Journal of virological methods最新文献_第6页

Rapid and sensitive SYBR Green-based RT-qPCR assays for the detection and quantification of mink coronavirus. 基于SYBR绿色的水貂冠状病毒快速、灵敏的RT-qPCR检测与定量

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-03-09 DOI: 10.1016/j.jviromet.2026.115383

Ruiling Niu, Xinrui Wang, Qiushu Wang, Shuyi Liu, Jingxuan Cui, Michael J Carr, Juan Li, Weifeng Shi, Cixiu Li

Mink coronavirus (MCoV) infection poses a serious threat to animal health, resulting in substantial economic losses in mink farming and impeding the sustainable development of the fur industry. Thus, developing both sensitive and specific molecular diagnostics is imperative for effective MCoV surveillance and disease control. In the present study, specific oligonucleotide primers were designed to target the 1b and N genes of MCoV, and two SYBR Green-based RT-qPCR assays - designated as MCoV-1b and MCoV-N, respectively - were established. The assays exhibited extremely high sensitivity, with lower limits of detection of 1.79 × 10 ¹ copies/μL for MCoV-1b and 1.29 × 10 ¹ copies/μL for MCoV-N. No cross-reactivity was observed with other common viruses and melting curve analyses revealed a single sharp peak for each assay, confirming specific amplification of MCoV targets. Reproducibility across three independent experimental runs exhibited low standard deviations and coefficients of variation (CV) of cycle threshold (Ct) values across broad dynamic ranges, indicating stable and consistent performance (CV < 5%). When applied to faecal clinical samples (n = 236), the assays yielded overall positivity rates of 77.97% for the 1b target and 77.54% for the N targets, which are consistent with that from high-throughput sequencing, validating their utility. Collectively, the two RT-qPCR assays developed in this study offer a reliable technical platform for rapid screening, accurate quantification, and epidemiological monitoring of MCoV. The reported molecular diagnostics hold significant practical value in mitigating the risks of MCoV transmission, supporting the sustainable growth of the global mink farming industry and diminishing potential global public health threats.

水貂冠状病毒（MCoV）感染对动物健康构成严重威胁，给水貂养殖业造成重大经济损失，阻碍毛皮产业的可持续发展。因此，开发敏感和特异的分子诊断方法对于有效监测和控制MCoV至关重要。本研究设计了针对MCoV 1b和N基因的特异性寡核苷酸引物，建立了两种基于SYBR green的RT-qPCR检测方法，分别命名为MCoV-1b和MCoV-N。该方法灵敏度极高，MCoV-1b和MCoV-N的检测下限分别为1.79×10¹copies/μL和1.29×10¹copies/μL。未观察到与其他常见病毒的交叉反应性，熔化曲线分析显示每次检测都有一个尖峰，证实了MCoV靶标的特异性扩增。在三个独立的实验运行中，再现性表现出低标准偏差和周期阈值（Ct）值的变异系数在很宽的动态范围内，表明性能稳定一致（CV < 5%）。当应用于粪便临床样本（n=236）时，检测结果显示1b靶点的总阳性率为77.97%，n靶点的总阳性率为77.54%，这与高通量测序结果一致，验证了其实用性。总的来说，本研究开发的两种RT-qPCR检测方法为MCoV的快速筛选、准确定量和流行病学监测提供了可靠的技术平台。报告的分子诊断在减轻MCoV传播风险、支持全球水貂养殖业的可持续增长和减少潜在的全球公共卫生威胁方面具有重要的实用价值。

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引用次数: 0

A split GFP approach to assay SARS-CoV-2 spike-dependent cell fusion 分裂GFP法检测SARS-CoV-2刺突依赖性细胞融合

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-01 DOI: 10.1016/j.jviromet.2025.115296

M. Jane Morwitzer, Ying Yi Zheng, Heather Friberg, Jeffrey R. Currier

The SARS-CoV-2 spike (S) protein plays a central role in viral entry through receptor binding and membrane fusion, making it a key target for therapeutic interventions. While existing assays for studying spike-mediated fusion can be complex and lack real-time monitoring, we present a split GFP-based cell fusion assay that provides a straightforward and adaptable platform for evaluating fusion dynamics. This assay utilizes a split GFP system in non-adherent 293-F cells to detect fusion events, allowing for reliable assessment of spike-targeting monoclonal antibodies and fusion inhibitors. Our study demonstrates the effectiveness of this approach in evaluating the inhibitory potential of therapeutics against multiple SARS-CoV-2 variants. The results highlight the assay’s ability to distinguish variant-specific fusion characteristics and inhibitor responses, particularly in the context of Omicron’s altered entry pathways. By enabling the quantitative, real-time assessment of spike-mediated fusion, this platform provides a valuable tool for therapeutic screening, supporting future efforts to develop antiviral strategies against SARS-CoV-2 and other emerging coronaviruses.

