Virology Journal

Foot-and-mouth disease (FMD) is a highly contagious viral disease that affects cloven-hoofed animals. Vaccination is the most effective measure to control FMD. However, FMDV particles are prone to dissociation, leading to insufficient potency of vaccine. Based on this characteristic, a combination of twenty percentage trehalose, 500 mM NaCl and 3 mM CuSO4·5H2O was developed to increase viral stability. Heating-resistance test showed that FMDV infectivity was maintained when formulated with formulation. Additionally, the half-life of FMDV inactivation was prolonged remarkably. Sequencing analysis demonstrated that viral genome could not be altered in serial passages. Vaccine stability was monitored for up to 1 year at 4 °C, with a higher level of 146S content remained. This study suggested that the formulation could protect FMDV against massive structural breakdown and extend the shelf life of vaccine. Our findings could provide strategy to develop more solutions for the stabilization of viral vaccine.

Các virus parechovirus (PeV-As), thuộc một giống mới trong họ Picornaviridae, đã được liên kết với nhiều vụ bùng phát bệnh nghiêm trọng tại địa phương, chẳng hạn như sổ mũi, viêm phổi, phát ban dạng sẩn và viêm kết mạc. Tuy nhiên, theo như chúng tôi biết, chỉ có một số ít phòng thí nghiệm toàn cầu tiến hành các xét nghiệm để xác định nhóm virus này. Do đó, trong nghiên cứu này, chúng tôi nhằm phát triển và xác thực một phương pháp RT-PCR thời gian thực để nhận diện PeV-As. Để thiết kế và xác thực một mồi – đầu dò PCR thời gian thực nhắm vào vùng 5′-UTR của PeV-As, chúng tôi đã căn chỉnh các trình tự 5′-UTR của PeV-As có sẵn trong GenBank bằng cách sử dụng thuật toán MUSCLE trong MEGA v7.0. Sau đó, vùng 5′-UTR được bảo tồn cao đã được chọn và trình tự mồi – đầu dò của nó được thiết kế bằng cách sử dụng Primer Premier v5.0. Trình tự mồi – đầu dò này sau đó được đánh giá về độ đặc hiệu, tính nhạy và độ lặp lại, và để xác thực, nó được thử nghiệm trên các mẫu phân của 728 trẻ em khỏe mạnh sống tại Bắc Kinh (Trung Quốc). Phương pháp RT-PCR thời gian thực PeV-A chỉ phát hiện các tiêu chuẩn dương tính RNA của các kiểu gene PeV-A (1-8, 14, 17 và 18), trong khi 72 serotype của các virus EV không phải PeV-A không được phát hiện. Thêm vào đó, vùng VP1 của 11 kiểu gene PeV-A này đã thử nghiệm dương tính được khuếch đại bằng các mồi được thiết kế trong nghiên cứu này. Kết quả phân loại cho thấy tám, một và hai chủng trong số 11 là PeV-A1, PeV-A4 và PeV-A6, tương ứng. Chúng tôi cũng đã xác định và trình bày các kết quả đặc trưng di truyền và phân tích hệ gen tương ứng với các trình tự vùng VP1 này. Hơn nữa, phương pháp RT-PCR thời gian thực cho thấy độ nhạy tốt với LOD là 102 bản sao/μL. Kết quả dương tính trong tám thử nghiệm song song ở mỗi gradient nồng độ từ 107 bản sao/μL đến 102 bản sao/μL, cho thấy độ lặp lại tốt. Những phát hiện của chúng tôi cho thấy phương pháp RT-PCR thời gian thực phát triển trong nghiên cứu này có thể được áp dụng để nhận diện PeV-A trong các trường hợp thường xuyên. Chúng tôi đã phát hiện các kiểu gene PeV-A1, 4 và 6 trong 728 mẫu phân bằng phương pháp này. Ngoài ra, chúng tôi tin rằng kết quả của chúng tôi sẽ tạo nền tảng cho các nghiên cứu tiếp theo về PeV-As và tạo điều kiện mở rộng thông tin trình tự gene có sẵn trong GenBank.

The editors of Virology Journal would like to thank all our reviewers who have contributed to the journal in Volume 9 (2012). The success of any scientific journal depends on an effective and strict peer review process and Virology Journal could not operate without your contribution. We look forward to your continuous support to this journal either as an invited reviewer or a contributing author in the years to come.

