Genetic engineering of vaccine manufacturing cell lines enhances poliovirus and enterovirus 71 production

Vaccine manufacturing costs and production limitations represent two fundamental challenges facing researchers, public health officials and vaccine manufacturers committed to global health solutions. To address these issues, we have investigated whether the cell lines employed by vaccine manufacturers can be engineered to enhance vaccine virus production. As a first step in a proof-of-principle study, a genome-wide RNA Interference (RNAi) screen was conducted to identify host gene modulation events that increased Sabin 2 poliovirus (PV) replication. Primary screen hits were validated in a Vero vaccine manufacturing cell line using both attenuated and wild type poliovirus strains. This approach identified multiple single and dual gene knockdown events that increased PV titers \u3e20-fold and \u3e50-fold, respectively. Top candidate genes did not affect virus antigenicity, cell viability, or cell doubling times. Moreover, CRISPR/Cas9-mediated knockout (KO) of the top three targets created stable cell substrates with improved viral vaccine strain production. Please click Additional Files below to see the full abstract

Results from the second WHO external quality assessment for the molecular detection of respiratory syncytial virus, 2019-2020

BACKGROUND: External quality assessments (EQAs) for the molecular detection of human respiratory syncytial virus (RSV) are necessary to ensure the standardisation of reliable results. The Phase II, 2019-2020 World Health Organization (WHO) RSV EQA included 28 laboratories in 26 countries. The EQA panel evaluated performance in the molecular detection and subtyping of RSV-A and RSV-B. This manuscript describes the preparation, distribution, and analysis of the 2019-2020 WHO RSV EQA. METHODS: Panel isolates underwent whole genome sequencing and in silico primer matching. The final panel included nine contemporary, one historical virus and two negative controls. The EQA panel was manufactured and distributed by the UK National External Quality Assessment Service (UK NEQAS). National laboratories used WHO reference assays developed by the United States Centers for Disease Control and Prevention, an RSV subtyping assay developed by the Victorian Infectious Diseases Reference Laboratory (Australia), or other in-house or commercial assays already in use at their laboratories. RESULTS: An in silico analysis of isolates showed a good match to assay primer/probes. The panel was distributed to 28 laboratories. Isolates were correctly identified in 98% of samples for detection and 99.6% for subtyping. CONCLUSIONS: The WHO RSV EQA 2019-2020 showed that laboratories performed at high standards. Updating the composition of RSV molecular EQAs with contemporary strains to ensure representation of circulating strains, and ensuring primer matching with EQA panel viruses, is advantageous in assessing diagnostic competencies of laboratories. Ongoing EQAs are recommended because of continued evolution of mismatches between current circulating strains and existing primer sets

Humoral and intestinal immunity induced by new schedules of bivalent oral poliovirus vaccine and one or two doses of inactivated poliovirus vaccine in Latin American infants: an open-label randomised controlled trial

© 2016 Elsevier Ltd Background Replacement of the trivalent oral poliovirus vaccine (tOPV) with bivalent types 1 and 3 oral poliovirus vaccine (bOPV) and global introduction of inactivated poliovirus vaccine (IPV) are major steps in the polio endgame strategy. In this study, we assessed humoral and intestinal immunity in Latin American infants after three doses of bOPV combined with zero, one, or two doses of IPV. Methods This open-label randomised controlled multicentre trial was part of a larger study. 6-week-old full-term infants due for their first polio vaccinations, who were healthy on physical examination, with no obvious medical conditions and no known chronic medical disorders, were enrolled from four investigational sites in Colombia, Dominican Republic, Guatemala, and Panama. The infants were randomly assigned by permuted block randomisation (through the use of a computer-generated list, block size 36) to nine groups, of which five will be discussed in this report. These fi

Inactivated poliovirus vaccine given alone or in a sequential schedule with bivalent oral poliovirus vaccine in Chilean infants: a randomised, controlled, open-label, phase 4, non-inferiority study