SARS-CoV-2刺突蛋白(S)在病毒通过受体结合和膜融合进入病毒中起着核心作用，使其成为治疗干预的关键靶点。虽然现有的研究尖峰介导的融合的方法可能很复杂，缺乏实时监测，但我们提出了一种基于gfp的分裂细胞融合实验，为评估融合动力学提供了一个简单且适应性强的平台。该试验利用非贴壁293-F细胞中的分裂GFP系统来检测融合事件，从而可以可靠地评估尖峰靶向单克隆抗体和融合抑制剂。我们的研究证明了这种方法在评估治疗方法对多种SARS-CoV-2变体的抑制潜力方面的有效性。结果强调了该检测方法区分变异特异性融合特征和抑制剂反应的能力，特别是在Omicron改变进入途径的背景下。通过对刺突介导的融合进行定量、实时评估，该平台为治疗筛选提供了有价值的工具，支持未来开发针对SARS-CoV-2和其他新兴冠状病毒的抗病毒策略。

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引用次数: 0

Optimization of rabies virus production in neuroblastoma cells using quality by design principles for vaccine and diagnostic applications 基于疫苗和诊断应用设计原则的神经母细胞瘤细胞狂犬病病毒生产质量优化

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-10-16 DOI: 10.1016/j.jviromet.2025.115289

Nayara Ugeda , Orlando Garcia Ribeiro , Elaine Raniero Fernandes , Fernanda Guedes , Sandriana Dos Ramos Silva , Iana Suly Santos Katz

Efficient production of high-titer rabies virus (RABV) stocks is critical for vaccine development, diagnostic assay design, and virological research. This study employed a Quality by Design (QbD) framework to optimize RABV propagation in Neuro-2a (N2a) cells, targeting infectious titers ≥ 8.0 log₁₀ FFU/mL and cell viability ≥ 85 %. Three genetically distinct RABV strains were evaluated: the laboratory-adapted CVS-11 strain for vaccine production and two field isolates, MN-18 and V-M, relevant to diagnostics and research. A 2 ³ full factorial Design of Experiments (DOE) was implemented to assess the impact of multiplicity of infection (MOI), incubation temperature, and culture medium composition on critical quality attributes (CQAs). Distinct strain-specific responses were observed: CVS-11 and V-M were significantly influenced by MOI (p < 0.05), whereas MN-18 demonstrated a more stable replication profile. Enriched medium significantly enhanced viral titers and cell viability across all strains (p < 0.001). Monte Carlo simulations (n = 10,000) delineated an optimal operational space—MOI 0.1–1.0, temperature 34–35°C, enriched medium—ensuring process robustness with 95 % confidence. This QbD-guided strategy provides a scalable, reproducible platform for generating high-quality RABV stocks, facilitating the manufacture of CVS-11-based vaccines and supporting the diagnostic use of field isolates as biologically standardized reference reagents for diagnostics.

高效生产高滴度狂犬病毒（RABV）库存对于疫苗开发、诊断试验设计和病毒学研究至关重要。本研究采用质量设计（QbD）框架优化RABV在神经-2a （N2a）细胞中的繁殖，目标是感染滴度≥8.0log₁₀FFU/mL，细胞存活率≥85%。评估了三种遗传上不同的RABV毒株：用于疫苗生产的实验室适应的CVS-11毒株和与诊断和研究相关的两种现场分离株MN-18和V-M。采用2³全因子实验设计（DOE）来评估感染次数（MOI）、孵育温度和培养基组成对关键质量属性（cqa）的影响。观察到不同的菌株特异性反应：CVS-11和V-M受MOI显著影响（p < 0.05），而MN-18表现出更稳定的复制谱。强化培养基显著提高了所有菌株的病毒滴度和细胞活力（p < 0.001）。蒙特卡罗模拟（n = 10,000）描绘了最佳操作空间- moi 0.1-1.0，温度34-35°C，富集介质-确保过程稳健性，置信度为95%。这种qbd指导的战略提供了一个可扩展的、可重复的平台，用于生成高质量的RABV库存，促进基于cvs -11的疫苗的生产，并支持将现场分离株作为诊断的生物标准化参考试剂进行诊断。