Oncolytic viruses are being studied and developed as novel cancer treatments. Using directed evolution technology, structural modification of the viral surface protein increases the specificity of the oncolytic virus for a particular cancer cell. Newcastle disease virus (NDV) does not show specificity for certain types of cancer cells during infection; therefore, it has low cancer cell specificity. Hemagglutinin is an NDV receptor-binding protein on the cell surface that determines host cell tropism. NDV selectivity for specific cancer cells can be increased by artificial amino acid changes in hemagglutinin neuraminidase HN proteins via directed evolution, leading to improved therapeutic effects. Sialic acid-binding sites (H domains) of the HN protein mutant library were generated using error-prone PCR. Variants of the H domain protein were screened by enzyme-linked immunosorbent assay using HCT 116 cancer cell surface molecules. The mutant S519G H domain protein showed the highest affinity for the surface protein of HCT 116 cells compared to that of different types of cancer cells. This showed that the S519G mutant H domain protein gene replaced the same part of the original HN protein gene, and S519G mutant recombinant NDV (rNDV) was constructed and recovered. S519G rNDV cancer cell killing effects were tested using the MTT assay with various cancer cell types, and the tumor suppression effect of the S519G mutant rNDV was tested in a xenograft mouse model implanted with cancer cells, including HCT 116 cells. S519G rNDV showed increased specificity and enhanced killing ability of HCT 116 cells among various cancer cells and a stronger suppressive effect on tumor growth than the original recombinant NDV. Directed evolution using an artificial amino acid change in the NDV HN (S519G mutant) protein increased its specificity and oncolytic effect in colorectal cancer without changing its virulence. These results provide a new methodology for the use of directed evolution technology for more effective oncolytic virus development.

Human papillomaviruses (HPVs) remain a serious world health problem due to their association with anogenital/oral cancers and warts. While over 100 HPV types have been identified, a subset is associated with malignancy. HPV16 and 18 are the most prevalent oncogenic types, while HPV6 and 11 are most commonly responsible for anogenital warts. While other quantitative PCR (qPCR) assays detect oncogenic HPV, there is no single tube assay distinguishing the most frequent oncogenic types and the most common types found in warts. A Sybr Green-based qPCR assay was developed utilizing degenerate primers to the highly conserved HPV E1 theoretically detecting any HPV type. A single tube multiplex qPCR assay was also developed using type-specific primer pairs and TaqMan probes that allowed for detection and quantitation of HPV6,11,16,18. Each HPV type was detected over a range from 2 × 101 to 2 × 106copies/reaction providing a reliable method of quantitating type-specific HPV in 140 anogenital/cutaneous/oral benign and malignant specimens. 35 oncogenic and low risk alpha genus HPV types were detected. Concordance was detected in previously typed specimens. Comparisons to the gold standard detected an overall sensitivity of 89% (95% CI: 77% - 96%) and specificity of 90% (95%CI: 52% - 98%). There was good agreement between the ability of the qPCR assays described here to identify HPV types in malignancies previously typed using standard methods. These novel qPCR assays will allow rapid detection and quantitation of HPVs to assess their role in viral pathogenesis.

Human papillomavirus (HPV) vaccines based on major capsid protein L1 are licensed in over 100 countries to prevent HPV infections. The yeast-derived recombinant quadrivalent HPV L1 vaccine, GARDASIL(R), has played an important role in reducing cancer and genital warts since its introduction in 2006. The L1 proteins self-assemble into virus-like particles (VLPs). VLPs were subjected to post-purification disassembly and reassembly (D/R) treatment during bioprocessing to improve VLP immunoreactivity and stability. The post-D/R HPV16 VLPs and their complex with H16.V5 neutralizing antibody Fab fragments were visualized by cryo electron microscopy, showing VLPs densely decorated with antibody. Along with structural improvements, post-D/R VLPs showed markedly higher antigenicity to conformational and neutralizing monoclonal antibodies (mAbs) H16.V5, H16.E70 and H263.A2, whereas binding to mAbs recognizing linear epitopes (H16.J4, H16.O7, and H16.H5) was greatly reduced. Strikingly, post-D/R VLPs showed no detectable binding to H16.H5, indicating that the H16.H5 epitope is not accessible in fully assembled VLPs. An atomic homology model of the entire HPV16 VLP was generated based on previously determined high-resolution structures of bovine papillomavirus and HPV16 L1 pentameric capsomeres. D/R treatment of HPV16 L1 VLPs produces more homogeneous VLPs with more virion-like antibody reactivity. These effects can be attributed to a combination of more complete and regular assembly of the VLPs, better folding of L1, reduced non-specific disulfide-mediated aggregation and increased stability of the VLPs. Markedly different antigenicity of HPV16 VLPs was observed upon D/R treatment with a panel of monoclonal antibodies targeting neutralization sensitive epitopes. Multiple epitope-specific assays with a panel of mAbs with different properties and epitopes are required to gain a better understanding of the immunochemical properties of VLPs and to correlate the observed changes at the molecular level. Mapping of known antibody epitopes to the homology model explains the changes in antibody reactivity upon D/R. In particular, the H16.H5 epitope is partially occluded by intercapsomeric interactions involving the L1 C-terminal arm. The homology model allows a more precise mapping of antibody epitopes. This work provides a better understanding of VLPs in current vaccines and could guide the design of improved vaccines or therapeutics.

