To Analyze the Sensitivity of RT-PCR Assays Employing S Gene Target Failure with Whole Genome Sequencing Data during Third Wave by SARS-CoV-2 Omicron Variant

Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the community by immune evasion mechanisms. Due to mutation within S gene, most Omicron variants have reported S gene target failure (SGTF) with some commercially available PCR kits. Such diagnostic features can be used as markers to screen Omicron. However, Whole Genome Sequencing (WGS) is the only gold standard approach to confirm novel microorganisms at genetically level as similar mutations can also be found in other variants that are circulating at low frequencies worldwide. This Retrospective study is aimed to assess RT-PCR sensitivity in the detection of S gene target failure in comparison with whole genome sequencing to detect variants of Omicron. Methods: We have analysed retrospective data of SARS-CoV-2 positive RT-PCR samples for S gene target failure (SGTF) with TaqPath COVID-19 RT-PCR Combo Kit (ThermoFisher) and combined with sequencing technologies to study the emerged pattern of SARS-CoV-2 variants during third wave at the tertiary care centre, Surat. Results: From the first day of December 2021 till the end of February 2022, a total of 321,803 diagnostic RT-PCR tests for SARS-CoV-2 were performed, of which 20,566 positive cases were reported at our tertiary care centre with an average cumulative positivity of 6.39% over a period of three months. In the month of December 21 samples characterized by the SGTF (70/129) were suggestive of being infected by the Omicron variant and identified as Omicron (B.1.1.529 lineage) when sequence. In the month of January, we analysed a subset of samples (n = 618) with SGTF (24%) and without SGTF (76%) with Ct values Conclusions: During the COVID-19 pandemic, it took almost more than 15 days to diagnose infection and identify pathogen by sequencing technology. In contrast to that molecular assay provided quick identification with the help of SGTF phenomenon within 5 hours of duration. This strategy helps scientists and health policymakers for the quick isolation and identification of clusters. That ultimately results in a decreased transmission of pathogen among the community.

1起由冷链产品引起的新型冠状病毒Omicron BA.2.3.7感染的传播链和基因序列分析

目的对福建省福州市M区1起由进口冷链产品引起的新型冠状病毒暴发疫情进行溯源及基因序列分析。方法收集2022年9月6日至9月9日期间M区新冠疫情中全部感染者(8名)以及1份阳性冷链产品样本拭子,进行流行病学调查,对采集到的样本进行实时荧光定量PCR检测,对3例阳性感染者和1份冷链产品样本进行全基因组序列测序分析。结果该起疫情共有8人感染,均为无症状感染者,4条全基因组序列高度同源,共享75个的核苷酸突变位点,共享55个氨基酸突变位点,且符合谱系BA.2.3.7的特征。结论本次疫情由未有效防护的工人暴露于污染了新型冠状病毒Omicron BA.2.3.7的进口冷链产品所引起。

Neutralizing Antibody Responses against Five SARS-CoV-2 Variants and T Lymphocyte Change after Vaccine Breakthrough Infections from the SARS-CoV-2 Omicron BA.1 Variant in Tianjin, China: A Prospective Study

Objective To investigate whether Omicron BA.1 breakthrough infection after receiving the SARS-CoV-2 vaccine could create a strong immunity barrier.Methods Blood samples were collected at two different time points from 124 Omicron BA.1 breakthrough infected patients and 124 controls matched for age,gender,and vaccination profile.Live virus-neutralizing antibodies against five SARS-CoV-2 variants,including WT,Gamma,Beta,Delta,and Omicron BA.1,and T-lymphocyte lymphocyte counts in both groups were measured and statistically analyzed.Results The neutralizing antibody titers against five different variants of SARS-CoV-2 were significantly increased in the vaccinated population infected with the Omicron BA.1 variant at 3 months after infection,but mainly increased the antibody level against the WT strain,and the antibody against the Omicron strain was the lowest.The neutralizing antibody level decreased rapidly 6 months after infection.The T-lymphocyte cell counts of patients with mild and moderate disease recovered at 3 months and completely returned to the normal state at 6 months.Conclusion Omicron BA.1 breakthrough infection mainly evoked humoral immune memory in the original strain after vaccination and hardly produced neutralizing antibodies specific to Omicron BA.1.Neutralizing antibodies against the different strains declined rapidly and showed features similar to those of influenza.Thus,T-lymphocytes may play an important role in recovery.

