Journal of Clinical Virology最新文献_第9页

Use of polyethylene glycol precipitation and ultracentrifugation to enhance the sensitivity of hepatitis B virus DNA detection 采用聚乙二醇沉淀法和超离心法提高乙型肝炎病毒DNA检测的灵敏度

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-05-03 DOI: 10.1016/j.jcv.2025.105802

Michael X. Fu , Osmany Larralde , Richard Mayne , Kai Kean , Kaitlin Reid , Monique Andersson , Tanya Golubchik , Jane A. McKeating , Lisa Jarvis , William L. Irving , Peter Simmonds , Heli Harvala

Background

Sensitive molecular detection of hepatitis B virus (HBV) DNA is crucial for diagnosing and managing occult hepatitis. To improve the sensitivity of HBV DNA detection, we compared the effectiveness of polyethylene glycol (PEG) precipitation and ultracentrifugation to concentrate DNA prior to extraction.

Methods

Twenty-three HBV DNA-positive samples with low viral loads were compared between the extraction of standard (0.2 mL) and larger volumes (5 mL) of plasma, through PEG precipitation of 10 mL and 20 mL of plasma, and ultracentrifugation from 35 mL of plasma. The effectiveness of the methods for HBV DNA detection was assayed by quantitative PCR. For genetic characterisation, Sanger sequencing of amplicons and targeted Illumina sequencing were used. Costs, sample capacities, and turnaround times were compared.

Results

DNA was detected in a greater number of samples using PEG and ultracentrifugation (detecting up to all 23 samples) compared to more standard extraction methods (detecting at least 18 samples). Efficiencies of recovery of HBV DNA from samples were comparable in all concentration methods. HBV and other DNA viruses, such as human herpesviruses and anelloviruses, were detected in samples and at higher read counts with PEG concentration than without. The availability, cost, relative simplicity, and throughput of PEG precipitation conferred further advantages to ultracentrifugation.

Conclusions

PEG precipitation from large volumes of plasma is a practical and economical alternative to ultracentrifugation and could be a similarly effective concentration method for low viral load samples in blood donation and clinical virology laboratories.

背景：乙型肝炎病毒（HBV） DNA的敏感分子检测对于诊断和治疗隐匿性肝炎至关重要。为了提高HBV DNA检测的灵敏度，我们比较了聚乙二醇（PEG）沉淀和超离心在提取前浓缩DNA的有效性。方法对23例低病毒载量HBV dna阳性样本，分别采用标准血浆（0.2 mL）和大容量血浆（5 mL）、PEG沉淀血浆（10 mL和20 mL）、超离心血浆（35 mL）进行比较。采用定量PCR方法检测HBV DNA的有效性。为了进行遗传表征，扩增子采用Sanger测序和靶向Illumina测序。比较了成本、样品容量和周转时间。结果与更多标准提取方法（检测至少18个样品）相比，使用PEG和超离心（检测最多23个样品）在更多样品中检测到dna。从样品中回收HBV DNA的效率在所有浓度方法中都是相当的。在样品中检测到HBV和其他DNA病毒，如人类疱疹病毒和无绒病毒，并且在PEG浓度下检测到更高的读取计数。聚乙二醇沉淀的可用性、成本、相对简单性和吞吐量赋予了超离心进一步的优势。结论从大量血浆中提取speg沉淀是一种实用且经济的超离心方法，对于献血和临床病毒学实验室的低病毒载量样品是一种同样有效的浓缩方法。

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引用次数: 0

Comparison of mid-turbinate nasal and combined nasal-throat specimen types for detection of respiratory viruses in children 儿童中鼻甲鼻与合并鼻咽标本类型检测呼吸道病毒的比较

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-29 DOI: 10.1016/j.jcv.2025.105801

Irem Onalan , Zheyi Teoh , Sarah L. Steele , Eileen J. Klein , Bonnie Strelitz , Kirsten Lacombe , Erin M. Sullivan , Arun K. Nalla , Danielle M. Zerr , Janet A. Englund

Background

The source of respiratory specimens may impact the detection of respiratory viruses. It may also have implications for research participant recruitment, caregiver acceptability due to concerns for patient comfort, and potential risk of aerosolization during epidemics/pandemics.

Objective

To determine the impact of collecting a throat swab (TS) in addition to a mid-turbinate nasal swab (MTS) by comparing agreement of viral detection and relative viral loads.

Study design

We reviewed molecular detection results from 2548 children enrolled in the New Vaccine Surveillance Network at Seattle Children’s Hospital from 11/2015–05/2019. Participants with a clinical MTS who agreed to collection of a combined TS and MTS (TS&MTS) for research were included. All specimens were tested using FilmArray^R Respiratory Panel (Biofire Diagnostics). Viral detection from MTS and TS&MTS were compared. Relative viral loads were compared between specimens with concordant (same viruses detected) and discordant (different or additional viruses detected) results.

