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SUMMARYFour UK strains of three different serotypes were found to differ by only 2-3% of their S1 amino acids. The S1 sequences were also very similar to those of three Dutch isolates (D207, D274 and D3896), the greatest difference between two of the seven isolates being 4.4%. The few amino acid differences between the seven isolates were located largely between residues 19-122 and 251-347 of the mature S1 subunit.The seven isolates could be differentiated using 16 monoclonal antibodies in an enzymelinked immunosorbent assay. Some virus neutralizing (VN) antibody-inducing epitopes were common to all seven isolates even though the strains had been differentiated into three serotypes by polyclonal sera. The results indicate that the most antigenic of the VN antibody-inducing epitopes are formed by very few amino acids and that these occur in the first and third quarters of the S1 subunit. We suggest that serology-based epizootiological studies of IBV should, therefore, be augmented by the inclusion of nucleic acid sequencing and/or monoclonal antibody analysis.

The spike glycoprotein (S) gene of IBV codes for a precursor protein which is cleaved into the N-terminal S1 and C-terminal S2 glycopolypeptides. The S1 glycopolypeptide, which induces neutralizing antibody, comprises approximately 520 amino acid residues. We have determined the nucleotide sequence of S1 of seven strains of the Massachusetts (Mass) serotype and the first 337 bases of two additional Mass strains. Despite the fact that the strains had been isolated over three decades in Europe and the U.S.A. there was only 4% base and 6% amino acid variation within the group. Nearly one third of the 32 amino acid differences in S1 were in two hypervariable regions (HVRs 1 and 2) comprising residues 38-51 and 99-115, identified by Niesters et al. (1986), showing that HVRs 1 and 2 are a feature of the Mass serotype. Amino acid variation within HVRs 1 and 2 was 29% and 40% respectively. Five vaccine strains could be distinguished from each other by sequencing of the first 337 nucleotides. Variants of M41 which resisted neutralization by two monoclonal antibodies (A13 and A38) had the same, single base change at position 134, resulting in substitution of proline residue 45 by histidine. This indicates that residues within HVR 1 are associated with epitopes which induce neutralizing antibody.

SUMMARYAvian infectious bronchitis coronavirus (IBV) inactivated by fl-propiolactone induced partial protection of the trachea in up to 40~ of chickens following one intramuscular inoculation 4 to 6 weeks prior to challenge. Retention of an intact tracheal ciliated epithelium 4 days after challenge was the criterion of protection. There was no correlation between protection and serum titres of virus-neutralizing (VN) and haemagglutination-inhibiting (HI) antibody, which were maximal at about 4 weeks after inoculation. Virus from which the S1 but not the $2 (spike-anchoring) spike glycopolypeptide had been removed by urea did not induce protection or VN or HI antibody. Four intramuscular inoculations of monomeric S1 induced VN and HI antibody in two and four chickens respectively. These results indicate that VN and HI antibodies are induced primarily by S1, that intact spikes are a major requirement for the induction of protective immunity and that this property is probably associated with S1.

SUMMARYUrea has been used to remove the S1 spike glycopolypeptide from avian infectious bronchitis virus (IBV) strains M41 and Beaudette, without removing the $2 spikeanchoring glycopolypeptide. Reduction of the pH to 2-9 did not cause release of S 1 although some S 1 was released spontaneously from IBV Beaudette at pH 7.4. Virus that lacked S1 was no longer infectious or able to cause haemagglutination (HA). However, radiolabelled IBV that lacked S 1 attached to erythrocytes and chick kidney cells to the same or similar extent as did intact virus. Treatment of IBV with a phospholipase C preparation, required to make IBV cause HA, did not increase binding of IBV to erythrocytes. The results indicate that while the attachment to cells of virus that lacks S1 is qualitatively different from that of intact virus, the decline in infectivity is the consequence of the loss of some other spike function.

BackgroundOesophagogastric cancer surgery is immunosuppressive. This may be modulated by omega-3 fatty acids (O-3FAs). The aim of this study was to assess the effect of perioperative O-3FAs on clinical outcome and immune function after oesophagogastric cancer surgery.MethodsPatients undergoing subtotal oesophagectomy and total gastrectomy were recruited and allocated randomly to an O-3FA enteral immunoenhancing diet (IED) or standard enteral nutrition (SEN) for 7 days before and after surgery, or to postoperative supplementation alone (control group). Clinical outcome, fatty acid concentrations, and HLA-DR expression on monocytes and activated T lymphocytes were determined before and after operation.ResultsOf 221 patients recruited, 26 were excluded. Groups (IED, 66; SEN, 63; control, 66) were matched for age, malnutrition and co-morbidity. There were no differences in morbidity (P = 0·646), mortality (P = 1·000) or hospital stay (P = 0·701) between the groups. O-3FA concentrations were higher in the IED group after supplementation (P < 0·001). The ratio of omega-6 fatty acid to O-3FA was 1·9:1, 4·1:1 and 4·8:1 on the day before surgery in the IED, SEN and control groups (P < 0·001). There were no differences between the groups in HLA-DR expression in either monocytes (P = 0·538) or activated T lymphocytes (P = 0·204).ConclusionDespite a significant increase in plasma concentrations of O-3FA, immunonutrition with O-3FA did not affect overall HLA-DR expression on leucocytes or clinical outcome following oesophagogastric cancer surgery. Registration number: ISRCTN43730758 (http://www.controlled-trials.com). Copyright © 2012 British Journal of Surgery Society Ltd. Published by John Wiley & Sons, Ltd.

