A survey of Mexican retail chain stores for fresh U.S. pork.

An overview of fresh US pork in the Mexican market was achieved by surveying fresh US pork packages (n=342) for sale in five Mexican cities. Data on cut, primal/sub-primal from which the cut was sourced, subcutaneous and seam fat thicknesses, marbling scores, and presence of bone were collated. The most prevalent identifiable retail cuts were milanesa (thin slice of pork, breaded or non-breaded) and trozos (diced pork) derived primarily from the leg and accounting for 68% of the total US pork on sale. Over 90% of the retail cuts were trimmed to 3.2mm or less of external fat and the average marbling score was 2.26. Differences in distribution and fat measures were observed with chain, location and socio-economic status of clientele indicating potential for a targeted marketing approach in Mexico.

Definition, willingness-to-pay, and ranking of quality attributes of U.S. pork as defined by importers in Asia and Mexico.

A survey was conducted from November 2009 to April 2010 to determine how importers of pork define 7 predetermined quality categories (food safety, customer service, eating quality, product specification, packaging, visual characteristics, and production history) and to estimate willingness-to-pay (WTP) and establish best-worst (B/W) scaling (rank) for the 7 quality categories. Interviews were conducted in Hong Kong/China (n = 83), Japan (n = 48), Mexico (n = 70) and Russia (n = 54) with importers of U.S. pork or those who had purchased U.S. pork from distributors in the last 3 yr. Interviews used dynamic routing software and were structured such that economic factors for purchase were addressed first, allowing all responses to focus on quality. Questions about WTP and B/W were asked and then each respondent was asked to define what each quality category meant to them. Generalized linear mixed models were used to analyze frequency data. Over 70% of interviewees in Hong Kong/China, Japan, and Mexico responded that purchase price was influential in deciding whether or not to purchase imported pork. This number was lower in Russia, where respondents stated tariff rates were also important, indicating market access was a larger issue in Russia. Food safety was the most important quality category (price was not included as a part of quality) for imported pork followed by specifications. Respondents indicated some form of government inspection was how they defined food safety, whereas product size, weight, and subcutaneous fat were all included in the definition of specifications. Interviewees were more likely to pay premiums for customer service and less likely to pay premiums for packaging (P < 0.05). The premiums that were willing to be paid for guarantees of quality for imported pork variety meats were numerically lower than for whole muscle cuts or processed products. A guarantee associated with food safety of processed pork products was found to be the quality attribute for which importers would be willing to pay the highest premium. Production history was found to be the least important quality attribute for importers of all types of U.S. pork, except those in Japan. Exporters could increase profitability if a guarantee of customer service was made. Price, tariffs, and exchange rates are important to pork importers; these results indicated that if certain quality attributes could be guaranteed, exporters could increase profitability.

Effects of dietary ractopamine hydrochloride and zilpaterol hydrochloride supplementation on performance, carcass traits, and carcass cutability in beef steers.

British × Continental steers (initial BW = 484.6 kg) were fed at a commercial feed yard to evaluate the effects of ß-agonists on live performance, carcass characteristics, and carcass subprimal yield. Weights and ultrasonic measurements were used to allocate steers to pens (n = 40) divided equally into 4 blocks, with 2 treatment replicates per block. Pens were randomly assigned to 1 of 5 treatments: control; ractopamine-HCl (RH) fed at 200 or 300 mg • steer(-1) • d(-1), or 400 mg • steer(-1) • d(-1) top dress for the final 30 d of feeding; or zilpaterol-HCl (ZH) fed at 7.5 mg/kg beginning 23 d before slaughter with a 3-d withdrawal period. Steers were harvested by block at a commercial facility over 4 wk. Carcass based performance measures were calculated using initial pen weights and actual DMI. From each pen, eight carcasses that were within ± 13.6 kg of the mean pen HCW were selected such that two carcasses were within each of the following four Yield Grade (YG) ranges: YG ≤ 2.8; 2.9-3.2; 3.3-3.5; YG > 3.5. Carcasses were fabricated by plant personnel to determine subprimal yield. Steers fed ZH had higher carcass-based ADG and carcass-based G:F compared with all other treatments (P < 0.05). Carcass-based ADG and carcass-based G:F were higher in RH treatments compared with controls (P < 0.05). Steers fed ZH had higher dressing percentages (1.0 to 1.6%) and larger LM area (4.3 to 6.7 cm(2)) than all other treatments (P < 0.05). Use of RH 400 and ZH increased HCW 6.3 and 11.1 kg, respectively compared with controls (P < 0.05). Compared with controls, RH 300 and ZH decreased marbling score and the frequency of carcasses qualifying for upper 2/3 Choice premiums (P < 0.05). Beta-agonists increased subprimal yield from the round and loin; however, blade meat was the only cut from the rib or chuck affected by ß-agonists. Results from this study indicated improvements in performance and carcass traits as a result of ß-agonist use; however, differences between ZH, RH 400, and RH 300 treatments were minimal for carcass traits and cutability. Increases in saleable yield following ß-agonist use were not uniformly distributed across the four major primals and the majority of weight gain occurred in the lower priced cuts of the round and chuck. Increased response of the lower priced cuts to ß-agonists could have economic implications to packers.

Effects of ractopamine hydrochloride and zilpaterol hydrochloride supplementation on longissimus muscle shear force and sensory attributes of calf-fed Holstein steers.