Results

Results from 743 participants with clinical MTS and research TS&MTS specimens were compared; Viral detections were similar between the two groups, including 596 (80.2 %) paired results that were concordant. The most common discordant viruses were rhinovirus/enterovirus, respiratory syncytial virus, and adenovirus. Mean relative viral loads were lower in discordant specimens compared to concordant specimens, regardless of specimen source.

Conclusion

Comparison of clinical and research specimens revealed that respiratory viral detection was similar with or without an added TS. Lower relative viral loads of discordant specimens suggest that a combined TS&MTS may not improve viral detection for clinically significant pathogens.

呼吸道标本的来源可能影响呼吸道病毒的检测。它还可能对研究参与者的招募、护理人员对患者舒适度的接受程度以及流行病/大流行期间雾化的潜在风险产生影响。目的通过比较病毒检测结果与相对病毒载量的一致性，确定除中鼻甲鼻拭子外还采集咽拭子（TS）的影响。我们回顾了2015年11月至2019年5月在西雅图儿童医院新疫苗监测网络中登记的2548名儿童的分子检测结果。患有临床MTS的参与者同意收集TS和MTS （TS&；MTS）联合用于研究。所有标本均使用FilmArrayR呼吸面板（Biofire Diagnostics）进行检测。比较MTS和TS&；MTS的病毒检测结果。比较结果一致（检测到相同病毒）和不一致（检测到不同或额外的病毒）的标本之间的相对病毒载量。结果对743例临床MTS和研究MTS标本的结果进行了比较；两组之间的病毒检测结果相似，其中596（80.2%）对结果一致。最常见的不一致病毒是鼻病毒/肠病毒、呼吸道合胞病毒和腺病毒。不一致标本的平均相对病毒载量低于一致标本，无论标本来源如何。结论临床标本与研究标本的比较表明，添加或不添加TS的呼吸道病毒检出率相似，不一致标本的相对病毒载量较低，表明联合使用TS和MTS可能不能提高对临床重要病原体的病毒检出率。

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引用次数: 0

Universal screening for congenital Cytomegalovirus infection using saliva from the neonate 利用新生儿唾液对先天性巨细胞病毒感染进行普遍筛查

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-20 DOI: 10.1016/j.jcv.2025.105800

Gabriele Halwachs-Baumann , Oliver Wagner , Yarub Salaheddin , Eveline Ziebermayr , Lukas Angleitner-Boubenizek , Georg Grüßenberger , Ulla Folger-Buchegger

Background

Congenital Cytomegalovirus infection (cCMV) is the most common intrauterine infection in developed countries. The diagnostic gold standard is detection of CMV in neonatal urine. Considering difficulties in urine sample collection, saliva is an alternative specimen providing supportive material for early diagnosis and timely intervention.

Objectives

Aim of this study was to evaluate a universal screening program using saliva as target specimen, and urine as confirmatory material.

Results

Between May 2019 and November 2024, 5168 newborns were screened for CMV shedding in saliva, 45 neonates were tested positive. From 44 neonates a urine sample was examined. Twelve out of forty-four neonates had concordant results (prevalence = 0,232 %) with five newborns showing CMV related symptoms. In saliva, the difference between the Ct-value of the cCMV negative and the cCMV positive group was significant (38.59 vs. 28.56, p-value < 0.0001). At a Ct-value cut-off of 38.8 sensitivity was 100 %, specificity 53.1 %, positive predictive value 44 %, and negative predictive value 100 %. ROC analysis yielded an AUC value of 0.930. The mean virus load in urine was 31,266,663 copies/ml.

Discussion

This study reports a prevalence of 0,232 % for cCMV infection in the catchment area of the hospital Steyr. PCR based CMV detection in saliva is a good screening tool, but one must be aware of false positive results, arising from possible contamination. In our cohort CMV positive vaginal or cervical secretion seems more likely being the source of contamination than CMV positive breast milk. The high false positive rate in saliva makes confirmatory testing in urine samples mandatory.

