pH inactivation of SARS-CoV-2 and SARS-CoV in virus spiked protein A eluates from a mAb purification process.

Biopharmaceutical manufacturing processes may include a low pH treatment step as a means of inactivating enveloped viruses. Small scale virus clearance studies are routinely performed using model enveloped viruses such as murine leukemia virus to assess inactivation at the pH range used in the downstream manufacturing process. Further, as a means of bioburden reduction, chromatography resins may be cleaned and stored using sodium hydroxide and this can also inactivate viruses. The susceptibility of SARS-CoV-2 and SARS-CoV to low pH conditions using protein A eluate derived material from a monoclonal antibody production process as well as high pH cleaning conditions was addressed. SARS-CoV-2 was effectively inactivated at pH 3.0, moderately inactivated at pH 3.4, but not inactivated at pH 3.8. Low pH was less effective at inactivating SARS-CoV. Both viruses were inactivated at a high pH of ca.13.4. These studies provide important information regarding the effectiveness of viral clearance and inactivation steps of novel coronaviruses when compared to other enveloped viruses.

Retrospective Evaluation of Cycled Resin in Viral Clearance Studies-A Multiple Company Collaboration.

The BioPhorum Development Group Viral Clearance Workstream performed a collaborative retrospective analysis to evaluate packed bed chromatographic resin performance after repeated cycling for two commonly used chromatography steps in biopharmaceutical manufacturing: protein A and anion exchange. Key variables evaluated in the assessment included virus type, resin type, number of reuse cycles, and virus challenge. In this retrospective analysis of viral clearance data on naïve versus cycled resin, powered by the availability of a decade's worth of accumulated industry data, clearance capability was not negatively impacted by resin cycling. This finding is consistent with publications showing that surrogates for viral clearance capabilities could be employed in lieu of testing the viral clearance of cycled resins for protein A and anion exchange chromatography. The rigorous analysis of the retrospective data supports the view that viral clearance studies for cycled resins are not necessary provided that appropriate cleaning methods are applied during repeated use of the chromatography columns.LAY ABSTRACT: The manufacturing processes for biopharmaceutical products often include reusable chromatographic resins that remove process- and product-related impurities as well as potential contaminating viruses. Typically, chromatography resin is "cycled" through repeated steps of resin conditioning, product purification, and resin cleaning. The cycling approach has been evaluated in both small- and full-scale studies that show the performance parameters are maintained. The ability to remove virus is demonstrated separately in a focused small-scale virus-spiking study that is resource-intensive and costly. This paper is a retrospective review of industry data comparing virus removal by naïve and repeatedly cycled resins that summarizes the viral clearance impact of re-using protein A and anion exchange chromatography resins. The key variables evaluated in the assessment included virus type, resin type, number of cycles, and virus challenge. In this retrospective analysis, it was found that the viral clearance capability is not negatively impacted by resin cycling. This finding is consistent with other publications and supports the view that viral clearance studies for cycled resins are not necessary if appropriate cleaning methods are applied during the repeated use of the chromatography columns.Abbreviations: AAV-2, Adeno-associated virus; A-MuLV, Amphotropic murine leukemia virus; AEX, Anion-exchange chromatography; B/E, Bind and elute; BVDV, Bovine viral diarrhea virus; C.P.G., Controlled pore glass; DEAE, Diethylaminoethanol; EMCV, Encephalomyocarditis virus; FT, Flow through; HAV, Hepatitis A virus; HSV-1, Herpes simplex virus type 1; LOD, Limit of detection; LOQ, Limit of quantification; LRF, Log10 reduction factor; mAb, Monoclonal antibody; MVM, Minute virus of mice; NaOH, Sodium hydroxide; PA, Protein A; PPV, Porcine parvovirus; QA, Quaternary amine; QP, Quaternized polyethyleneimine; qPCR, Quantitative polymerase chain reaction; Reo3, Reovirus type 3; SuHV-1, Suid herpesvirus; SV40, Simian virus 40; X-MuLV, Xenotropic murine leukemia virus.

Turning up the heat: Effect of new vaccine for children's (VFC) program recommendations for use of temperature monitors upon incorrect product storage adverse event reporting.