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引用次数: 0

Metagenomic surveillance of emerging viruses in mosquito populations from high-risk regions of Iran 伊朗高危地区蚊群中新发病毒的宏基因组监测

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-17 DOI: 10.1016/j.jviromet.2025.115301

Ebrahim Abbasi

Background

Mosquito-borne arboviruses pose a growing threat to public health, particularly in ecologically vulnerable and climatically dynamic regions. This study aimed to investigate the diversity of emerging arboviruses in mosquito populations from high-risk provinces in southern and southeastern Iran using a metagenomic surveillance approach.

Methods

Adult mosquitoes were collected from 36 sites across Hormozgan, Sistan and Baluchestan, and Khuzestan provinces. Specimens were pooled by species and location, followed by RNA extraction and high-throughput sequencing. Bioinformatics analysis was performed to identify viral taxa and assess phylogenetic relationships.

Results

A total of 4275 mosquitoes representing six species were analyzed. Virome analysis revealed 43 viral taxa, including medically important arboviruses such as dengue virus serotype 2 (DENV-2), chikungunya virus (CHIKV), and West Nile virus (WNV). Multiple novel viral sequences were also detected, including putative members of Phenuiviridae and Orthomyxoviridae. Viral diversity was highest in Hormozgan province and positively correlated with ambient temperature.

Conclusion

This study provides the first comprehensive metagenomic insight into mosquito viromes in Iran, revealing both endemic and potentially novel arboviruses. These findings underscore the need for integrated genomic surveillance and regional vector-borne disease preparedness.

背景：蚊媒虫媒病毒对公众健康构成日益严重的威胁，特别是在生态脆弱和气候动态地区。本研究旨在利用宏基因组监测方法调查伊朗南部和东南部高风险省份蚊子种群中新发虫媒病毒的多样性。方法：在霍尔木兹甘省、锡斯坦省、俾路支斯坦省和胡齐斯坦省36个地点采集成蚊。标本按物种和地点汇总，然后进行RNA提取和高通量测序。进行生物信息学分析以确定病毒分类群并评估系统发育关系。结果：共捕获蚊虫6种4275只。病毒组分析揭示了43个病毒分类群，包括医学上重要的虫媒病毒，如2型登革热病毒（DENV-2）、基孔肯雅病毒（CHIKV）和西尼罗河病毒（WNV）。此外，还检测到多个新的病毒序列，包括疑似的苯病毒科和正粘病毒科成员。病毒多样性在霍尔木兹甘省最高，且与环境温度呈正相关。结论：本研究首次对伊朗蚊子病毒群进行了全面的宏基因组研究，揭示了地方性和潜在的新型虫媒病毒。这些发现强调了综合基因组监测和区域媒介传播疾病防范的必要性。

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引用次数: 0

A novel methodology for assessing contact tracing precision: Phylogenetic validation of a contact tracing program for COVID-19 in Belgium 评估接触者追踪精度的新方法：比利时COVID-19接触者追踪计划的系统发育验证。

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-10-14 DOI: 10.1016/j.jviromet.2025.115279

Jonathan Thibaut , Fabiana Gámbaro , Caspar Geenen , Samuel L. Hong , Joren Raymenants , Sarah Gorissen , Lize Cuypers , Piet Maes , Guy Baele , Simon Dellicour , Emmanuel André

During the COVID-19 pandemic, contact tracing was widely used to limit virus propagation and implement targeted disease control measures. It can however be difficult to assess whether infected cases are actually linked to the traced index case. In this study, we developed and applied an analytical pipeline using genomic data to assess the precision of contact tracing, defined as the proportion of transmission events suggested by contact tracing that are not contradicted by genomic analysis. We exemplify our approach by examining the transmission of SARS-CoV-2 among students at Belgium’s largest university, in Leuven, during the Omicron BA.1 and BA.2 epidemic waves. We analysed 459 case-contact pairs identified through contact tracing. We then aimed to determine whether the pairs, where patients were infected with the same variant, clustered together within a time-scaled phylogeny. Our findings indicate that 34.6 % of transmission events suggested by contact tracing were not invalidated by our combined phylogenetic and single nucleotide polymorphism analysis. Only considering non invalidated transmission events, we estimated serial intervals with a smaller standard deviation than with all case-contact pairs. Our genomic-based pipeline allows us to assess the ability of our contact tracing program to correctly identify transmission chains. While contact tracing is crucial for early outbreak detection and control, monitoring its precision is vital for using this data in public health decisions.