先天性巨细胞病毒感染（cCMV）是发达国家最常见的宫内感染。诊断的金标准是新生儿尿液中巨细胞病毒的检测。考虑到尿液样本采集的困难，唾液是一种可替代的样本，为早期诊断和及时干预提供支持材料。目的探讨一种以唾液为目标样本，尿液为确认材料的通用筛查方案。结果2019年5月至2024年11月，对5168名新生儿进行唾液巨细胞病毒脱落筛查，45名新生儿检测呈阳性。对44名新生儿进行了尿样检查。44名新生儿中有12名结果一致（患病率= 0,232%），其中5名新生儿出现巨细胞病毒相关症状。在唾液中，cCMV阴性组和cCMV阳性组的ct值差异有统计学意义(38.59比28.56，p值<；0.0001)。ct值为38.8时，敏感性为100%，特异性为53.1%，阳性预测值为44%，阴性预测值为100%。ROC分析的AUC值为0.930。尿中平均病毒载量为31,266,663拷贝/ml。本研究报告了斯太尔医院集水区cCMV感染率为0,232%。基于PCR的唾液巨细胞病毒检测是一个很好的筛选工具，但必须注意假阳性结果，可能引起污染。在我们的队列中，巨细胞病毒阳性的阴道或宫颈分泌物似乎比巨细胞病毒阳性的母乳更可能是污染源。唾液的高假阳性率使得尿样的确证测试成为强制性的。

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引用次数: 0

Human cytomegalovirus (HCMV) glycoprotein M occurs in three distinct genotypes in laboratory strains and clinical isolates 人巨细胞病毒（HCMV）糖蛋白M在实验室菌株和临床分离株中出现三种不同的基因型

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-20 DOI: 10.1016/j.jcv.2025.105799

Alia Al-Alfard , Zahrah Buhamad , Julia Duffey , Simon Lovell , Paul Klapper , Pamela Vallely

Background

HCMV carries virus-encoded envelope glycoproteins important for viral attachment, entry into host cells and replication. These glycoproteins have polymorphic features resulting in distinct genotypes some of which have been associated with disease outcome. Glycoprotein complex II comprises glycoproteins M (gM) and N (gN) and is essential for viral replication. Glycoprotein N is highly polymorphic while gM is highly conserved. The existence of distinct gM genotypes has not previously been reported.

Study design

PCR amplification of the entire gM gene (1298 bp) was carried out on five HCMV laboratory strains (AD169, Towne, Davis, Toledo, Merlin) and ten clinical isolates. PCR product was sequenced and nucleotide and aminoacid phylogenetic trees constructed for these samples and for an additional 287 HCMV gM sequences obtained from NCBI database. The hydrophobicity of the predicted protein sequences was compared.

Results

Three distinct genotypes were identified (gM1, gM2 and gM3) with every sequence aligning with one genotype, suggesting they are stable polymorphisms rather than random nucleotide substitutions. Aminoacid translation of the nucleotide sequences merged gM1 and gM2, but gM3 remained as a distinct and separate type. The aminoacid substitutions led to changes in the hydrophobicity of the 3 glycoprotein types particularly between gM3 and the other two types.

Conclusions

Glycoprotein M is highly conserved, but we have identified three distinct genotypes, arising from variability in a small region of the genome. These changes altered the predicted hydrophobicity of the glycoprotein potentially altering the conformational structure of the glycoprotein and affecting its function in vivo.

dhcmv携带病毒编码的包膜糖蛋白，对病毒附着、进入宿主细胞和复制很重要。这些糖蛋白具有多态性特征，导致不同的基因型，其中一些与疾病结果相关。糖蛋白复合物II包括糖蛋白M （gM）和N (gN)，是病毒复制所必需的。糖蛋白N具有高度多态性，而gM具有高度保守性。存在不同的转基因基因型以前没有报道过。对5株HCMV实验室菌株（AD169、Towne、Davis、Toledo、Merlin）和10株临床分离株进行了gM全基因扩增（1298 bp）。对PCR产物进行测序，并对这些样本和NCBI数据库中另外287个HCMV gM序列构建核苷酸和氨基酸系统发育树。比较了预测蛋白序列的疏水性。结果鉴定出三种不同的基因型（gM1、gM2和gM3），每个序列都与一个基因型一致，表明它们是稳定的多态性，而不是随机的核苷酸替换。氨基酸翻译的核苷酸序列融合了gM1和gM2，但gM3仍然是一个独特的和独立的类型。氨基酸的取代导致3种糖蛋白的疏水性发生变化，特别是在gM3和其他两种糖蛋白之间。结论糖蛋白M是高度保守的，但我们已经确定了三种不同的基因型，这些基因型是由基因组一小部分区域的变异性引起的。这些变化改变了糖蛋白的预测疏水性，可能改变糖蛋白的构象结构并影响其在体内的功能。

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引用次数: 0

Insights into the clinical and molecular epidemiology of an infections outbreak of human parvovirus B19 in France, 2023–2024 2023-2024年法国人细小病毒B19感染爆发的临床和分子流行病学分析

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-18 DOI: 10.1016/j.jcv.2025.105798

Elisa Creuzet , Wendy Pulby , Christine Archimbaud , Amélie Brebion , Hélène Chabrolles , Audrey Mirand , Ophélie Perruche , Mathilde Picard , Christel Regagnon , Céline Lambert , Siméon Bakyono , Jean-Luc Bailly , Maxime Bisseux , Cécile Henquell

Background

The human parvovirus B19 (B19V) infections cycle occurs in 3- to 4-year periods and is responsible for benign childhood erythema infectiosum. It is also associated with transient aplastic crisis in patients with underlying hemolytic diseases and with severe fetal sometimes fatal infection. This study investigated the epidemiological, clinical and molecular characteristics of an unusually large 2023–2024 outbreak of B19V.

