxicillin-induced flare was observed in 7 patients with DRESS induced by other drugs. In two cases HHV-6 reactivation was studied and was demonstrated by PCR. Amoxicillin neither modified cell viability nor cell proliferation for the range of tested concentrations. Amoxicillin increased the replication of HHV-6 at 25 microg*mL-1 and 50 microg*mL-1. Amoxicillin may induce a flare of DRESS, possibly by acting directly on herpesvirus replication.

The survival of herpes simplex virus in preserved murine corneas.

BACKGROUND/AIMS: This study investigates the survival of HSV in infected mouse corneas, in the conditions of normal human eye bank preservation. METHODS: Hundred seventy-two BALB/C mice infected with herpes simplex virus 1 (HSV-1) (KOS) were randomly assigned to either: no preservation in group 1 (n = 62), 31 degrees C preservation for 3 weeks in group 2 (n = 70) or 4 degrees C preservation for 8 days in group 3 (n = 40). The presence of HSV-1 was thereafter detected by viral culture and PCR. RESULTS: In groups 1, 2 and 3, HSV-1 was detected by culture in 22 (35.5%), 1 (1.4%) and 0 (0.0%) of the corneas, and by PCR in 27 (43.7%), 3 (4.2%) and 7 (17.5%) of the corneas respectively. When compared to group 1, HSV was detected significantly less often in groups 2 (p < 0.0001) and 3 (p < 0.0001). CONCLUSIONS: HSV-1 DNA undergoes a degradation during corneal preservation.

HIV-2 Primary Infection in a French 69-Year-Old Bisexual Man.

More than 1 million individuals, mainly in West Africa, are thought to be infected with HIV-2. Acute HIV-2 infection is rarely observed, only 2 primary infections have been described to date. We report a detailed case of HIV-2 primary infection in a 69-year-old French bisexual Caucasian man, thereby providing valuable insights into HIV-2 early infection.

Multiplex real-time PCR assay for simultaneous quantitation of human cytomegalovirus and herpesvirus-6 in polymorphonuclear and mononuclear cells of transplant recipients.

Human cytomegalovirus (HCMV) and human herpesvirus-6 (HHV-6) are two closely related viruses, which belong to the Herpesviridae family. Following primary infection, they are thought to persist for life as latent forms in mononuclear cells. HCMV and HHV-6 can cause considerable morbidity in immunocompromised individuals, such as transplant patients. A sensitive and specific LightCycler multiplex real-time PCR assay based on fluorescence energy transfer (known as FRET) was developed. This assay, by using two sets of hybridization probes specific for HHV-6 (A and B) and HCMV, can differentiate reliably and quantify simultaneously both viruses in order to diagnose reactivation processes. The assay was optimized and the lower limit of detection for both viruses was determined to be 10 viral genome copies per reaction. Both viruses were quantified in 83 peripheral blood mononuclear cells (PBMCs) and 87 polymorphonuclear leukocytes (PMNLs) collected from 32 transplant recipients. This multiplex real-time quantitative PCR was finally compared with two other quantitation and detection assays used daily in laboratory (PCR DIG detection and antigenemia for HCMV, TaqMan Assay for HHV-6). This technique can be useful for the differentiation and quantitation of HCMV and HHV-6 for monitoring transplant patients.

A reverse transcription-nested PCR assay for HHV-6 mRNA early transcript detection after transplantation.

Monitoring of human herpesvirus-6 (HHV-6) reactivation is important, especially in immunocompromised patients such as transplant recipients. Reverse transcription PCR (RT-PCR) is a useful method to distinguish between latent and active infection. Here, a RT-nested PCR coupled with a colorimetric plate hybridization assay was established to detect HHV-6 types A and B U79/80 mRNAs. After confirming the reliability of the assay on HHV-6 cultures, it was applied to the detection of HHV-6 reactivation after renal (27 patients), bone marrow (14 patients) or heart (7 patients) transplantation. A total of 206 blood samples were tested from renal (137), bone marrow (58) and heart (11) transplant recipients. U79/80 mRNAs were found in 32 samples that were considered as indicative of HHV-6 reactivation: 15, 13 and 5 from kidney, bone marrow and heart transplant recipients, respectively. Finally, U79/80 mRNA detection was correlated with clinical manifestations including leucopenia, skin rash, graft rejection or dysfunction and diarrhoea.

