Live attenuated vaccines: Historical successes and current challenges

AbstractLive attenuated vaccines against human viral diseases have been amongst the most successful cost effective interventions in medical history. Smallpox was declared eradicated in 1980; poliomyelitis is nearing global eradication and measles has been controlled in most parts of the world. Vaccines function well for acute diseases such as these but chronic infections such as HIV are more challenging for reasons of both likely safety and probable efficacy. The derivation of the vaccines used has in general not been purely rational except in the sense that it has involved careful clinical trials of candidates and subsequent careful follow up in clinical use; the identification of the candidates is reviewed

High resolution identity testing of inactivated poliovirus vaccines

AbstractBackgroundDefinitive identification of poliovirus strains in vaccines is essential for quality control, particularly where multiple wild-type and Sabin strains are produced in the same facility. Sequence-based identification provides the ultimate in identity testing and would offer several advantages over serological methods.MethodsWe employed random RT-PCR and high throughput sequencing to recover full-length genome sequences from monovalent and trivalent poliovirus vaccine products at various stages of the manufacturing process.ResultsAll expected strains were detected in previously characterised products and the method permitted identification of strains comprising as little as 0.1% of sequence reads. Highly similar Mahoney and Sabin 1 strains were readily discriminated on the basis of specific variant positions. Analysis of a product known to contain incorrect strains demonstrated that the method correctly identified the contaminants.ConclusionRandom RT-PCR and shotgun sequencing provided high resolution identification of vaccine components. In addition to the recovery of full-length genome sequences, the method could also be easily adapted to the characterisation of minor variant frequencies and distinction of closely related products on the basis of distinguishing consensus and low frequency polymorphisms

Human coronavirus 229E encodes a single ORF4 protein between the spike and the envelope genes

BACKGROUND: The genome of coronaviruses contains structural and non-structural genes, including several so-called accessory genes. All group 1b coronaviruses encode a single accessory protein between the spike and envelope genes, except for human coronavirus (HCoV) 229E. The prototype virus has a split gene, encoding the putative ORF4a and ORF4b proteins. To determine whether primary HCoV-229E isolates exhibit this unusual genome organization, we analyzed the ORF4a/b region of five current clinical isolates from The Netherlands and three early isolates collected at the Common Cold Unit (CCU) in Salisbury, UK. RESULTS: All Dutch isolates were identical in the ORF4a/b region at amino acid level. All CCU isolates are only 98% identical to the Dutch isolates at the nucleotide level, but more closely related to the prototype HCoV-229E (>98%). Remarkably, our analyses revealed that the laboratory adapted, prototype HCoV-229E has a 2-nucleotide deletion in the ORF4a/b region, whereas all clinical isolates carry a single ORF, 660 nt in size, encoding a single protein of 219 amino acids, which is a homologue of the ORF3 proteins encoded by HCoV-NL63 and PEDV. CONCLUSION: Thus, the genome organization of the group 1b coronaviruses HCoV-NL63, PEDV and HCoV-229E is identical. It is possible that extensive culturing of the HCoV-229E laboratory strain resulted in truncation of ORF4. This may indicate that the protein is not essential in cell culture, but the highly conserved amino acid sequence of the ORF4 protein among clinical isolates suggests that the protein plays an important role in vivo

Novel Parvovirus and Related Variant in Human Plasma

We report a novel parvovirus (PARV4) and related variants in pooled human plasma used in the manufacture of plasma-derived medical products. Viral DNA was detected by using highly selective polymerase chain reaction assays; 5% of pools tested positive, and amounts of DNA ranged from <500 copies/mL to >106 copies/mL plasma

Isolation of Vaccine-Like Poliovirus Strains in Sewage Samples From the United Kingdom.

Background: Environmental surveillance (ES) is a sensitive method for detecting human enterovirus (HEV) circulation, and it is used worldwide to support global polio eradication. We describe a novel ES approach using next-generation sequencing (NGS) to identify HEVs in sewage samples collected in London, United Kingdom, from June 2016 to May 2017. Methods: Two different methods were used to process raw sewage specimens: a 2-phase aqueous separation system and size exclusion by filtration and centrifugation. HEVs were isolated using cell cultures and analyzed using NGS. Results: Type 1 and 3 vaccine-like poliovirus (PV) strains were detected in samples collected from September 2016 through January 2017. NGS analysis allowed us to rapidly obtain whole-genome sequences of PV and non-PV HEV strains. As many as 6 virus strains from different HEV serotypes were identified in a single cell culture flask. PV isolates contained only a small number of mutations from vaccine strains commonly seen in early isolates from vaccinees. Conclusions: Our ES setup has high sensitivity for polio and non-PV HEV detection, generating nearly whole-genome sequence information. Such ES systems provide critical information to assist the polio eradication endgame and contribute to the improvement of our understanding of HEV circulation patterns in humans