Methods

Laboratory-confirmed cases were retrospectively and prospectively recorded at the Clermont-Ferrand University Hospital, France, between January, 2018 and November, 2023 and between December 2023 and May 2024 (2023/2024), respectively. Demographical and clinical data were investigated for the 2023/2024 period. Subgenome sequences (2690 nt) were obtained by next generation sequencing for virus genotyping and temporal molecular analysis.

Results

The positive rate of B19V positive laboratory-confirmed cases was seven times higher between December 2023 and May 2024 than in the previous 5-year period (14.6 % vs 2.1 %, p < 0.001). No atypical clinical presentation or increased pathogenicity were observed, but this large outbreak resulted in a higher number of severe infections in pregnant women (8/16, 50.0 % of fetal complications) and those with chronic anemia. Phylogenetic analysis revealed that the 2023/2024 outbreak in France and Europe was mainly driven by a pre-existing lineage of B19V 1a subgenotype that emerged in 2017 (95 % highest posterior density interval: 2000–2018).

Conclusions

The recent epidemic of B19V infections re-illustrates the immunity gap of the post-pandemic COVID-19 pandemic. This highlight the impact of any outbreak on at-risk population and the need for a more global and genomic surveillance.

人细小病毒B19 （B19V）感染周期发生在3- 4年，是造成儿童良性感染性红斑的原因。它也与潜在溶血性疾病患者的短暂性再生危象和严重的胎儿有时致命的感染有关。本研究调查了2023-2024年一次异常大规模B19V暴发的流行病学、临床和分子特征。方法回顾性和前瞻性记录2018年1月至2023年11月和2023年12月至2024年5月（2023/2024）法国克莱蒙费朗大学医院实验室确诊病例。调查了2023/2024年期间的人口统计学和临床数据。下一代测序获得2690 nt亚基因组序列，用于病毒基因分型和时间分子分析。结果2023年12月- 2024年5月B19V阳性实验室确诊病例阳性率比前5年同期(14.6% vs 2.1%, p <；0.001)。没有观察到不典型的临床表现或增加的致病性，但这次大爆发导致孕妇（8/16,50.0%的胎儿并发症）和慢性贫血的严重感染人数增加。系统发育分析显示，法国和欧洲2023/2024年爆发的疫情主要是由2017年出现的B19V 1a亚基因型预先存在的谱系驱动的（95%最高后验密度区间：2000-2018年）。结论近期B19V感染的流行再次说明了大流行后COVID-19大流行的免疫缺口。这突出了任何疫情对高危人群的影响，以及加强全球和基因组监测的必要性。

{"title":"Insights into the clinical and molecular epidemiology of an infections outbreak of human parvovirus B19 in France, 2023–2024","authors":"Elisa Creuzet , Wendy Pulby , Christine Archimbaud , Amélie Brebion , Hélène Chabrolles , Audrey Mirand , Ophélie Perruche , Mathilde Picard , Christel Regagnon , Céline Lambert , Siméon Bakyono , Jean-Luc Bailly , Maxime Bisseux , Cécile Henquell","doi":"10.1016/j.jcv.2025.105798","DOIUrl":"10.1016/j.jcv.2025.105798","url":null,"abstract":"<div><h3>Background</h3><div>The human parvovirus B19 (B19V) infections cycle occurs in 3- to 4-year periods and is responsible for benign childhood erythema infectiosum. It is also associated with transient aplastic crisis in patients with underlying hemolytic diseases and with severe fetal sometimes fatal infection. This study investigated the epidemiological, clinical and molecular characteristics of an unusually large 2023–2024 outbreak of B19V.</div></div><div><h3>Methods</h3><div>Laboratory-confirmed cases were retrospectively and prospectively recorded at the Clermont-Ferrand University Hospital, France, between January, 2018 and November, 2023 and between December 2023 and May 2024 (2023/2024), respectively. Demographical and clinical data were investigated for the 2023/2024 period. Subgenome sequences (2690 nt) were obtained by next generation sequencing for virus genotyping and temporal molecular analysis.</div></div><div><h3>Results</h3><div>The positive rate of B19V positive laboratory-confirmed cases was seven times higher between December 2023 and May 2024 than in the previous 5-year period (14.6 % vs 2.1 %, p < 0.001). No atypical clinical presentation or increased pathogenicity were observed, but this large outbreak resulted in a higher number of severe infections in pregnant women (8/16, 50.0 % of fetal complications) and those with chronic anemia. Phylogenetic analysis revealed that the 2023/2024 outbreak in France and Europe was mainly driven by a pre-existing lineage of B19V 1a subgenotype that emerged in 2017 (95 % highest posterior density interval: 2000–2018).</div></div><div><h3>Conclusions</h3><div>The recent epidemic of B19V infections re-illustrates the immunity gap of the post-pandemic COVID-19 pandemic. This highlight the impact of any outbreak on at-risk population and the need for a more global and genomic surveillance.</div></div>","PeriodicalId":15517,"journal":{"name":"Journal of Clinical Virology","volume":"178 ","pages":"Article 105798"},"PeriodicalIF":4.0,"publicationDate":"2025-04-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143864515","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Emergence of influenza A(H1N1)pdm09 6B.1A.5a.2a and 6B.1A.5a.2a.1 subclades leading to subtyping failure in a commercial molecular assay 甲型H1N1流感pdm09 6B.1A.5a的出现。a. a. a. a. a. a. a. b。在商业分子分析中，1个亚枝导致分型失败