SARS-CoV-2 Omicron变异株多重微滴式数字PCR定量方法的建立及应用

【目的】为建立精准、高效的多重微滴式数字PCR(droplet digital PCR,ddPCR)定量分析检测方法,开发可同时鉴别新型冠状病毒(severe acute respiratory syndrome coronavirus 2,SARS-CoV-2)ORF1ab基因、N基因、E基因以及Omicron变异株S基因的检测体系,提高病毒性传染疾病的诊断效率及传播风险监测能力。【方法】通过筛选保守基因序列并设计特异性引物与探针,优化反应体系和扩增程序,对该方法的特异性、灵敏度及稳定性进行评价。以临床样本为实验材料,利用建立的ddPCR方法进行检测和验证,确定阳性检出率。【结果】多重ddPCR反应体系中,各对引物探针对目的片段均能有效扩增,SARS-CoV-2 ORF1ab基因、N基因、E基因以及Omicron变异株S基因灵敏度检测下限分别为0.59、0.68、1.44、1.03 copies/μL。在20份临床样本的核酸检测中,共检出16份阳性样本,阳性率达80%(16/20),经荧光定量PCR方法复测符合率一致。【结论】研究建立的多重ddPCR方法特异性强、灵敏度高,可实现对临床样本中微量新冠病毒的精确定量检测。

PCR-HRM for Genomic Surveillance of SARS-CoV-2: A Variant Detection Tool in Côte d’Ivoire, West Africa

The rise of new viruses, like SARS-CoV-2 causing the COVID-19 outbreak, along with the return of antibiotic resistance in harmful bacteria, demands a swift and efficient reaction to safeguard the health and welfare of the global population. It is crucial to have effective measures for prevention, intervention, and monitoring in place to address these evolving and recurring risks, ensuring public health and international security. In countries with limited resources, utilizing recombinant mutation plasmid technology in conjunction with PCR-HRM could help differentiate the existence of novel variants. cDNA synthesis was carried out on 8 nasopharyngeal samples following viral RNA extraction. The P1 segment of the SARS-CoV-2 Spike S protein was amplified via conventional PCR. Subsequently, PCR products were ligated with the pGEM-T Easy vector to generate eight recombinant SARS-CoV-2 plasmids. Clones containing mutations were sequenced using Sanger sequencing and analyzed through PCR-HRM. The P1 segment of the S gene from SARS-CoV-2 was successfully amplified, resulting in 8 recombinant plasmids generated from the 231 bp fragment. PCR-HRM analysis of these recombinant plasmids differentiated three variations within the SARS-CoV-2 plasmid population, each displaying distinct melting temperatures. Sanger sequencing identified mutations A112C, G113T, A114G, G214T, and G216C on the P1 segment, validating the PCR-HRM findings of the variations. These mutations led to the detection of L452R or L452M and F486V protein mutations within the protein sequence of the Omicron variant of SARS-CoV-2. In summary, PCR-HRM is a vital and affordable tool for distinguishing SARS-CoV-2 variants utilizing recombinant plasmids as controls.

Contribution of Genomic Surveillance in the Detection and Monitoring of SARS-CoV-2 Variants during the 6 Pandemic Waves in the Central African Republic from 2020 to 2023

Objective: The COVID-19 pandemic has highlighted the need to strengthen diagnosis and genomic surveillance capacities. In 2021, Central African managed five waves of COVID-19 by integrating genomic surveillance into their health monitoring system. This study sought to report surveillance data from the National Laboratory of Clinical Biology and Public Health and describe the circulation of SARS-CoV-2 variants. Materials and Methods: This retrospective, descriptive observational study spans three years, from April 2020 to November 2023. It was conducted on a population of consenting volunteers from across the Central African Republic, who were tested using RT-PCR on nasopharyngeal samples. Data with sufficient information were obtained from the National Laboratory of Clinical Biology and Public Health (LNBCSP) databases. Sequencing was largely carried out at the National Institute of Biomedical Research (INRB) in Kinshasa until May 2023, and subsequently at the LNBCSP. Results and Discussion: Out of 97,864 RT-PCR tests performed, 9,764 were positive, resulting in a prevalence of 9.98%. The average age of the patients was 39.97 years ± 13.76, and the male-to-female sex ratio was 2.12. RT-PCR test positivity was significantly associated with age (p = 0.001), sex (p = 0.013) and clinical manifestations. Ten variants circulated during the five recorded waves, with Omicron (B.1.1.529), Delta (B.1.617.2) variants being predominant. Notably, the B.1.620 and B.640 variants were prominent during the second wave. Conclusion: This retrospective study provides key insights into the COVID-19 pandemic in the CAR. It identifies risk factors and details the circulation of various SARS-CoV-2 variants. Enhancing national genomic surveillance capacities would enable the country to better respond to future pandemic challenges.

