Effect of time at temperature on wild poliovirus titers in stool specimens

AbstractBackgroundThe effect of transport temperature on the viability of poliovirus in stool specimens from paralyzed cases has not been tested. Quality assurance of programmatic indicators will be necessary in the final phase of polio eradication.ObjectiveTo estimate the effect of time at elevated temperatures on wild poliovirus titers in stool specimens.MethodsWe exposed aliquots of pooled wild poliovirus type 1 specimens to elevated temperatures (27°C, 31°C, and 35°C) for varying time periods up to 14 days. We determined the virus titer of these aliquots and created decay curves at each temperature to estimate the relationship between time at temperature and virus titer.ResultsWe found significantly different slopes of decay at each temperature. The negative slopes increased as the temperature increased.ConclusionsWhile poliovirus in stool remains relatively stable at moderately elevated temperature, transport at higher temperatures could impact sample integrity and virus isolation results

Cold Chain and Virus‐Free Chloroplast‐Made Booster Vaccine to Confer Immunity Against Different Poliovirus Serotypes

The WHO recommends complete withdrawal of oral polio vaccine (OPV) type 2 by April 2016 globally and replacing with at least one dose of inactivated poliovirus vaccine (IPV). However, high‐cost, limited supply of IPV, persistent circulating vaccine‐derived polioviruses transmission and need for subsequent boosters remain unresolved. To meet this critical need, a novel strategy of a low‐cost cold chain‐free plant‐made viral protein 1 (VP1) subunit oral booster vaccine after single IPV dose is reported. Codon optimization of the VP1 gene enhanced expression by 50‐fold in chloroplasts. Oral boosting of VP1 expressed in plant cells with plant‐derived adjuvants after single priming with IPV significantly increased VP1‐IgG1 and VP1‐IgA titres when compared to lower IgG1 or negligible IgA titres with IPV injections. IgA plays a pivotal role in polio eradication because of its transmission through contaminated water or sewer systems. Neutralizing antibody titres (~3.17–10.17 log2 titre) and seropositivity (70–90%) against all three poliovirus Sabin serotypes were observed with two doses of IPV and plant‐cell oral boosters but single dose of IPV resulted in poor neutralization. Lyophilized plant cells expressing VP1 stored at ambient temperature maintained efficacy and preserved antigen folding/assembly indefinitely, thereby eliminating cold chain currently required for all vaccines. Replacement of OPV with this booster vaccine and the next steps in clinical translation of FDA‐approved antigens and adjuvants are discussed

The Coxsackievirus B 3Cpro Protease Cleaves MAVS and TRIF to Attenuate Host Type I Interferon and Apoptotic Signaling

The host innate immune response to viral infections often involves the activation of parallel pattern recognition receptor (PRR) pathways that converge on the induction of type I interferons (IFNs). Several viruses have evolved sophisticated mechanisms to attenuate antiviral host signaling by directly interfering with the activation and/or downstream signaling events associated with PRR signal propagation. Here we show that the 3Cpro cysteine protease of coxsackievirus B3 (CVB3) cleaves the innate immune adaptor molecules mitochondrial antiviral signaling protein (MAVS) and Toll/IL-1 receptor domain-containing adaptor inducing interferon-beta (TRIF) as a mechanism to escape host immunity. We found that MAVS and TRIF were cleaved in CVB3-infected cells in culture. CVB3-induced cleavage of MAVS and TRIF required the cysteine protease activity of 3Cpro, occurred at specific sites and within specialized domains of each molecule, and inhibited both the type I IFN and apoptotic signaling downstream of these adaptors. 3Cpro-mediated MAVS cleavage occurred within its proline-rich region, led to its relocalization from the mitochondrial membrane, and ablated its downstream signaling. We further show that 3Cpro cleaves both the N- and C-terminal domains of TRIF and localizes with TRIF to signalosome complexes within the cytoplasm. Taken together, these data show that CVB3 has evolved a mechanism to suppress host antiviral signal propagation by directly cleaving two key adaptor molecules associated with innate immune recognition

Evaluation of vaccine derived poliovirus type 2 outbreak response options: A randomized controlled trial, Karachi, Pakistan