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-09 DOI: 10.1016/j.jcv.2025.105797

Bosung Park, Eun Jeong Won, Heungsup Sung, Mi-Na Kim

Background

During the 2023–2024 and early 2024–2025 influenza seasons, several influenza A-positive specimens in our laboratory failed subtyping for H1, H1pdm09, and H3 using the Allplex Respiratory Panel 1 (Allplex RP1) (Seegene Inc.). This study aimed to identify the cause of these subtyping failures.

Materials and methods

Between August 2023 and December 2024, 23 nasopharyngeal specimens tested positive for influenza A but were unsubtypeable for H1, H1pdm09, and H3. Confirmatory testing by the manufacturer included target-specific PCR for the M and HA genes, followed by sequencing to determine subclades.

Results

Among the 23 unsubtypeable specimens, 22 yielded PCR products for sequencing. Of these, 21 belonged to subclade 6B.1A.5a.2a.1 and one to 6B.1A.5a.2a. Sequence analysis revealed mismatches in the H1pdm09 primer/probe-binding regions of Allplex RP1, explaining the subtyping failures. Despite testing negative for H1pdm09 in Allplex RP1, sequencing confirmed their classification as H1N1pdm09 subclades with HA gene mutations.

Conclusions

Subclades 6B.1A.5a.2a.1 and 6B.1A.5a.2a harbour mutations that contributed to subtyping failures in some specimens tested with a commercial assay. While unsubtypeable influenza A results often raise concerns about emerging strains, sequencing confirmed that all unsubtypeable specimens tested with Allplex RP1 belonged to H1N1pdm09 within recognised subclades. Thus, such subtyping failures in this assay do not necessarily indicate a novel or zoonotic virus, though genomic surveillance remains essential.

背景在2023-2024年和2024-2025年初的流感季节，我们实验室的一些甲型流感阳性标本未能通过Allplex Respiratory Panel 1 (Allplex RP1) (Seegene Inc.)进行H1、H1pdm09和H3亚型检测。材料与方法在 2023 年 8 月至 2024 年 12 月期间，23 份鼻咽标本的甲型流感检测结果呈阳性，但无法对 H1、H1pdm09 和 H3 进行亚型鉴定。制造商进行的确证检测包括 M 和 HA 基因的目标特异性 PCR，然后进行测序以确定亚型。其中，21 个属于 6B.1A.5a.2a.1 亚支系，1 个属于 6B.1A.5a.2a。序列分析表明，Allplex RP1 的 H1pdm09 引物/探针结合区存在错配，这解释了亚型划分失败的原因。结论6B.1A.5a.2a.1和6B.1A.5a.2a亚支系存在突变，导致一些用商业检测方法检测的样本亚型鉴定失败。虽然无法分型的甲型流感结果往往会引起人们对新出现菌株的担忧，但测序结果证实，用 Allplex RP1 检测的所有无法分型的样本都属于公认亚支系中的 H1N1pdm09。因此，尽管基因组监测仍然非常重要，但该检测方法的亚型鉴定失败并不一定表明存在新型病毒或人畜共患病毒。

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引用次数: 0

Sustained circulation of enterovirus D68 in Europe in 2023 and the continued evolution of enterovirus D68 B3-lineages associated with distinct amino acid substitutions in VP1 protein 2023年欧洲肠道病毒D68的持续传播以及与VP1蛋白不同氨基酸替换相关的肠道病毒D68 b3谱系的持续进化