RT-PCR/CRISPR-Cas12a快速检测和区分SARS-CoV-2 Omicron BA.4/5变异株

目的建立一种基于CRISPPR-Cas12a基因编辑技术对新型冠状病毒(SARS-CoV-2)奥密克戎BA.4/5变异株快速检测与鉴别的方法。方法结合逆转录聚合酶链式反应(RT-PCR)与CRISPR基因编辑技术,基于次优原间隔区相邻基序(suboptimal-PAM)设计奥密克戎BA.4/5变异株特异性的CRISPRRNA(crRNA),从而建立RT-PCR/CRISPR-Cas12a快速检测新冠病毒BA.4/5变异株的方法。本研究检测了43例新冠病毒阳性的临床样本(包括新冠病毒野生株以及变异株Alpha、Beta、Delta、奥密克戎BA.1和BA.4/5)以及20例新冠病毒阴性的临床样本(包括11种常见呼吸道病原体),并以测序结果为金标准计算PCR/CRISPR-Cas12a检测方法的ROC曲线下面积(AUC)以及灵敏度(SEN)、特异性(SPE)、一致性(Kappa)评价检测方法的性能。结果本研究筛选到两条特异性crRNA-1和crRNA-2,所建立的方法可在30min内快速特异地检测出SARS-CoV-2奥密克戎BA.4/5变异株,最低检测限为10copies/μL。此外,在检测感染11种常见呼吸道病原体的临床样本时均未观察到非特异性交叉反应。基于crRNA-1和crRNA-2的检测结果的灵敏度分别为97.83%、100%;特异性均为100%;ROC曲线下面积分别为0.9989和1.0;与Sanger测序方法的一致性分别为92.83%、96.41%。结论本研究采用CRISPPR-Cas12a基因编辑技术与逆转录聚合酶链式反应相结合,成功建立了一种快速检测与鉴别SARS-CoV-2奥密克戎BA.4/5变异株的新方法。其特异性强、灵敏度高、稳定性好,可用于新冠病毒奥密克戎BA.4/5变异株的批量检测与分型,可用于常规监测和跟踪SARS-CoV-2变异的传播。

Chest computed tomography findings of the Omicron variants of SARS-CoV-2 with different cycle threshold values

BACKGROUND The Omicron variant of severe acute respiratory syndrome coronavirus 2(SARSCoV-2)mainly infects the upper respiratory tract.This study aimed to determine whether the probability of pulmonary infection and the cycle threshold(Ct)measured using the fluorescent polymerase chain reaction(PCR)method were related to pulmonary infections diagnosed via computed tomography(CT).AIM To analyze the chest CT signs of SARS-CoV-2 Omicron variant infections with different Ct values,as determined via PCR.METHODS The chest CT images and PCR Ct values of 331 patients with SARS-CoV-2Omicron variant infections were retrospectively collected and categorized into low(<25),medium(25.00-34.99),and high(≥35)Ct groups.The characteristics of chest CT images in each group were statistically analyzed.RESULTS The PCR Ct values ranged from 13.36 to 39.81,with 99 patients in the low,155 in the medium,and 77 in the high Ct groups.Six abnormal chest CT signs were detected,namely,focal infection,patchy consolidation shadows,patchy groundglass shadows,mixed consolidation ground-glass shadows,subpleural interstitial changes,and pleural changes.Focal infections were less frequent in the low Ct group than in the medium and high Ct groups;these infections were the most common sign in the medium and high Ct groups.Patchy consolidation shadows and pleural changes were more frequent in the low Ct group than in the other two groups.The number of patients with two or more signs was greater in the low Ct group than in the medium and high Ct groups.CONCLUSION The chest CT signs of patients with pulmonary infection caused by the Omicron variants of SARSCoV-2 varied depending on the Ct values.Identification of the characteristics of Omicron variant infection can help subsequent planning of clinical treatment.