The editors of Virology Journal would like to thank all our reviewers who have contributed to the journal in Volume 10 (2013). The success of any scientific journal depends on an effective and strict peer review process and Virology Journal could not operate without your contribution. We are grateful to the large number of reviewers (1026 to be exact!), who have done a great job in not only lifting the quality of the journal’s scientific peer reviewing process, but also helped us to achieve our goal of a median time to first decision of just 35 days. Our record time from submission to online, open access, publication in 2013 was 22 days for a Research Article [1] and 28 days for a Review [2]. This is a great achievement by any standard. We look forward to your continuous support of Virology Journal either as an invited reviewer or a contributing author in the years to come.

Containment of influenza A H1N1 virus spread was implemented successfully in Xiamen, with large-scale inoculation to reduce morbidity. To identify beneficial elements and to guide decision-making in epidemic containment, we analyzed the epidemiologic parameters and evaluated the control measures. We determined various parameters from laboratory-confirmed cases, including incubation period, duration of illness and reproductive number (R0), and evaluated the control measures. There were1414 cases with dates of onset between June 14, 2009 and March 22, 2010. The incidence was 56.79/100,000, and mortality was 0.12/100,000. The incidence during the community epidemic phase was 6.23 times higher than in the containment phase. A total of 296,888 subjects were inoculated with domestic influenza H1N1 virus cleavage vaccine. An epidemic curve showed that vaccination in students cut the peak incidence of illness significantly. Men (relative risk (RR) = 1.30, 95% confidence interval (CI): 1.17-1.45) and persons aged 0-14 years were at greater risk of infection. The incidence increased with younger age (χ2 = 950.675, p = ∞). Morbidity was lower in urban than in rural areas (RR = 0.56, 95%CI: 0.50-0.62). The median incubation time was 2 days, median duration of symptoms was 7 days, and the within-school reproductive number was 1.35. Our analysis indicated that the characteristics of this novel influenza virus were similar to those of seasonal influenza. The principle of "interception of imported cases" applied at Xiamen ports, and vaccination of students effectively limited the spread of the influenza pandemic and reduced the epidemic peak.

Hepatitis C virus (HCV) is highly infectious pathogen which is responsible for causing Hepatitis around 200 million individuals worldwide. In Pakistan, 4.7% of HCV prevalence has been reported and HCV genotype 3a has been found to be the major source of infection in Pakistan but still there is lack of information on distribution of HCV genotypes and viral load in various geographical regions of Pakistan. Therefore, current study was designed to determine distribution of HCV genotypes as well viral load in different areas of Punjab province of Pakistan. A total of 995 serum samples were taken from those individuals in which antibodies against HCV were detected through ELISA, from different regions of Punjab i.e. Lahore 317(31.85%), Faisalabad 70(7.03%), Gujranwala 129(12.96%), Gujrat 106(10.65%), Sialkot 94(9.44%), Sargodha 60(6.03%), Mandibaha-ud-din 135(13.56%), Jhang 86(8.64%). Qualitative PCR was performed to determine viral load and genotyping was performed using Nested PCR. Chi-square test was used to determine the age and sex-wise prevalence of HCV. Out of 995 samples, 888 samples were found positive for HCV RNA. In all regions, genotype 3a showed highest prevalence (82.81%) followed by genotype 1 (3.41%), mixed genotypes (2.41%), genotype 2 (0.50%), genotype 5 (0.1%) and unclassified genotypes (10.75%). Viral load in 29.5% patients infected with genotype 3a was less than 600,000 IU/mL, while it was between 600,000-800,000 IU/mL in 27.9% patients and 25.22% patients had more than 800,000 IU/mL viral load. HCV genotype 3a is the most prevalent genotype in various regions of Punjab. Viral load of HCV patients in these different regions of Punjab are reported for the first time. Moreover, based upon these results the Patients having viral load below 800,000 IU/mL would be expected to show better response of anti-HCV therapy.