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-05 DOI: 10.1016/j.jcv.2025.105785

Aurora Hirvonen , Caroline Klint Johannesen , Peter Simmonds , Thea K Fischer , Heli Harvala , Kimberley S.M. Benschop , on behalf of ENPEN study collaborators

Background

Enterovirus D68 (EV-D68) causes respiratory disease ranging from mild to severe and in rare cases a paralytic syndrome, called acute flaccid myelitis (AFM). Since the global EV-D68 outbreak in 2014, the virus has mainly circulated in biennial epidemic cycles with peaks detected during even years. However, following the COVID-19 pandemic, the seasonal pattern of EV-D68 has been characterized by large yearly upsurges. Here, we describe the circulation of EV-D68 in Europe in 2023 and track its genetic evolution.

Study design

Data was compiled from members of the European Non-Polio Network (ENPEN). This included monthly data on the total number of EV samples tested, EV positive samples, EV-D68 positive samples and cases, and other EV positive samples detected in 2023. Information on sample types and surveillance system was recorded. Sequence data from the VP1 gene was used for phylogenetic and amino acid sequence analysis.

Results

EV was detected in 13585 out of 203622 diagnostic samples tested (6.7%), of which 402 (3.0%) were determined as EV-D68, representing 386 cases. EV-D68 infections peaked in October 2023 (136/386; 35.2%). 267/386 (69.2%) of EV-D68 cases were captured through clinical EV surveillance, almost all of which (202/204 of positive samples with sample type information) were detected in respiratory specimens. Phylogenetic analysis performed on 99 VP1 sequences revealed a distinct B3-derived lineage with a previously undescribed residue change, D554E, in Europe.

Conclusions

The study documents sustained circulation of EV-D68 in Europe in 2023, the evolution of B3-derived lineages, and appearance of previously undescribed amino acid substitutions in Europe. This stresses the need for continuous EV-D68 surveillance and harmonization of EV-D68 detection practices towards better data comparability across countries.

背景肠病毒D68 （EV-D68）引起从轻度到严重的呼吸系统疾病，在极少数情况下引起麻痹综合征，称为急性弛缓性脊髓炎（AFM）。自2014年全球暴发EV-D68以来，该病毒主要以两年一次的流行周期传播，在偶数年发现高峰。然而，在2019冠状病毒病大流行之后，EV-D68的季节性特征是每年大幅上升。在这里，我们描述了EV-D68在2023年欧洲的循环，并跟踪其基因进化。研究设计数据来自欧洲非脊髓灰质炎网络（ENPEN）的成员。这包括2023年检测到的EV样本总数、EV阳性样本、EV- d68阳性样本和病例以及其他EV阳性样本的月度数据。记录了样本类型和监测系统的信息。利用VP1基因序列数据进行系统发育和氨基酸序列分析。结果203622份诊断样本中检出13585份（6.7%），其中402份（3.0%）检出EV-D68，共386例。EV-D68感染在2023年10月达到高峰(136/386；35.2%)。临床监测捕获EV- d68病例中有267/386例（69.2%），其中绝大多数（202/204例有标本类型信息的阳性标本）为呼吸道标本。对99个VP1序列进行的系统发育分析显示，在欧洲，具有先前未描述的残基变化D554E的独特b3衍生谱系。该研究记录了2023年EV-D68在欧洲的持续循环，b3衍生谱系的进化以及之前未描述的氨基酸取代在欧洲的出现。这强调需要持续监测EV-D68并统一EV-D68检测做法，以提高各国数据的可比性。

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引用次数: 0

Feasibility and performance evaluations of Alinity m quantitative NAT for HBV and HCV Alinity m定量NAT检测HBV和HCV的可行性及性能评价

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-01 DOI: 10.1016/j.jcv.2025.105784

Sana Sajid, Zaira Rehman, Fouzia Naseer, Sana Faisal Raza, Javeria Aijaz

Background

Rapid and accurate testing for Hepatitis B Virus (HBV) and Hepatitis C Virus (HCV) is essential for management of these infections. Selecting the most appropriate testing methodology entails feasibility considerations of cost and throughput in addition to performance evaluations.

Objective

In this study, we determined the feasibility and analytical performance of Alinity m in comparison with Roche Cobas 6800 for both assays according to College of American Pathologists (CAP) standards, and Clinical and Laboratory Standards Institute (CLSI) guidelines.