Artículo de publicación ISIBackground Bivalent oral poliovirus vaccine (bOPV; types 1 and 3) is expected to replace trivalent OPV (tOPV) globally by April, 2016, preceded by the introduction of at least one dose of inactivated poliovirus vaccine (IPV) in routine immunisation programmes to eliminate vaccine-associated or vaccine-derived poliomyelitis from serotype 2 poliovirus. Because data are needed on sequential IPV bOPV schedules, we assessed the immunogenicity of two different IPV bOPV schedules compared with an all-IPV schedule in infants. Methods We did a randomised, controlled, open-label, non-inferiority trial with healthy, full-term (>2.5 kg birthweight) infants aged 8 weeks (+/- 7 days) at six well-child clinics in Santiago, Chile. We used supplied lists to randomly assign infants (1:1:1) to receive three polio vaccinations (IPV by injection or bOPV as oral drops) at age 8,16, and 24 weeks in one of three sequential schedules: IPV bOPV bOPV, IPV IPV bOPV, or IPV IPV IPV. We did the randomisation with blocks of 12 stratified by study site. All analyses were done in a masked manner. Co-primary outcomes were non-inferiority of the bOPV-containing schedules compared with the all-IPV schedule for seroconversion (within a 10% margin) and antibody titres (within two-thirds log(2) titres) to poliovirus serotypes land 3 at age 28 weeks, analysed in the per-protocol population. Secondary outcomes were seroconversion and titres to serotype 2 and faecal shedding for 4 weeks after a monovalent OPV type 2 challenge at age 28 weeks. Safety analyses were done in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, number NCT01841671, and is dosed to new participants. Findings Between April 25 and August 1,2013, we assigned 570 infants to treatment: 190 to IPV bOPV bOPV, 192 to IPV IPV bOPV, and 188 to IPV IPV IPV. 564 (99%) were vaccinated and included in the intention-to-treat cohort, and 537 (94%) in the per-protocol analyses. In the IPV bOPV bOPV, IPV IPV bOPV, and IPV IPV IPV groups, respectively, the proportions of children with seroconversion to type 1 poliovirus were 166 (98.8%) of 168,95% CI 95-8-99.7; 178 (100%), 97.9-100-0; and 175 (100%), 97.9-100.0. Proportions with seroconvsion to type 3 poliovirus were 163 (98.2%) of 166,94.8-99.4; 177(100%), 97.9-100-0, and 172(98.9%) of 174,95.9-99.7. Non-inferiority was thus shown for the bOPV-containing schedules compared with the all-IPV schedule, with no significant differences between groups. In the IPV bOPV bOPV, IPV IPV bOPV, and IPV IPV IPV groups, respectively, the proportions of children with seroprotective antibody titres to type 1 poliovirus were 168 (98-8%) of 170, 95% CI 95-8-99.7; 181 (100%), 97.9-100.0; and 177 (100%), 97.9-100.0. Proportions to type 3 poliovirus were 166 (98-2%) of 169, 94.9-99.4; 180 (100%), 97-9400.0; and 174 (98.9%) of 176,96.0-99.7. Non-inferiority comparisons could not be done for this outcome because median litres for the groups receiving OPV were greater than the assay's upper limit of detection (log2 titres >10.5). The proportions of children seroconverting to type 2 poliovirus in the IPV bOPV bOPV, IPV IPV bOPV, and IPV IPV IPV groups, respectively, were 130 (77.4%) of 168, 95% CI 70.5-83.0; 169 (96.0%) of 176,92.0-98.0; and 175 (100%), 97.8-100. IPV bOPV schedules resulted in almost a 0.3 log reduction of type 2 faecal shedding compared with the IPV-only schedule. No participants died during the trial; 81 serious adverse events were reported, of which one was thought to be possibly vaccine-related (intestinal intussusception). Interpretation Seroconversion rates against polioviruses types 1 and 3 were non-inferior in sequential schedules containing IPV and bOPV, compared with an all-IPV schedule, and proportions of infants with protective antibodies were high after all three schedules. One or two doses of bOPV after IPV boosted intestinal immunity for poliovirus type 2, suggesting possible cross protection. Additionally, there was evidence of humoral priming for type 2 from one dose of IPV. Our findings could give policy makers flexibility when choosing a vaccination schedule, especially when trying to eliminate vaccine-associated and vaccine-derived poliomyelitis

Pathologic Studies of Fatal Cases in Outbreak of Hand, Foot, and Mouth Disease, Taiwan

In 1998, an outbreak of enterovirus 71-associated hand, foot, and mouth disease occurred in Taiwan. Pathologic studies of two fatal cases with similar clinical features revealed two different causative agents, emphasizing the need for postmortem examinations and modern pathologic techniques in an outbreak investigation