在2019冠状病毒病大流行期间，接触者追踪被广泛用于限制病毒传播和实施有针对性的疾病控制措施。然而，很难评估感染病例是否确实与追踪到的指示病例有关。在这项研究中，我们开发并应用了一种使用基因组数据的分析管道来评估接触者追踪的精度，定义为接触者追踪所提示的与基因组分析不相矛盾的传播事件的比例。我们通过研究在Omicron BA.1和BA.2流行波期间，比利时最大的大学鲁汶大学的学生中SARS-CoV-2的传播来举例说明我们的方法。我们分析了通过接触者追踪确定的459对病例-接触者。然后，我们的目标是确定患者感染相同变体的对是否在一个时间尺度的系统发育中聚集在一起。我们的研究结果表明，通过系统发育和单核苷酸多态性联合分析，34.6%的接触者追踪提示的传播事件没有被证实。我们基于基因组学的途径使我们能够评估接触者追踪项目正确识别传播链的能力。虽然接触者追踪对早期发现和控制疫情至关重要，但监测其准确性对于在公共卫生决策中使用这些数据至关重要。

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引用次数: 0

Development of a dual-label time-resolved fluorescence immunoassay platform for simultaneous detection of canine distemper virus and parvovirus 同时检测犬瘟热病毒和细小病毒的双标记时间分辨荧光免疫分析平台的建立。

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-19 DOI: 10.1016/j.jviromet.2025.115302

Laiqing Li , Hongrui Lai , Huankun Liang , Guiling Guo , Cuicui Chen , Qiang Jia

Objective

Canine distemper virus (CDV) and canine parvovirus type 2 (CPV-2) are two highly contagious and severely pathogenic viruses commonly found in dogs, particularly lethal to puppies. This study aimed to establish a double-labeling time-resolved fluorescence immunoassay (TRFIA) to test and distinguish CDV and CPV-2 infection.

Methods

A sandwich TRFIA method was established and optimized using europium(III) (Eu^3 +)/samarium(III) (Sm³⁺) chelates dual labeling. Subsequently, it was formulated into a kit and constructed the TRFIA platform, and both its laboratory performance (limit of detection, specificity, accuracy and stability) and its ability to detect target viruses in canine clinical samples were evaluated.

Results

A dual-label TRFIA platform for simultaneous CDV and CPV-2 detection was constructed and rigorously validated. The TRFIA platform exhibited limit of detection of 0.43 ng/mL for CDV and 0.73 ng/mL for CPV-2, with high specificity for both targets. Clinical cut-offs were established at 5.47 ng/mL (CDV) and 6.96 ng/mL (CPV-2). Intra-assay coefficients of variation ranged from 1.11 % to 5.53 %, with spike-recoveries between 104.74 % and 108.50 %. Concordance with PCR was excellent (Pearson’s χ² test, P < 0.001). Clinical validation yielded diagnostic sensitivities/specificities of 90.24 %/97.37 % for CDV and 88.37 %/98.65 % for CPV-2, respectively.

Conclusion

A TRFIA platform for simultaneous detection of CDV and CPV-2 demonstrated robust limit of detection, specificity, accuracy, plus reliable clinical sensitivity and specificity. It offers an additional tool for distinguishing CDV from CPV-2 infections and may enhance future disease-prevention strategies.