BACKGROUND: The Centers for Disease Control and Prevention (CDC) coordinates the Vaccines For Children (VFC) program, which provides free vaccines to qualified children in the US. In 2009, the CDC issued Vaccine Storage Requirements, which were later replaced (2012) with an interim guidance and toolkit for vaccine storage and handling. The guidance called for use of Digital Data Loggers (DDL) to monitor vaccine storage temperatures. We describe a change in frequency of Incorrect Product Storage Reports (IPSRs) following issuance of the 2009 CDC guidance. METHODS: Merck & Co., Inc., Kenilworth, NJ, USA, systematically evaluates vaccine safety concerns for all products. The safety database was queried (01-Jan-2004 through 31-December-2016) to identify all IPSRs associated with 10 vaccines. We compared IPSRs received prior to and following the 2009 CDC guidance, comparing reports received from the US with those received from international sources during the same period. RESULTS: Following the release of the DDL guidance, a progressive increase in IPSRs was identified in the US (1 report received in 2004, 12,993 reports in 2016). In contrast, non-US IPSRs - have not had a similar increase: no reports received in 2004, 216 reports received in 2016. US reports of IPSRs 2004 through 2016 account for 96% of reports worldwide. There were no serious reports found in the database in conjunction with IPSRs, nor were there any additional safety findings in any of the reports with additional events reported. CONCLUSION: VFC DDL guidance was followed by an increase in IPSRs. No similar trend was seen outside the US (where no broad change in DDL guidance occurred). Despite the increase in IPSRs, there have been few associated adverse events (AEs) reported; no new safety concerns were identified. These findings suggest that the increase in IPSRs was associated with the introduction of use of DDLs, and suggests the need for further impact assessment.

Retrospective Evaluation of Low-pH Viral Inactivation and Viral Filtration Data from a Multiple Company Collaboration.

Considerable resources are spent within the biopharmaceutical industry to perform viral clearance studies, which are conducted for widely used unit operations that are known to have robust and effective retrovirus clearance capability. The collaborative analysis from the members of the BioPhorum Development Group Viral Clearance Working Team considers two common virus reduction steps in biopharmaceutical processes: low-pH viral inactivation and viral filtration. Analysis included eight parameters for viral inactivation and nine for viral filtration. The extensive data set presented in this paper provides the industry with a reference point for establishing robust processes in addition to other protocols available in the literature (e.g., ASTM Std. E2888-12 for low-pH inactivation). In addition, it identifies points of weakness in the existing data set and instructs the design and interpretation of future studies. Included is an abundance of data that would have been difficult to generate individually but collectively will help support modular viral clearance claims.

Effect of beta lactam antibiotic use in children on pneumococcal resistance to penicillin: prospective cohort study.

OBJECTIVE: To examine the relation between use of antibiotics in a cohort of preschool children and nasal carriage of resistant strains of pneumococcus. DESIGN AND PARTICIPANTS: Prospective cohort study over two years of 461 children aged under 4 years living in Canberra, Australia. MAIN OUTCOME MEASURES: Use of drugs, respiratory symptoms, and visits to doctors were documented in a daily diary by parents of the children during 25 months of observation. Isolates of pneumococci, which were cultured from nasal swabs collected approximately six monthly, were tested for antibiotic resistance. RESULTS: From the four swab collections 631 positive pneumococcal isolates from 461 children were found, of which 13.6% were resistant to penicillin. Presence of penicillin resistant pneumococci was significantly associated with children's use of a beta lactam antibiotic in the two months before each swab collection (odds ratio 2.03 (95% confidence interval 1.15 to 3.56, P=0.01)). The odds ratio of the association remained >1 (though did not reach significance at the 0.05 level) for use in the six months before swab collection. The association was seen in children who received only penicillin or only cephalosporin antibiotics in that period. The odds ratio was 4.67 (1.29 to 17.09, P=0.02) in children who had received both types of beta lactam in the two months before their nasal swab. The modelled odds of carrying penicillin resistant pneumococcus was 4% higher for each additional day of use of beta lactam antibiotics in the six months before swab collection. CONCLUSIONS: Reduction in beta lactam use could quickly reduce the carriage rates of penicillin resistant pneumococci in early childhood. In view of the propensity of these organisms to be spread among children in the community, the prevalence of penicillin resistant organisms may fall as a consequence.

Varicella vaccine exposure during pregnancy: data from 10 Years of the pregnancy registry.