Background: Outbreaks of circulating vaccine derived polioviruses type 2 (cVDPV2) remain a risk to poliovirus eradication in an era without live poliovirus vaccine containing type 2 in routine immunization. We evaluated existing outbreak response strategies recommended by the World Health Organization (WHO) for control of cVDPV2 outbreaks.Methods: Seronegative children for poliovirus type 2 (PV2) at 22 weeks of life were assigned to one of four study groups and received respectively (1) one dose of trivalent oral poliovirus vaccine (tOPV); (2) monovalent OPV 2 (mOPV2); (3) tOPV together with a dose of inactivated poliovirus vaccine (IPV); or (4) mOPV2 with monovalent high-potency IPV type 2. Stool and blood samples were collected and assessed for presence of PV2 (stool) and anti-polio antibodies (sera).Results: We analyzed data from 265 children seronegative for PV2. Seroconversion to PV2 was achieved in 48, 76, 98 and 100% in Groups 1–4 respectively. mOPV2 was more immunogenic than tOPV alone (p \u3c 0.001); and OPV in combination with IPV was more immunogenic than OPV alone (p \u3c 0.001). There were 33%, 67%, 20% and 43% PV2 excretors in Groups 1–4 respectively. mOPV2 resulted in more prevalent shedding of PV2 than when tOPV was used (p \u3c 0.001); and tOPV together with IPV resulted in lower excretion of PV2 than tOPV alone (p = 0.046).Conclusion: mOPV2 was a more potent vaccine than tOPV. Adding IPV to OPV improved immunological response; adding IPV also seemed to have shortened the duration of PV2 shedding. mIPV2 did not provide measurable improvement of immune response when compared to conventional IPV. WHO recommendation to use mOPV2 as a vaccine of first choice in cVDPV2 outbreak response was supported by our findings

A search-based geographic metadata curation pipeline to refine sequencing institution information and support public health

BackgroundThe National Center for Biotechnology Information (NCBI) Sequence Read Archive (SRA) has amassed a vast reservoir of genetic data since its inception in 2007. These public data hold immense potential for supporting pathogen surveillance and control. However, the lack of standardized metadata and inconsistent submission practices in SRA may impede the data’s utility in public health.MethodsTo address this issue, we introduce the Search-based Geographic Metadata Curation (SGMC) pipeline. SGMC utilized Python and web scraping to extract geographic data of sequencing institutions from NCBI SRA in the Cloud and its website. It then harnessed ChatGPT to refine the sequencing institution and location assignments. To illustrate the pipeline’s utility, we examined the geographic distribution of the sequencing institutions and their countries relevant to polio eradication and categorized them.ResultsSGMC successfully identified 7,649 sequencing institutions and their global locations from a random selection of 2,321,044 SRA accessions. These institutions were distributed across 97 countries, with strong representation in the United States, the United Kingdom and China. However, there was a lack of data from African, Central Asian, and Central American countries, indicating potential disparities in sequencing capabilities. Comparison with manually curated data for U.S. institutions reveals SGMC’s accuracy rates of 94.8% for institutions, 93.1% for countries, and 74.5% for geographic coordinates.ConclusionSGMC may represent a novel approach using a generative AI model to enhance geographic data (country and institution assignments) for large numbers of samples within SRA datasets. This information can be utilized to bolster public health endeavors

Antibodies to Enteroviruses in Cerebrospinal Fluid of Patients with Acute Flaccid Myelitis.

Acute flaccid myelitis (AFM) has caused motor paralysis in >560 children in the United States since 2014. The temporal association of enterovirus (EV) outbreaks with increases in AFM cases and reports of fever, respiratory, or gastrointestinal illness prior to AFM in >90% of cases suggest a role for infectious agents. Cerebrospinal fluid (CSF) from 14 AFM and 5 non-AFM patients with central nervous system (CNS) diseases in 2018 were investigated by viral-capture high-throughput sequencing (VirCapSeq-VERT system). These CSF and serum samples, as well as multiple controls, were tested for antibodies to human EVs using peptide microarrays. EV RNA was confirmed in CSF from only 1 adult AFM case and 1 non-AFM case. In contrast, antibodies to EV peptides were present in CSF of 11 of 14 AFM patients (79%), significantly higher than controls, including non-AFM patients (1/5 [20%]), children with Kawasaki disease (0/10), and adults with non-AFM CNS diseases (2/11 [18%]) (P = 0.023, 0.0001, and 0.0028, respectively). Six of 14 CSF samples (43%) and 8 of 11 sera (73%) from AFM patients were immunoreactive to an EV-D68-specific peptide, whereas the three control groups were not immunoreactive in either CSF (0/5, 0/10, and 0/11; P = 0.008, 0.0003, and 0.035, respectively) or sera (0/2, 0/8, and 0/5; P = 0.139, 0.002, and 0.009, respectively).IMPORTANCE The presence in cerebrospinal fluid of antibodies to EV peptides at higher levels than non-AFM controls supports the plausibility of a link between EV infection and AFM that warrants further investigation and has the potential to lead to strategies for diagnosis and prevention of disease

Aseptic Meningitis Epidemic during a West Nile Virus Avian Epizootic

While enteroviruses have been the most commonly identified cause of aseptic meningitis in the United States, the role of the emerging, neurotropic West Nile virus (WNV) is not clear. In summer 2001, an aseptic meningitis epidemic occurring in an area of a WNV epizootic in Baltimore, Maryland, was investigated to determine the relative contributions of WNV and enteroviruses. A total of 113 aseptic meningitis cases with onsets from June 1 to September 30, 2001, were identified at six hospitals. WNV immunoglobulin M tests were negative for 69 patients with available specimens; however, 43 (61%) of 70 patients tested enterovirus-positive by viral culture or polymerase chain reaction. Most (76%) of the serotyped enteroviruses were echoviruses 13 and 18. Enteroviruses, including previously rarely detected echoviruses, likely caused most aseptic meningitis cases in this epidemic. No WNV meningitis cases were identified. Even in areas of WNV epizootics, enteroviruses continue to be important causative agents of aseptic meningitis