A Complete Spectroscopic Survey of the Milky Way Satellite Segue 1: The Darkest Galaxy

We present the results of a comprehensive Keck/DEIMOS spectroscopic survey of the ultra-faint Milky Way satellite galaxy Segue 1. We have obtained velocity measurements for 98.2% of the stars within 67 pc (10 arcmin, or 2.3 half-light radii) of the center of Segue 1 that have colors and magnitudes consistent with membership, down to a magnitude limit of r=21.7. Based on photometric, kinematic, and metallicity information, we identify 71 stars as probable Segue 1 members, including some as far out as 87 pc. After correcting for the influence of binary stars using repeated velocity measurements, we determine a velocity dispersion of 3.7^{+1.4}_{-1.1} km/s, with a corresponding mass within the half-light radius of 5.8^{+8.2}_{-3.1} x 10^5 Msun. The stellar kinematics of Segue 1 require very high mass-to-light ratios unless the system is far from dynamical equilibrium, even if the period distribution of unresolved binary stars is skewed toward implausibly short periods. With a total luminosity less than that of a single bright red giant and a V-band mass-to-light ratio of 3400 Msun/Lsun, Segue 1 is the darkest galaxy currently known. We critically re-examine recent claims that Segue 1 is a tidally disrupting star cluster and that kinematic samples are contaminated by the Sagittarius stream. The extremely low metallicities ([Fe/H] < -3) of two Segue 1 stars and the large metallicity spread among the members demonstrate conclusively that Segue 1 is a dwarf galaxy, and we find no evidence in favor of tidal effects. We also show that contamination by the Sagittarius stream has been overestimated. Segue 1 has the highest measured dark matter density of any known galaxy and will therefore be a prime testing ground for dark matter physics and galaxy formation on small scales.Comment: 24 pages, 4 tables, 11 figures (10 in color). Submitted for publication in ApJ. V3 revised according to comments from the refere

Development of a candidate reference material for adventitious virus detection in vaccine and biologicals manufacturing by deep sequencing.

Unbiased deep sequencing offers the potential for improved adventitious virus screening in vaccines and biotherapeutics. Successful implementation of such assays will require appropriate control materials to confirm assay performance and sensitivity. A common reference material containing 25 target viruses was produced and 16 laboratories were invited to process it using their preferred adventitious virus detection assay. Fifteen laboratories returned results, obtained using a wide range of wet-lab and informatics methods. Six of 25 target viruses were detected by all laboratories, with the remaining viruses detected by 4-14 laboratories. Six non-target viruses were detected by three or more laboratories. The study demonstrated that a wide range of methods are currently used for adventitious virus detection screening in biological products by deep sequencing and that they can yield significantly different results. This underscores the need for common reference materials to ensure satisfactory assay performance and enable comparisons between laboratories

Global urban environmental change drives adaptation in white clover

Urbanization transforms environments in ways that alter biological evolution. We examined whether urban environmental change drives parallel evolution by sampling 110,019 white clover plants from 6169 populations in 160 cities globally. Plants were assayed for a Mendelian antiherbivore defense that also affects tolerance to abiotic stressors. Urban-rural gradients were associated with the evolution of clines in defense in 47% of cities throughout the world. Variation in the strength of clines was explained by environmental changes in drought stress and vegetation cover that varied among cities. Sequencing 2074 genomes from 26 cities revealed that the evolution of urban-rural clines was best explained by adaptive evolution, but the degree of parallel adaptation varied among cities. Our results demonstrate that urbanization leads to adaptation at a global scale

Assaying the Potency of Influenza Vaccines

The potency of vaccines must be determined to ensure that the appropriate dose is given. The manufacture and assessment of influenza vaccines are complicated by the continuously changing nature of the pathogen, which makes efficacy estimates difficult but also confounds attempts to produce a well-validated, consistent potency assay. Single radial diffusion has been used for decades and provides a relatively simple way to measure the amount of biologically active materials present in the vaccine. It requires reagents, which are updated on a regular, frequently yearly, basis and alternative methods continue to be sought