目的：犬瘟热病毒（Canine犬瘟热病毒，CDV）和犬细小病毒2型（Canine parvovirus type 2, CPV-2）是犬类中常见的两种高传染性、高致病性病毒，对幼犬具有致命性。本研究旨在建立一种双标记时间分辨荧光免疫分析法（TRFIA）来检测和区分CDV和CPV-2感染。方法：采用铕（III） (Eu3+)/钐（III）（Sm3+）螯合物双标记，建立并优化夹层TRFIA方法。随后，将其配制成试剂盒，构建TRFIA平台，并对其实验室性能（检出限、特异性、准确性和稳定性）以及犬临床样本中检测目标病毒的能力进行评价。结果：建立了同时检测CDV和CPV-2的双标签TRFIA平台，并进行了严格的验证。TRFIA平台对CDV和CPV-2的检出限分别为0.43ng/mL和0.73ng/mL，特异性均较高。临床临界值分别为5.47ng/mL （CDV）和6.96ng/mL （CPV-2）。测定内变异系数为1.11% ~ 5.53%，加样回收率为104.74% ~ 108.50%。与PCR的一致性极好（Pearson χ 2检验，P < 0.001）。临床验证结果显示，CDV和CPV-2的诊断敏感性和特异性分别为90.24%/97.37%和88.37%/98.65%。结论：TRFIA平台同时检测CDV和CPV-2具有较强的检出限、特异性、准确性，临床敏感性和特异性可靠。它为区分CDV和CPV-2感染提供了一种额外的工具，并可能加强未来的疾病预防策略。

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引用次数: 0

Development of a LAMP-based diagnostic method for HTLV-1 using Iranian clinical samples 利用伊朗临床样本建立基于lamp的HTLV-1诊断方法

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-16 DOI: 10.1016/j.jviromet.2025.115303

Yousef Douzandegan , Ali Kargar Kheirabad , Sayed‑Hamidreza Mozhgani , Gholamreza Tavoosidana , Abbas Rahimi Foroushani , Mehdi Norouzi

Background

Human T-lymphotropic virus type 1 (HTLV-1) is a delta-retrovirus responsible for severe diseases such as adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Accurate and accessible diagnostic tools are required, especially in endemic regions such as Iran, to manage reasonably and prevent HTLV-1 infections.

Objective

This study aimed to develop and validate an HTLV-1 detection protocol using loop-mediated isothermal amplification (LAMP) method on clinical samples from Iranian patients.

Methods

A LAMP assay was designed using optimized primers targeting the HTLV-1 genome. It was validated with clinical samples, and its sensitivity and specificity were evaluated compared to the polymerase chain reaction (PCR) method. Reaction conditions were optimized for fast and specific amplification, and results were visually analyzed utilizing a Luminator device and a real-time thermocycler.

Results

The LAMP assay revealed a sensitivity of 100 % and specificity of 94.7 % for HTLV-1 detection. Within 15–30 min, the method produced results in isothermal conditions with good performance. The fluorescence-based detection approach is easy to use. It does not need a sophisticated laboratory facility and has a limit of detection of 2.5 copies/µL.

Conclusion

This study demonstrates the feasibility of a LAMP-based method for HTLV-1 detection that is rapid, cost-effective, and accessible compared to PCR. For this reason, this alternative could enhance HTLV-1 diagnosis in both qualitative or quantitative form in endemic and low-resource scenarios. Future research should explore further validation in more diverse populations and integration with new technologies for digital detection.

背景：人类嗜t淋巴病毒1型（HTLV-1）是一种delta逆转录病毒，可导致成人t细胞白血病/淋巴瘤（ATL）和HTLV-1相关脊髓病/热带痉挛性截瘫（HAM/TSP）等严重疾病。需要准确和可获得的诊断工具，特别是在伊朗等流行地区，以便合理管理和预防HTLV-1感染。目的：本研究旨在建立并验证环介导等温扩增（LAMP）方法对伊朗患者临床样品的HTLV-1检测方案。方法：采用优化后的引物，设计针对HTLV-1基因组的LAMP检测方法。用临床样本进行验证，并与聚合酶链反应（PCR）法比较其敏感性和特异性。优化反应条件以实现快速特异扩增，并利用Luminator装置和实时热循环仪对扩增结果进行可视化分析。结果：LAMP法检测HTLV-1的灵敏度为100%，特异性为94.7%。该方法在等温条件下15-30min内生成结果，性能良好。基于荧光的检测方法易于使用。它不需要复杂的实验室设施，检测限为2.5拷贝/µL。结论：本研究证明了基于lamp的HTLV-1检测方法的可行性，与PCR相比，该方法快速，经济，易于获取。因此，在流行和资源匮乏的情况下，这种替代方法可以提高HTLV-1的定性或定量诊断。未来的研究应该探索在更多样化的人群中进一步验证，并与数字检测的新技术相结合。