BACKGROUND: The Pregnancy Registry for Varivax (Merck) was established to monitor for congenital varicella syndrome or other birth defects in the offspring of women who were exposed to varicella vaccine while pregnant. METHODS: The registry receives voluntary reports from health care providers or consumers about women given the vaccine 3 months before or during pregnancy. Follow-up is conducted to obtain and classify pregnancy outcomes. All reports are evaluated for the presence of birth defects. Outcomes from prospectively reported pregnancy exposures are used to calculate rates and 95% confidence intervals. RESULTS: From 17 March 1995 through 16 March 2005, 981 women were enrolled. Pregnancy outcomes were available for 629 prospectively enrolled women. Among the 131 live births to varicella-zoster virus-seronegative women, there was no evidence of congenital varicella syndrome (rate, 0% [95% confidence interval [CI], 0%-6.7%]), and major birth defects were observed in 3 infants (rate, 3.7% [95% CI, 0.8%-10.7%]). CONCLUSIONS: Although the numbers of exposures are not sufficient to rule out a very low risk, data collected in the pregnancy registry to date do not support a relationship between the occurrence of congenital varicella syndrome or other birth defects and varicella vaccine exposure during pregnancy.

Anxiety, post-traumatic stress disorder and depression in Korean War veterans 50 years after the war.

BACKGROUND: There has been no comprehensive investigation of psychological health in Australia's Korean War veteran population, and few researchers are investigating the health of coalition Korean War veterans into old age. AIMS: To investigate the association between war service, anxiety, post-traumatic stress disorder (PTSD) and depression in Australia's 7525 surviving male Korean War veterans and a community comparison group. METHOD: A survey was conducted using a self-report postal questionnaire which included the PTSD Checklist, the Hospital Anxiety and Depression scale and the Combat Exposure Scale. RESULTS: Post-traumatic stress disorder (OR 6.63, P<0.001), anxiety (OR 5.74, P<0.001) and depression (OR 5.45, P<0.001) were more prevalent in veterans than in the comparison group. These disorders were strongly associated with heavy combat and low rank. CONCLUSIONS: Effective intervention is necessary to reduce the considerable psychological morbidity experienced by Korean War veterans. Attention to risk factors and early intervention will be necessary to prevent similar long-term psychological morbidity in veterans of more recent conflicts.

Changing GPs' antibiotic prescribing: a randomised controlled trial.

A randomised controlled trial involving 54 general practitioners (GPs) was conducted in Canberra, Australian Capital Territory from September 1997 to November 1999. In the first year of the study, 24 GPs, who constituted the active arm of the intervention group, were involved in the consideration of evidence and the development and implementation of a set of clinical guidelines for the treatment of acute respiratory infections. These guidelines were then endorsed in a meeting together with specialist colleagues. In the second year of the study the group of GPs who had been acting as controls, received a moderate intervention consisting of a brief educational event and distribution of the locally developed guidelines. We obtained data from January 1997 to December 1999 from the Health Insurance Commission on prescribing rates for 40 of the doctors in the study. The rate of prescribing was calculated as the number of antibiotic prescriptions per 100 Medicare services. The average yearly prescribing decreased significantly in the intensive intervention group and increased in the moderate intervention group, (p=0.026). A mixed effects longitudinal time series model was fitted to the data to account for seasonal variation of antibiotic prescribing and trends over time. The intensive intervention group significantly reduced their antibiotic prescribing over time compared to the moderate intervention group, (p<0.001). This study has shown that an intensive intervention in which general practitioners were actively engaged in development and consideration of the evidence base for the guidelines resulted in a significant fall in general antibiotic prescribing.

Crystallization and preliminary X-ray analysis of the acyl carrier protein synthase (AcpS) from Staphylococcus aureus.

Acyl carrier protein synthase (AcpS) catalyzes the transfer of 4'-phophopantetheine from coenzyme A to the acyl carrier protein (ACP) to activate it for fatty-acid biosynthesis. Two crystal forms of Staphylococcus aureus AcpS have been generated at 277 K using either NaCl or PEG 6000 as a precipitant. The diffraction patterns of the crystals extend to 1.65 and 1.8 A, respectively. Full sets of X-ray diffraction data were collected from native crystals and the crystal structures were solved by molecular replacement.