Surveillance of emerging SARS-CoV-2 variants by nanopore technology-based genome sequencing

Objective:To surveill emerging variants by nanopore technology-based genome sequencing in different COVID-19 waves in Sri Lanka and to examine the association with the sample characteristics,and vaccination status.Methods:The study analyzed 207 RNA positive swab samples received to sequence laboratory during different waves.The N gene cut-off threshold of less than 30 was considered as the major inclusion criteria.Viral RNA was extracted,and elutes were subjected to nanopore sequencing.All the sequencing data were uploaded in the publicly accessible database,GISAID.Results:The Omicron,Delta and Alpha variants accounted for 58%,22%and 4%of the variants throughout the period.Less than 1%were Kappa variant and 16%of the study samples remained unassigned.Omicron variant was circulated among all age groups and in all the provinces.Ct value and variants assigned percentage was 100%in Ct values of 10-15 while only 45%assigned Ct value over 25.Conclusions:The present study examined the emergence,prevalence,and distribution of SARS-CoV-2 variants locally and has shown that nanopore technology-based genome sequencing enables whole genome sequencing in a low resource setting country.

Comparison of the replication and neutralization of different SARS-CoV-2 Omicron subvariants in vitro

Background:New Omicron subvariants are emerging rapidly from BA.1 to BA.4 and BA.5.Their pathogenicity has changed from that of wild-type(WH-09)and Omicron variants have over time become globally dominant.The spike proteins of BA.4 and BA.5 that serve as the target for vaccine-induced neutralizing antibodies have also changed compared to the previous subvariants,which is likely to cause immune es-cape and the reduction of the protective effect of the vaccine.Our study addresses the above issues and provides a basis for formulating relevant prevention and control strategies.Methods:We collected cellular supernatant and cell lysates and measured the viral titers,viral RNA loads,and E subgenomic RNA(E sgRNA)loads in different Omicron subvariants grown in Vero E6 cells,using WH-09 and Delta variants as a reference.Additionally,we evaluated the in vitro neutralizing activity of different Omicron sub-variants and compared it to the WH-09 and Delta variants using macaque sera with different types of immunity.Results:As the SARS-CoV-2 evolved into Omicron BA.1,the replication ability in vitro began to decrease.Then with the emergence of new subvariants,the replication ability gradually recovered and became stable in the BA.4 and BA.5 subvariants.In WH-09-inactivated vaccine sera,geometric mean titers of neutralization antibodies against different Omicron subvariants declined by 3.7~15.4-fold compared to those against WH-09.In Delta-inactivated vaccine sera,geometric mean titers of neutrali-zation antibodies against Omicron subvariants declined by 3.1~7.4-fold compared to those against Delta.Conclusion:According to the findings of this research,the replication efficiency of all Omicron subvariants declined compared with WH-09 and Delta variants,and was lower in BA.1 than in other Omicron subvariants.After two doses of inactivated(WH-09 or Delta)vaccine,cross-neutralizing activities against various Omicron subvariants were seen despite a decline in neutralizing titers.