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引用次数: 0

Development of monoclonal antibody based SPCE logistic regression model to predict the protective status of animals vaccinated against FMD virus type A 建立基于单克隆抗体的SPCE logistic回归模型预测动物接种口蹄疫A型病毒的保护状况。

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-07 DOI: 10.1016/j.jviromet.2025.115299

Rajamanickam Hema Sayee , Madhusudan Hosamani , Narayanan Krishnaswamy , Subramaniyan Shanmuganathan , Manchikanti Sri Sai Charan , Dasu Ramadoss Vignesh , Veerakyathappa Bhanuprakash

In enzootic regions like India, foot-and-mouth disease (FMD) is managed through vaccination and strict biosecurity. Traditionally, FMD vaccine potency is assessed via in vivo challenge, which raises animal welfare concerns. As an alternative, this study evaluated a serology-based indirect potency test using 87 serum samples derived from calves 28 days post-vaccination. Samples were analyzed via virus neutralization test (VNT) and SPCEs employing polyclonal (pAb) and monoclonal antibodies (mAbs 2C4G11 and 6E8D11). Log₁₀ serum titres (ti) were correlated with protection outcomes from live virus challenge (FMDV serotype A/40/2000) using logistic regression. Four logit models were developed, and ROC analysis determined ti cut-offs for 50 %, 75 %, and 95 % protection probabilities (pi). VNT and mAb SPCEs showed sigmoid logit curves, indicating strong ti–pi correlation. Based on AIC and Somers’ D, mAb 2C4G11 SPCE was the best-fit model. At 75 % pi, ti thresholds for VNT, pAb, mAb 2C4G11, and mAb 6E8D11 were > 1.204, > 1.041, > 1.204, and > 1.204, respectively, with mAb 6E8D11 showing highest accuracy. Thus, mAbs 2C4G11 and 6E8D11 SPCEs are promising tools for predicting homologous protection. Further validation with larger sample sizes is recommended.

在印度等动物地方病流行地区，通过接种疫苗和严格的生物安全措施来控制口蹄疫。传统上，口蹄疫疫苗效力是通过体内挑战来评估的，这引起了对动物福利的关注。作为替代方案，本研究评估了基于血清学的间接效力测试，使用了接种后28天小牛的87份血清样本。采用多克隆抗体（pAb）和单克隆抗体（mAbs 2C4G11和6E8D11）对样品进行病毒中和试验（VNT）和spce分析。log₁0血清滴度（ti）与活病毒攻击（FMDV血清型A/40/2000）的保护结果相关。建立了四个logit模型，ROC分析确定了50%、75%和95%保护概率（pi）的截止值。VNT和mAb spce呈s形logit曲线，具有较强的ti-pi相关性。基于AIC和Somers' D， mAb 2C4G11 SPCE是最适合的模型。在75%的pi值下，VNT、pAb、mAb 2C4G11和mAb 6E8D11的阈值分别为>.204、>1.041、>1.204和>1.204，其中mAb 6E8D11的准确度最高。因此，单抗2C4G11和6E8D11 spce是预测同源保护的有效工具。建议使用更大的样本量进行进一步验证。

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引用次数: 0

Development and evaluation of a molecular assay for the detection and quantification of SARS-CoV-2, influenza A, influenza B, and respiratory syncytial virus from municipal wastewater in British Columbia 不列颠哥伦比亚省城市污水中SARS-CoV-2、甲型流感、乙型流感和呼吸道合胞病毒分子检测方法的建立和评价

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-05 DOI: 10.1016/j.jviromet.2025.115297

Jennifer Kopetzky , Sarah C. Mansour , Rachel Floyd , Liam Byrne , Christine Tchao , Natalie Prystajecky

The ability to monitor the prevalence of respiratory viruses in a community depends on testing infrastructure, surveillance programs, and community engagement. Wastewater-based surveillance has recently evolved as a tool to monitor disease at a population level with reduced costs, diverse surveillance coverage, and inherently passive population participation. This study designed a molecular assay for the detection and quantification of multiple respiratory viruses—SARS-CoV-2, influenza A, influenza B, and Respiratory Syncytial Virus (RSV)—from wastewater samples. The presence of inhibitory substances in the wastewater matrix and multiple molecular targets did not prove to be limiting factors for the performance of the assay. Evaluation of the assay using municipal wastewater treatment plant samples showed a strong concordance with publicly available surveillance and clinical case data in British Columbia, Canada. Existing and future community testing programs for respiratory viruses should integrate wastewater-based surveillance to capture asymptomatic/non-testing populations, expand surveillance coverage, and provide greater insight into the community dynamics of respiratory viruses.