Results

Unit costs remain relatively stable until around a monthly test volume of 1200. Below this volume, cost escalation pattern varies, such that at 60 tests/month, unit costs increase 9-fold for tests on Roche Cobas 6800, but relatively modestly (2.5-fold) for tests on Alinity m. The 24-h throughput of Alinity m, however, is lower than that of Roche Cobas 6800. The overall concordance between the Alinity m and Cobas 6800 was 97.5 % (39/40) for HBV and 87.5 % (35/40) for HCV. The reportable range was from 1.765 to 8.460 log IU/mL (58.2 to 2.8 × 10⁸ IU/mL) for HBV, and from 1.975 to 7.250 log IU/mL (94.4 to 1.7 × 10⁷ IU/mL) for HCV. The complete reportable range remained undetermined on account of non-availability of extreme viral titer samples. Both intra and inter-run precision, as evaluated by SD falling within the manufacturer-reported, were within acceptable limits. No evidence of cross-contamination was found.

Conclusion

Alinity m demonstrated acceptable performance in comparison with Roche Cobas 6800 for HBV and HCV NAT. Additionally, it demonstrated suitability for lower throughput labs in terms of unit test costs.

背景快速准确地检测乙型肝炎病毒（HBV）和丙型肝炎病毒（HCV）对这些感染的管理至关重要。选择最合适的测试方法除了性能评估之外，还需要考虑成本和吞吐量的可行性。目的根据美国病理学家学会（CAP）标准和临床与实验室标准协会（CLSI）指南，确定Alinity m与罗氏Cobas 6800两种检测方法的可行性和分析性能。结果在每月测试量1200次左右之前，成本保持相对稳定。低于此数量，成本上升模式有所不同，例如，在每月60个测试时，罗氏Cobas 6800测试的单位成本增加了9倍，但Alinity m测试的单位成本相对较低（2.5倍）。然而，Alinity m的24小时吞吐量低于罗氏Cobas 6800。Alinity m和Cobas 6800在HBV和HCV方面的总体一致性分别为97.5%（39/40）和87.5%（35/40）。HBV报告范围为1.765 ~ 8.460 log IU/mL (58.2 ~ 2.8 × 108 IU/mL)， HCV报告范围为1.975 ~ 7.250 log IU/mL （94.4 ~ 1.7 × 107 IU/mL）。由于无法获得极端病毒滴度样本，完整的报告范围仍未确定。运行内和运行间的精度，通过SD评估落在制造商报告的范围内，都在可接受的范围内。没有发现交叉污染的证据。与罗氏Cobas 6800相比，alinity m在HBV和HCV NAT方面表现出可接受的性能。此外，就单位检测成本而言，alinity m也适用于低通量实验室。

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引用次数: 0

Molecular epidemiology of Kyasanur forest disease employing ONT-NGS a field forward sequencing 利用ONT-NGS进行野地正向测序的Kyasanur森林病害分子流行病学研究

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-04-01 DOI: 10.1016/j.jcv.2025.105783

Shashi Sharma , Pooja Yadav , Paban Kumar Dash, Suman Dhankher

The future of infectious agent detection and molecular characterization lies in field-forward, on-site strategies. The lack of genomic information for recently circulating Kyasanur Forest Disease virus strains is critical. Kyasanur Forest Virus Disease virus PCR-positive samples from 2018 to 2020 were selected for sequencing. Detailed molecular phylogenetic analyses were performed. In this study, we deciphered KFDV whole genomes using the ONT-NGS technique to analyze targeted KFD surveillance from 2018–2020. This study is the first to report recently circulating KFDV strains employing a simple on-site field-forward approach for viral surveillance. Altogether, 19 KFDV genomes were sequenced, and 28 non-synonymous variants were detected in the viral strains circulating from 2018–2020 in the Shivamogga district of Karnataka state in India. The prevailing Variant was detected in more than 10 changes in 80 % of the samples in the viral envelope protein. Recently, circulating KFDV has been the predominant lineage over the past years. India reports seasonal outbreaks almost every year from the Karnataka state of the KFD. The genomic sequences deciphered here belong to the period (2018–2020) that covers the KFDV sequences as the first information. This will contribute to the development and revisiting of diagnostic and vaccine strategies.