上海市某方舱医院2897例新型冠状病毒Omicron变异株轻型/无症状感染者住院时间影响因素分析

目的探讨上海市某方舱医院新型冠状病毒奥密克戎(Omicron)变异株轻型/无症状感染者住院时间的影响因素。方法回顾性收集上海宝山泾灿路(罗泾京东)方舱医院收治的2897例Omicron变异株轻型/无症状感染者的病历资料,分析其一般资料、住院时间等基本情况,探究不同特征的新冠病毒Omicron变异株感染者住院时间差异,采用多重线性回归方法分析住院时间的影响因素。根据单因素分析结果,采用二元logistic回归分析住院时间≥14 d可能的影响因素。结果2897例Omicron变异株轻型/无症状感染者的平均住院时间为(9.1±4.6)d,不同年龄段、入舱前核酸阳性时间以及伴有咳嗽、咳痰、咽痛、发热、流涕、鼻塞、口干咽燥、头痛、乏力、肌痛、腹泻、恶寒、恶心呕吐、头晕症状的感染者住院时间不同,差异具有统计学意义(P<0.05)。多重线性回归结果显示,年龄增加、患有糖尿病史以及咽痛、发热、流涕、肌痛症状会增加住院天数,而入舱前核酸阳性时间长则会减少住院天数。住院时间≥14 d者,高龄、糖尿病史以及出现咽痛、发热、口干咽燥、肌痛、恶心呕吐症状的占比更高,入舱前核酸阳性时间相对更短,接种2针及以上疫苗占比更低,差异具有统计学意义(P<0.05)。二元Logistic回归结果显示,高龄、入舱前核酸阳性时间短、糖尿病史以及发热和口干咽燥症状可能是导致住院时间超过14 d的危险因素,接种2针及以上疫苗可能是保护因素。结论Omicron变异株轻型/无症状感染者平均住院时间约为9 d,高龄、疫苗接种2针及以上、患有糖尿病史、入舱前核酸阳性时间短以及咽痛、发热、流涕、口干咽燥、肌痛等正气虚衰、病邪入里、化热伤阴表现对于住院时间可能产生重要影响。通过尽早识别核酸转阴慢、住院时间长的高危患者,以期为缩短核酸转阴时间、指导重点人群精准防控提供指导方向。

新型冠状病毒Omicron I1566V突变位点MS2噬菌体病毒样颗粒的构建

目的构建新型冠状病毒Omicron I1566V突变位点MS2噬菌体病毒样颗粒。方法选取并合成带有6×His标签的MS2噬菌体的衣壳蛋白(CP)、成熟蛋白(A蛋白)及Omicron I1566V突变基因序列,插入pACYCDuet-1质粒构建重组载体,通过原核系统诱导表达目的蛋白,纯化重组蛋白后利用透射电镜对蛋白质进行物理表征,最后通过反转录聚合酶链反应(RT-PCR)检测病毒样颗粒的热稳定性及耐核酸酶水解能力。结果成功构建包含有6×His标签的CP、A蛋白和Omicron I1566V突变基因序列的重组载体,经限制性内切酶BamHⅠ和KpnⅠ酶切鉴定和测序验证,结果均与预期相符。经诱导并纯化后,通过电镜观察到了大小均匀、直径为23~28 nm的病毒样颗粒,该病毒样颗粒经核酸酶消化后可在37℃条件下稳定储存20 d以上。结论该研究成功利用MS2噬菌体的CP和A蛋白构建了Omicron I1566V突变位点病毒样颗粒,为该突变位点的RT-PCR检测体系提供了可靠的质量保障。

Symptomatic COVID-19 in University Students: A School-Wide Web-Based Questionnaire Survey during the Omicron Variant Outbreak

Aim: To detect risk and preventive factors associated with the Omicron variant infection in university students, a combination of a web-based survey and multivariate logistic regression analysis was introduced as the front-line initiatives by the school health practitioners. Design: Questionnaire survey. Methods: The school-wide web-based questionnaire survey was conducted among our university students as a part of the annual health check-up in April, 2023. The positive outcome was confined to the first symptomatic COVID-19 onset during the Omicron variant outbreak. Results: In this self-administered survey, risk or protective associations were merely estimated statistically in university students (n = 5406). In measured factors, karaoke and club/group activities could maintain the statistical significance in adjusted odds ratios (ORs) as relative risk factors, and science course, measles/ rubella (MR) vaccination, and COVID-19 vaccination remained as relative protective factors in adjusted OR analyses. Club/group activities with member gathering and karaoke sing-along sessions in university students may frequently have WHO’s three Cs. These risk factors are still important topics for the infection control of COVID-19 in university students. Together with some recent reports from other researchers, the significant protective role of MR vaccine in our survey warrants further clinical investigation. If the breakthrough infection continuously constitutes the majority of infection, real data in test-negative case-control or web-based questionnaire design continue to be important for statistical analysis to determine the minimal requirement of our strategies which may be equivalent to or replace COVID-19 vaccines.