监测社区中呼吸道病毒流行的能力取决于检测基础设施、监测规划和社区参与。基于废水的监测最近已发展成为一种在人口层面监测疾病的工具，其成本较低，监测覆盖范围多样化，而且人口参与本身就是被动的。本研究设计了一种用于从废水样品中检测和定量多种呼吸道病毒（sars - cov -2、甲型流感、乙型流感和呼吸道合胞病毒（RSV））的分子分析方法。废水基质中存在的抑制物质和多个分子靶标并未被证明是测定性能的限制因素。使用城市污水处理厂样本进行的分析评估显示，与加拿大不列颠哥伦比亚省公开获得的监测和临床病例数据高度一致。现有和未来的社区呼吸道病毒检测规划应整合基于废水的监测，以捕获无症状/未检测人群，扩大监测覆盖范围，并更深入地了解呼吸道病毒的社区动态。

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引用次数: 0

Development of monoclonal antibodies against goose astrovirus 2 ORF2 protein and establishment of an indirect competitive ELISA detection method 鹅星状病毒2 ORF2蛋白单克隆抗体的制备及间接竞争ELISA检测方法的建立。

IF 1.6 4区医学 Q3 BIOCHEMICAL RESEARCH METHODS

Journal of virological methods

Pub Date : 2026-02-01 Epub Date: 2025-11-19 DOI: 10.1016/j.jviromet.2025.115306

Anping Wang, Zhi Wu, Qingkang Zhou, Xiaolu Zhang, Li Liu, Shanyuan Zhu

Gosling gout, caused by goose astrovirus 2 (GAstV-2), poses a significant threat to the goose industry due to its high morbidity and mortality, while effective control measures remain unavailable. The development of specific countermeasures is hindered by a limited characterization of monoclonal antibodies (mAbs) and epitopes against the major ORF2 antigen. To address this, we generated two mAbs (A12G1 and A10D12) against GAstV-2 ORF2 using hybridoma technology. Epitope mapping identified two novel linear B-cell epitopes, ₁MADRA₅ (A12G1) and ₃₆YKPQKLPMKA₄₅ (A10D12), residing in the conserved S domain. These epitopes were highly conserved among GAstV-2 isolates but demonstrated notable divergence from other avian astroviruses. The specific reactivity of both mAbs with GAstV-2 was confirmed by Western blotting, immunofluorescence, and immunohistochemistry. Furthermore, leveraging mAb A10D12, we established a highly sensitive and specific indirect competitive ELISA (icELISA) for detecting GAstV-2 antibodies in goose serum. The icELISA demonstrated excellent reproducibility, high sensitivity, and no cross-reactivity with antisera against other common waterfowl pathogens. This study not only expands the known epitope repertoire of GAstV-2 ORF2 but also provides valuable mAbs and a robust serological tool for functional studies of ORF2 and for monitoring GAstV-2 infections.

由鹅星状病毒2型（GAstV-2）引起的小鹅痛风因其高发病率和死亡率对鹅业构成重大威胁，而目前尚无有效的控制措施。针对主要ORF2抗原的单克隆抗体（mab）和表位的有限表征阻碍了特异性对策的发展。为了解决这个问题，我们使用杂交瘤技术生成了两个针对GAstV-2 ORF2的单克隆抗体（A12G1和A10D12）。表位定位鉴定出两个新的线性b细胞表位，1MADRA5 （A12G1）和36YKPQKLPMKA45 (A10D12)，位于保守的S结构域。这些表位在GAstV-2分离株中高度保守，但与其他禽星状病毒表现出显著的差异。两种单抗与GAstV-2的特异性反应性均通过Western blotting、免疫荧光和免疫组织化学证实。此外，利用mAb A10D12，我们建立了一种高灵敏度和特异性的间接竞争ELISA （icELISA）检测鹅血清中GAstV-2抗体。该酶联免疫吸附试验重现性好，灵敏度高，与抗血清无交叉反应性。这项研究不仅扩大了已知的GAstV-2 ORF2表位库，而且为ORF2的功能研究和监测GAstV-2感染提供了有价值的单克隆抗体和强大的血清学工具。

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引用次数: 0