Factors associated with poor outcomes among adults hospitalized for influenza in France: A three-year prospective multicenter study.

BACKGROUND: Influenza is an important cause of serious illness and death, particularly in elderly and high-risk groups. OBJECTIVES: Aim of this study was to identify factors associated with poor outcomes among adults hospitalized in France for laboratory-confirmed seasonal influenza. STUDY DESIGN: Patients hospitalized for influenza were identified in a prospective, multicenter study carried out in French hospitals during three consecutive influenza seasons (2012-2015). Influenza virus infection was confirmed by reverse transcription polymerase chain reaction. Sociodemographic and clinical variables were compared according to the virus type and subtype. Risk factors for complications, intensive care unit (ICU) admission and death were analyzed by backward stepwise logistic regression. RESULTS: The study population consisted of 566 patients, of whom 56% were older than 65 years and 82% had underlying chronic illnesses. Type A influenza viruses infected 422 patients (75%), including subtype H3N2 in 239 patients (57%). The prior vaccine coverage rate was 38%. Complications occurred in 255 patients (45%), consisting mainly of pneumonia (n=143, 30%) and respiratory failure (n=116, 20%). Eighty-three patients (15%) were admitted to an ICU, and the in-hospital mortality rate was 4% (n=21). Sixty-six patients (12%) received oseltamivir. Age over 65 years was the only identified risk factor for complications. Risk factors for ICU admission were an absence of vaccination, no oseltamivir administration before admission, pre-existing chronic respiratory disease, and current smoking. Age over 65 years and ICU admission were risk factors for death. CONCLUSIONS: Older individuals and patients with underlying conditions are most at risk of influenza complications. Vaccination and early oseltamivir administration, both of which are recommended for these patients, appear to reduce ICU admissions.

Transmission of viruses through corneal transplantation.

Transmission of viruses through transplantation has become a major concern for surgeons and tissue banks. Keratoplasty has been the first transplantation procedure in which transmission of a virus from the donor to the recipient has been proven, and recently, herpes has been suggested as a transmittable disease via corneal transplantation. In this article we review the history of viral transmission through transplantation, especially focusing on keratoplasty, and develop the recent updates on transmission of HSV through corneal grafting, its implications for surgeons and eye banks, and further insights into viral security in eye banks.

Influenza burden of illness: estimates from a national prospective survey of household contacts in France.

BACKGROUND: The burden of influenza among ambulatory patients is still relatively unknown, although this knowledge is crucial for evaluating strategies against influenza. We estimated the impact of influenza in terms of uncomplicated morbidity and its consequences on health care utilization and lost workdays. METHODS: A national prospective household contact study between January 4, 2000, and March 15, 2000, in France recruited the households of 946 persons who visited a physician (index cases); 395 households with influenza-positive index cases completed the follow-up, which assessed the clinical impact of influenza, medical visits, treatment, and lost workdays in these index cases and their contacts. RESULTS: Of 817 assessable household contacts, 313 developed clinical influenza (secondary cases); 178 (57%) of them visited a physician at least once (consulting secondary cases). The median duration of illness was 8 days (95% confidence interval [CI], 7-8 days) in index cases, 7 days (95% CI, 7-8 days) in consulting secondary cases, and 4 days (95% CI, 3-5 days) in nonconsulting secondary cases (P<.001); the median duration of treatment in these groups was 8 days (95% CI, 8-9 days), 8 days (95% CI, 7-10 days), and 5 days (95% CI, 4-6 days), respectively (P<.001); and their mean +/- SD number of lost workdays was 4.0 +/- 2.8, 2.9 +/- 2.5, and 0.3 +/- 0.6, respectively, in working adults (P<.001). CONCLUSIONS: These results confirm the substantial burden of illness of influenza. The results should be useful for evaluating the cost-effectiveness of strategies against influenza.

Serological pattern 'anti-hepatitis B core alone' in HIV or hepatitis C virus-infected patients is not fully explained by hepatitis B surface antigen mutants.

We determined the frequency of hepatitis B virus (HBV) occult infection and studied S gene mutations in HIV or hepatitis C virus-co-infected patients with persistent pattern 'anti-hepatitis B core alone'. HBV DNA was particularly frequent in HIV-positive patients. Sequencing of the S gene showed that only two HIV-infected patients harboured mutants in the major hydrophilic loop; the absence of hepatitis B surface antigen detection could not be explained by diagnostic escape mutants.