Genomic Profile of SARS-COV-2 Associated with COVID-19 Outbreaks in N’Djamena, Chad

Background: SARS-CoV-2 has circulated worldwide with dramatic consequences. In Chad, we have no data reported of variants. The aim of this study was to identify the SARS-CoV-2 variants that circulated during the epidemic from 2020 to 2021. Methods: This is a cross-sectional, descriptive study carried out between 2020 and 2021. Samples from patients with suspected COVID-19 were tested in five laboratories in N’Djamena. One hundred quality samples of the positives were sequenced in Kinshasa using Oxford nanopore technologies minion and the Protocol Midnight SARS-CoV2. Data were processed using Excel version 16 software. Results: Of the 100 samples sequenced, 77 (77%) produced sequences, 23 (23%) did not. The genomic profiles were wild-type Wuhan and minor mutations (19A, 19B (A), 20A (B.1, B.2), 20B (AV.1), 20D (B.1.1.1 /C.36), 20C), variant of concern Alpha (20I), variant of concern Delta (21A/J), variant of interest Eta (21D), variant of concern Omicron (21K) and unclassified variant under surveillance (B.1.640). Of these variants, the maximums were detected in patients aged 26 - 35 with 30.26% and 25.26% in 36 - 45. However, 24.67% were in travelers and 75.32% in residents, 35.06% in those vaccinated against COVID-19 and 62.33% in non-vaccinates. The estimated case-fatality rate was 2.44% (107/4374). Conclusion: This work has provided preliminary data on COVID-19 and SARS-CoV-2 variants circulating during the 2020-2021 epidemics in Chad.

Accurate Diagnosis of SARS-CoV-2 JN.1 by Sanger Sequencing of Receptor-Binding Domain Is Needed for Clinical Evaluation of Its Immune Evasion

Background: Omicron JN.1 has become the dominant SARS-CoV-2 variant in recent months. JN.1 has the highest number of amino acid mutations in its receptor binding domain (RBD) and has acquired a hallmark L455S mutation. The immune evasion capability of JN.1 is a subject of scientific investigation. The US CDC used SGTF of TaqPath COVID-19 Combo Kit RT-qPCR as proxy indicator of JN.1 infections for evaluation of the effectiveness of updated monovalent XBB.1.5 COVID-19 vaccines against JN.1 and recommended that all persons aged ≥ 6 months should receive an updated COVID-19 vaccine dose. Objective: Recommend Sanger sequencing instead of proxy indicator to diagnose JN.1 infections to generate the data based on which guidelines are made to direct vaccination policies. Methods: The RNA in nasopharyngeal swab specimens from patients with clinical respiratory infection was subjected to nested RT-PCR, targeting a 398-base segment of the N-gene and a 445-base segment of the RBD of SARS-CoV-2 for amplification. The nested PCR amplicons were sequenced. The DNA sequences were analyzed for amino acid mutations. Results: The N-gene sequence showed R203K, G204R and Q229K, the 3 mutations associated with Omicron BA.2.86 (+JN.1). The RBD sequence showed 24 of the 26 known amino acid mutations, including the hallmark L455S mutation for JN.1 and the V483del for BA.2.86 lineage. Conclusions: Sanger sequencing of a 445-base segment of the SARS-CoV-2 RBD is useful for accurate determination of emerging variants. The CDC may consider using Sanger sequencing of the RBD to diagnose JN.1 infections for statistical analysis in making vaccination policies.

SARS-CoV-2与Omicron变异株感染者血常规和生化指标的比较

目的分析新型冠状病毒SARS-CoV-2与Omicron变异株感染者血常规及生化指标的差异。方法收集44例SARS-CoV-2感染者(SARS-CoV-2组)以及49例Omicron变异株感染者(Omicron组)的临床资料,比较两组患者的首次血常规及生化指标结果。结果Omicron组患者的白细胞计数、中性粒细胞计数、中性粒细胞百分比、钾离子、钠离子、氯离子、尿素、总蛋白和白蛋白水平均高于SARS-CoV-2组(P<0.05或0.01),而淋巴细胞百分比、球蛋白水平以及钾离子、钠离子的异常率低于SARS-CoV-2感染组(P<0.05)。两组患者的淋巴细胞计数、尿素、肌酐、总胆红素等比较差异无统计学意义(P>0.05)。结论Omicron感染者早期炎症和部分生化指标变化较SARS-CoV-2患者差异显著。

Global challenge with the SARS-CoV-2 omicron BA.2(B.1.1.529.2)subvariant:Should we be concerned?