Epifluorescence microscopy was used to investigate the effect of cholesterol on monolayers of dipalmitoylphosphatidylcholine (DPPC) and 1 -palmitoyl-2-oleoyl phosphatidylcholine (POPC) at 21 +/- 2 degrees C using 1 mol% 1-palmitoyl-2-[12-[(7-nitro-2-1, 3-benzoxadizole-4-yl)amino]dodecanoyl]phosphatidylcholine (NBD-PC) as a fluorophore. Up to 30 mol% cholesterol in DPPC monolayers decreased the amounts of probe-excluded liquid-condensed (LC) phase at all surface pressures (pi), but did not effect the monolayers of POPC, which remained in the liquid-expanded (LE) phase at all pi. At low pi (2-5 mN/m), 10 mol% or more cholesterol in DPPC induced a lateral phase separation into dark probe-excluded and light probe-rich regions. In POPC monolayers, phase separation was observed at low pi when > or =40 mol% or more cholesterol was present. The lateral phase separation observed with increased cholesterol concentrations in these lipid monolayers may be a result of the segregation of cholesterol-rich domains in ordered fluid phases that preferentially exclude the fluorescent probe. With increasing pi, monolayers could be transformed from a heterogeneous dark and light appearance into a homogeneous fluorescent phase, in a manner that was dependent on pi and cholesterol content. The packing density of the acyl chains may be a determinant in the interaction of cholesterol with phosphatidylcholine (PC), because the transformations in monolayer surface texture were observed in phospholipid (PL)/sterol mixtures having similar molecular areas. At high pi (41 mN/m), elongated crystal-like structures were observed in monolayers containing 80-100 mol% cholesterol, and these structures grew in size when the monolayers were compressed after collapse. This observation could be associated with the segregation and crystallization of cholesterol after monolayer collapse.

The spike protein of avian infectious bronchitis coronavirus comprises two glycopolypeptides S1 and S2 derived by cleavage of a proglycopolypeptide So, the nucleotide sequence of which has recently been determined for the Beaudette strain (Binns, M.M. et al., 1985, J. Gen. Virol. 66, 719-726). The order of the two glycopolypeptides within So is aminoterminus(N)-S1-S2-carboxyterminus(C). To locate the N-terminus of S2 we have performed partial amino acid sequencing on S2 from IBV-Beaudette labelled with [3H]serine and from the related strain labelled with [3H]valine, leucine and isoleucine. The residues identified and their positions relative to the N-terminus of S2 were: serine, 13; valine, 6, 12; leucine, none in the first 20 residues; isoleucine, 2, 19. These results identified the N-terminus of S2 of IBV-Beaudette as serine, 520 residues from the N-terminus of S1, excluding the signal sequence. Immediately to the N-terminal side of residue 520 So has the sequence Arg-Arg-Phe-Arg-Arg; similar basic connecting peptides are a feature of several other virus spike glycoproteins. It was deduced that for IBV-Beaudette S1 comprises 519 residues (Mr 57.0K) or 514 residues (56.2K) if the connecting peptide was to be removed by carboxypeptidase-like activity in vivo while S2 has 625 residues (69.2K). Nucleotide sequencing of the cleavage region of the So gene of IBV-M41 revealed the same connecting peptide as IBV-Beaudette and that the first 20 N-terminal residues of S2 of IBV-M41 were identical to those of the Beaudette strain. IBV-Beaudette grown in Vero cells had some uncleaved So; this was cleavable by 10 micrograms/ml of trypsin and of chymotrypsin. Partial N-terminal analysis of S1 from IBV-M41 identified leucine and valine residues at positions 2 and 9 respectively from the N-terminus. This confirms the identification, made by Binns et al. (1985), of the N-terminus of S1 and the end of the signal sequence of the IBV-Beaudette spike propolypeptide. N-terminal sequencing of [3H]leucine-labelled IBV-Beaudette membrane (M) polypeptide showed leucine residues at positions 8, 16 and 22 from the N-terminus; these results confirm the open reading frame identified by M.E.G. Boursnell et al. (1984, Virus Res. 1, 303-313) in the nucleotide sequence of M. The N-terminus of the nucleocapsid (N) polypeptide appeared to be blocked.