Effect of substituting IPV for tOPV on immunity to poliovirus in Bangladeshi infants: An open-label randomized controlled trial

AbstractBackgroundThe Polio Endgame strategy includes phased withdrawal of oral poliovirus vaccines (OPV) coordinated with introduction of inactivated poliovirus vaccine (IPV) to ensure population immunity. The impact of IPV introduction into a primary OPV series of immunizations in a developing country is uncertain.MethodsBetween May 2011 and November 2012, we enrolled 700 Bangladeshi infant-mother dyads from Dhaka slums into an open-label randomized controlled trial to test whether substituting an injected IPV dose for the standard Expanded Program on Immunization (EPI) fourth tOPV dose at infant age 39 weeks would reduce fecal shedding and enhance systemic immunity. The primary endpoint was mucosal immunity to poliovirus at age one year, measured by fecal excretion of any Sabin virus at five time points up to 25 days post-52 week tOPV challenge, analyzed by the intention to treat principle.FindingsWe randomized 350 families to the tOPV and IPV vaccination arms. Neither study arm resulted in superior intestinal protection at 52 weeks measured by the prevalence of infants shedding any of three poliovirus serotypes, but the IPV dose induced significantly higher seroprevalence and seroconversion rates. This result was identical for poliovirus detection by cell culture or RT-qPCR. The non-significant estimated culture-based shedding risk difference was −3% favoring IPV, and the two vaccination schedules were inferred to be equivalent within a 95% confidence margin of −10% to +4%. Results for shedding analyses stratified by poliovirus type were similar.ConclusionsNeither of the vaccination regimens is superior to the other in enhancing intestinal immunity as measured by poliovirus shedding at 52 weeks of age and the IPV regimen provides similar intestinal immunity to the four tOPV series, although the IPV regimen strongly enhances humoral immunity. The IPV-modified regimen may be considered for vaccination programs without loss of intestinal protection

Comparison of fast-track diagnostics respiratory pathogens multiplex real-time RT-PCR assay with in-house singleplex assays for comprehensive detection of human respiratory viruses

Fast-track Diagnostics respiratory pathogens (FTDRP) multiplex real-time RT-PCR assay was compared with in-house singleplex real-time RT-PCR assays for detection of 16 common respiratory viruses. The FTDRP assay correctly identified 26 diverse respiratory virus strains, 35 of 41 (85%) external quality assessment samples spiked with cultured virus and 232 of 263 (88%) archived respiratory specimens that tested positive for respiratory viruses by in-house assays. Of 308 prospectively tested respiratory specimens selected from children hospitalized with acute respiratory illness, 270 (87.7%) and 265 (86%) were positive by FTDRP and in-house assays for one or more viruses, respectively, with combined test results showing good concordance (K=0.812, 95% CI = 0.786-0.838). Individual FTDRP assays for adenovirus, respiratory syncytial virus and rhinovirus showed the lowest comparative sensitivities with in-house assays, with most discrepancies occurring with specimens containing low virus loads and failed to detect some rhinovirus strains, even when abundant. The FTDRP enterovirus and human bocavirus assays appeared to be more sensitive than the in-house assays with some specimens. With the exceptions noted above, most FTDRP assays performed comparably with in-house assays for most viruses while offering enhanced throughput and easy integration by laboratories using conventional real-time PCR instrumentation. Published by Elsevier B.V.High Priority Pandemic and Seasonal Influenza Scientific proposal request initiativ

Characterization of poliovirus variants selected for resistance to the antiviral compound V-073

V-073, a small-molecule capsid inhibitor originally developed for nonpolio enterovirus indications is considerably more potent against polioviruses. All poliovirus isolates tested to date (n = 45), including wild, vaccine, vaccine-derived, and laboratory strains, are susceptible to the antiviral capsid inhibitor V-073. We grew poliovirus in the presence of V-073 to allow for the identification of variants with reduced susceptibility to the drug. Sequence analysis of 160 independent resistant variants (80 isolates of poliovirus type 1,40 isolates each of types 2 and 3) established that V-073 resistance involved a single amino acid change in either of two virus capsid proteins, VP1 (67 of 160 [42%]) or VP3 (93 of 160 [58%]). In resistant variants with a VP1 change, the majority (53 of 67 [79%]) exhibited a substitution of isoleucine at position 194 (equivalent position 192 in type 3) with either methionine or phenylalanine. Of those with a VP3 change, alanine at position 24 was replaced with valine in all variants (n = 93). The resistance phenotype was relatively stable upon passage of viruses in cell culture in the absence of drug. Single-step growth studies showed no substantial differences between drug-resistant variants and the virus stocks from which they were derived, while the resistant viruses were generally more thermally labile than the corresponding drug-susceptible parental viruses. These studies provide a foundation from which to build a greater understanding of resistance to antiviral compound V-073