BA.2 is a novel omicron offshoot of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)that has gone viral.There is limited knowledge regarding this variant of concern.Current evidence suggests that this variant is more contagious but less severe than previous SARS-CoV-2 variants.However,there is concern regarding the virus mutations that could influence pathogenicity,transmissibility,and immune evasion.

Omicron variant (B.1.1.529) of SARS-CoV-2: Mutation, infectivity, transmission, and vaccine resistance

The appearance of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)variant Omicron(B.1.1.529)has caused panic responses around the world because of its high transmission rate and number of mutations.This review summarizes the highly mutated regions,the essential infectivity,transmission,vaccine breakthrough and antibody resistance of the Omicron variant of SARSCoV-2.The Omicron is highly transmissible and is spreading faster than any previous variant,but may cause less severe symptoms than previous variants.The Omicron is able to escape the immune system’s defenses and coronavirus disease 2019 vaccines are less effective against the Omicron variant.Early careful preventive steps including vaccination will always be key for the suppression of the Omicron variant.

Clinical outcomes of the omicron variant compared with previous SARS-CoV-2 variants;meta-analysis of current reports

BACKGROUND Omicron(B.1.1.529)is a severe acute respiratory syndrome coronavirus 2(SARSCoV-2)variant of concern;however,there is no comprehensive analysis regarding clinical features,disease severity,or clinical outcomes of this variant.AIM To compare the clinical characteristics of infection with omicron and previous variants of SARS-CoV-2.METHODS We searched major international databases consisting ISI Web of Science,PubMed,Scopus,MedRxiv,and Reference Citation Analysis to collect the potential relevant documents.Finally,clinical features,e.g.,death rate,intensive care unit(ICU)admission,length of hospitalization,and mechanical ventilation,of infection with SARS-CoV-2 omicron variant compared with previous variants were assessed using odds ratio and 95%confidence intervals by Comprehensive Meta-Analysis software version 2.2.RESULTS A total of 12 articles met our criteria.These investigated the clinical outcomes of infection with omicron variant compared with other variants such as alpha,beta and delta.Our results suggested that ICU admission,need for mechanical ventilation,and death rate were significantly lower for omicron than previous variants.In addition,the average length of hospitalization during the omicron wave was significantly shorter than for other variants.CONCLUSION The infectivity of omicron variant was higher than for previous variants due to several mutations,particularly in the spike protein.However,disease severity was mild to moderate compared previous variants.

Sanger Sequencing for Molecular Diagnosis of SARS-CoV-2 Omicron Subvariants and Its Challenges

Large population passages of the SARS-CoV-2 in the past two and a half years have allowed the circulating virus to accumulate an increasing number of mutations in its genome. The most recently emerging Omicron subvariants have the highest number of mutations in the Spike (S) protein gene and these mutations mainly occur in the receptor-binding domain (RBD) and the N-terminal domain (NTD) of the S gene. The European Centre for Disease Prevention and Control (eCDC) and the World Health Organization (WHO) recommend partial Sanger sequencing of the SARS-CoV-2 S gene RBD and NTD on the polymerase chain reaction (PCR)-positive samples in diagnostic laboratories as a practical means of determining the variants of concern to monitor possible increased transmissibility, increased virulence, or reduced effectiveness of vaccines against them. The author’s diagnostic laboratory has implemented the eCDC/WHO recommendation by sequencing a 398-base segment of the N gene for the definitive detection of SARS-CoV-2 in clinical samples, and sequencing a 445-base segment of the RBD and a 490 - 509-base segment of the NTD for variant determination. This paper presents 5 selective cases to illustrate the challenges of using Sanger sequencing to diagnose Omicron subvariants when the samples harbor a high level of co-existing minor subvariant sequences with multi-allelic single nucleotide polymorphisms (SNPs) or possible recombinant Omicron subvariants containing a BA.2 RBD and an atypical BA.1 NTD, which can only be detected by using specially designed PCR primers. In addition, Sanger sequencing may reveal unclassified subvariants, such as BA.4/BA.5 with L84I mutation in the S gene NTD. The current large-scale surveillance programs using next-generation sequencing (NGS) do not face similar problems because NGS focuses on deriving consensus sequence.