Hepatitis A virus (HAV) packaging size limit

<p>Abstract</p> <p>Background</p> <p>Hepatitis A virus (HAV), an atypical Picornaviridae that causes acute hepatitis in humans, grows poorly in cell culture and in general does not cause cytopathic effect. Foreign sequences have been inserted into different parts of the HAV genome. However, the packaging size limit of HAV has not been determined. The purpose of the present study is to investigate the maximum size of additional sequences that the HAV genome can tolerate without loosing infectivity.</p> <p>Results</p> <p><it>In vitro </it>T7 polymerase transcripts of HAV constructs containing a 456-nt fragment coding for a blasticidin (Bsd) resistance gene, a 1,098-nt fragment coding for the same gene fused to GFP (GFP-Bsd), or a 1,032-nt fragment containing a hygromycin (Hyg) resistance gene cloned into the 2A-2B junction of the HAV genome were transfected into fetal Rhesus monkey kidney (FRhK4) cells. After antibiotic selection, cells transfected with the HAV construct containing the resistance gene for Bsd but not the GFP-Bsd or Hyg survived and formed colonies. To determine whether this size limitation was due to the position of the insertion, a 606 bp fragment coding for the Encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) sequence was cloned into the 5' nontranslated (NTR) region of HAV. The resulting HAV-IRES retained the EMCV IRES insertion for 1-2 passages. HAV constructs containing both the EMCV IRES at the 5' NTR and the Bsd-resistance gene at the 2A-2B junction could not be rescued in the presence of Bsd but, in the absence of antibiotic, the rescued viruses contained deletions in both inserted sequences.</p> <p>Conclusion</p> <p>HAV constructs containing insertions of approximately 500-600 nt but not 1,000 nt produced viable viruses, which indicated that the HAV particles can successfully package approximately 600 nt of additional sequences and maintain infectivity.</p

Pancreatic surgery outcomes: multicentre prospective snapshot study in 67 countries

Background: Pancreatic surgery remains associated with high morbidity rates. Although postoperative mortality appears to have improved with specialization, the outcomes reported in the literature reflect the activity of highly specialized centres. The aim of this study was to evaluate the outcomes following pancreatic surgery worldwide.Methods: This was an international, prospective, multicentre, cross-sectional snapshot study of consecutive patients undergoing pancreatic operations worldwide in a 3-month interval in 2021. The primary outcome was postoperative mortality within 90 days of surgery. Multivariable logistic regression was used to explore relationships with Human Development Index (HDI) and other parameters.Results: A total of 4223 patients from 67 countries were analysed. A complication of any severity was detected in 68.7 percent of patients (2901 of 4223). Major complication rates (Clavien-Dindo grade at least IIIa) were 24, 18, and 27 percent, and mortality rates were 10, 5, and 5 per cent in low-to-middle-, high-, and very high-HDI countries respectively. The 90-day postoperative mortality rate was 5.4 per cent (229 of 4223) overall, but was significantly higher in the low-to-middle-HDI group (adjusted OR 2.88, 95 per cent c.i. 1.80 to 4.48). The overall failure-to-rescue rate was 21 percent; however, it was 41 per cent in low-to-middle-compared with 19 per cent in very high-HDI countries.Conclusion: Excess mortality in low-to-middle-HDI countries could be attributable to failure to rescue of patients from severe complications. The authors call for a collaborative response from international and regional associations of pancreatic surgeons to address management related to death from postoperative complications to tackle the global disparities in the outcomes of pancreatic surgery (NCT04652271; ISRCTN95140761)