感染因子检测和分子表征的未来在于现场前进，现场策略。缺乏最近流行的恰萨努尔森林病病毒株的基因组信息是至关重要的。选取2018 ~ 2020年恰萨努尔森林病毒病病毒pcr阳性样本进行测序。进行了详细的分子系统发育分析。在这项研究中，我们使用ONT-NGS技术破译了KFDV的全基因组，分析了2018-2020年KFD的靶向监测。这项研究首次报道了最近流行的KFDV菌株采用简单的现场现场前向方法进行病毒监测。总共对19个KFDV基因组进行了测序，并在印度卡纳塔克邦Shivamogga地区2018-2020年流行的病毒株中检测到28个非同义变体。在80%的病毒包膜蛋白样品中，在超过10个变化中检测到流行的变体。最近，在过去的几年中，流行KFDV已成为主要的谱系。印度几乎每年都报告卡纳塔克邦的季节性疫情。这里破译的基因组序列属于覆盖KFDV序列作为第一信息的时期（2018-2020）。这将有助于制定和重新审视诊断和疫苗战略。

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引用次数: 0

Nucleocapsid-directed antibody testing is unsuitable to estimate hybrid immunity against SARS-CoV-2, a longitudinal cross-border study in the Meuse-Rhine Euroregion 在欧洲默兹-莱茵地区进行的一项纵向跨境研究表明，核衣壳定向抗体检测不适合评估对SARS-CoV-2的混合免疫

IF 4 3区医学 Q2 VIROLOGY

Journal of Clinical Virology

Pub Date : 2025-03-20 DOI: 10.1016/j.jcv.2025.105780

D.A.T. Hanssen , C.J.A. van Bilsen , C.D.J. den Heijer , C. Stabourlos , C.P.B. Moonen , R. de Vries , S. Brinkhues , D. Philipssen , B.A.M. van der Zanden , N.H.T.M. Dukers-Muijrers , C.J.P.A. Hoebe , P.H.M. Savelkoul , I.H.M. van Loo

Introduction

Understanding immunity from previous natural SARS-CoV-2 infection is important for booster vaccination strategies. A longitudinal study conducted in 2021 within the Meuse-Rhine Euroregion, bordering the Netherlands, Belgium, and Germany, aimed to assess seroprevalence of spike-directed (anti-S) and nucleocapsid-directed (anti-N) antibodies to demonstrate immunity as a result of both vaccination and natural infection (hybrid immunity), and to evaluate the dynamics of the anti-N response.

Materials and methods

Questionnaires and self-finger-prick blood samples from 3110 participants were collected at two time points: weeks 22–29 (June–July, round 1) and weeks 40–45 (October–November, round 2) of 2021. Individuals with anti-S antibodies were additionally tested for anti-N antibodies.

Results

In total, 4366 samples tested positive for anti-S; n = 1291 for round 1 and n = 3075 for round 2. Of these, 10.1 % of Dutch (32/316), 3.1 % of Belgian (9/294), and 2.8 % of German participants (13/466) were anti-N positive in round 1 (p < 0.001). In round 2, this was 4.6 % (69/1510), 3.3 % (20/607), and 1.5 % (14/912), respectively (p < 0.001). In 45.1 % (23/51) of anti-N positive participants in round 1, the result reversed to negative in round 2. In 42.1 % (16/38) of anti-N positive participants with a self-reported positive PCR result, anti-N reversed to negative in round 2.

Conclusion

Variations in anti-N seroprevalence across EMR countries may reflect differences in vaccination campaign enrollment. Over 40 % of participants experienced seroreversion of anti-N within six months, indicating anti-N testing is unsuitable for diagnosing past infection or estimating hybrid immunity within a population. However, anti-N testing may be used as a proxy for increased circulation of SARS-CoV-2 in a population.

了解以前自然感染SARS-CoV-2的免疫对加强疫苗接种策略很重要。2021年在毗邻荷兰、比利时和德国的欧洲默兹-莱茵河地区进行的一项纵向研究旨在评估尖刺导向（抗s）和核衣壳导向（抗n）抗体的血清阳性率，以证明疫苗接种和自然感染（混合免疫）的免疫效果，并评估抗n反应的动态。材料与方法在2021年6 - 7月22-29周（第1轮）和10 - 11月40-45周（第2轮）两个时间点采集3110名参与者的问卷和自指刺血样本。抗s抗体的个体另外检测抗n抗体。结果共检出抗s阳性4366份；第1轮N = 1291，第2轮N = 3075。其中，10.1%的荷兰人（32/316）、3.1%的比利时人（9/294）和2.8%的德国人（13/466）在第一轮中呈抗- n阳性(p <；0.001)。在第二轮中，分别为4.6%（69/1510）、3.3%（20/607）和1.5% (14/912)(p <；0.001)。在第一轮中，45.1%（23/51）的抗n阳性参与者在第二轮中结果逆转为阴性。在42.1%（16/38）自我报告PCR阳性的抗n阳性参与者中，抗n在第2轮逆转为阴性。结论EMR国家抗n血清阳性率的差异可能反映了疫苗接种活动登记的差异。超过40%的参与者在6个月内经历了抗n血清逆转，这表明抗n检测不适合诊断过去的感染或估计人群中的混合免疫。然而，抗n检测可作为SARS-CoV-2在人群中传播增加的指标。

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引用次数: 0