The effect of ractopamine hydrochloride (RH) and zilpaterol hydrochloride (ZH) on slice shear force (SSF) and sensory characteristics of beef from calf-fed Holstein steers was evaluated. All steers were implanted with a progesterone (100 mg) plus estradiol benzoate (10 mg) implant followed by a terminal trenbolone acetate (200 mg) plus estradiol (40 mg) implant. Steers were blocked by weight into pens (n = 32) randomly assigned to 1 of 4 treatments: control, RH fed at 300 mg·steer(-1)·d(-1) (RH 300) or RH fed at 400 mg·steer(-1)·d(-1)(RH 400) for the final 31 d of finishing, or ZH fed at 6.8 g/t for 21 d with a 5-d withdrawal before harvest. Fourteen carcasses were randomly selected from each pen, and two LM samples (1 per side) were excised and aged either 14 or 21 d before SSF testing. For trained panel evaluation, two steaks were collected from each of 60 low Choice strip loins (20 each from control, RH 300, and ZH treatments) and aged either 14 or 21 d. Steers fed RH and ZH produced steaks with SSF values that were 9% to 25% higher than controls. No difference in SSF was detected between the two levels of RH (P > 0.05). Compared to controls, the probability of steaks aged 14 d failing to meet SSF requirements to be certified tender (SSF < 20 kg) was increased 0.15, 0.17, and 0.26 in steers fed RH 300, RH 400, and ZH, respectively. Compared to controls, the probability of steaks aged 21 d having SSF values >20 kg was increased 0.03, 0.08, and 0.16 in steers fed RH 300, RH 400, and ZH, respectively. Steaks from Select carcasses of steers fed ZH aged 21 d postmortem had double the probability (0.39 vs. 0.17) of having SSF values >20 kg compared to steaks from steers fed either level of RH (P < 0.05). This difference tended to be identical in steaks from Select carcasses 14 d postmortem (0.50 vs. 0.33; P = 0.11); however, no difference was found in low Choice samples at 14 or 21 d postmortem. Trained panelists rated steaks aged 14 d from steers fed ZH lower for overall tenderness and flavor compared to controls (P < 0.05); however, no difference was found between controls and those fed RH 300. Steaks from steers fed ZH aged 21 d were rated lower for overall tenderness and juiciness compared to controls and those from steers fed RH 300 (P < 0.05). This study suggests RH and ZH negatively impact sensory attributes of beef from calf-fed Holstein steers.

Effects of ractopamine hydrochloride and zilpaterol hydrochloride supplementation on carcass cutability of calf-fed Holstein steers.

Effects of ractopamine hydrochloride (RH) and zilpaterol hydrochloride (ZH) on saleable yield of carcass sides from calf-fed Holstein steers were evaluated using steers implanted with a progesterone (100 mg) plus estradiol benzoate (10 mg) implant followed by a terminal trenbolone acetate (200 mg) plus estradiol (40 mg) implant. Steers were blocked by weight into pens (n = 32) randomly assigned to one of four treatments: control, RH fed at 300 mg•steer(-1)/d(-1) (RH 300) or RH fed at 400 mg•steer(-1)/d(-1) (RH 400) the final 31 d of finishing, and ZH fed at 60 to 90 mg•steer(-1)/d(-1) (7.56 g/ton on a 100% DM basis) for 21 d with a 5 d withdrawal before harvest. Eight to nine carcass sides were randomly selected from each pen; carcass sides with excessive hide pulls, fat pulls or bruises were avoided. Cutout data were collected within a commercial facility using plant personnel to fabricate sides at a rate of one every 3 to 4 min into items typically merchandised by the facility. All lean, fat and bone were weighed and summed back to total chilled side weight with a sensitivity of ± 2% to be included in the data set. Compared to controls, ß-agonists increased saleable yield of whole-muscle cuts by 0.61%, 0.86% and 1.95% for RH 300, RH 400 and ZH, respectively (P < 0.05). Percent fat was less in carcasses from the ZH treatment compared to controls (P < 0.05); however, this difference was not observed between RH treatments and controls (P > 0.05). Percent bone was less in the ZH treatment due to increased muscle (P < 0.05). The percent of chilled side weight comprised of trimmings was unchanged between treatments, but on a 100% lean basis, RH 400 and ZH increased trim yields (P < 0.05). Analysis of saleable yield by primal showed a fundamental shift in growth and development. Beta-agonists caused a shift in proportion of saleable yield within individual primals, with a greater portion produced from the hindquarter relative to the forequarter, specifically in those muscles of the round (P < 0.05). Beta-agonists increased saleable yield, but these effects were not constant between all major primals. The cutout value gained by packers as a result of ß-agonist use may be influenced more by reduced fatness and increased absolute weight if musculature is primarily increased in the lower priced cuts of the carcass.

Effects of ractopamine hydrochloride and zilpaterol hydrochloride supplementation on longissimus muscle shear force and sensory attributes of beef steers.

Effect of ractopamine hydrochloride (RH) and zilpaterol hydrochloride (ZH) on LM shear force and sensory attributes was determined using pens (n = 40) British × Continental crossbred steers randomly allocated to one of the following treatments: control; RH fed at 200 (RH 200) or 300 mg • steer(-1) • d(-1) (RH 300), or 400 mg • steer(-1) • d(-1) (RH 400) top-dressed for the final 30 d of feeding; or ZH fed at 7.5 mg/kg, beginning 23 d before slaughter with a 3-d withdrawal. Two replicates (pens) per treatment were represented in four blocks. Eighteen carcasses per pen were randomly selected and one 5-cm LM sample was removed from both carcass sides to be used for shear force and sensory evaluation. Samples were aged for 14 d, frozen at -28.8 °C, and cut into 2.5-cm steaks. All steaks were cooked to an internal temperature of 71.1 °C before being evaluated for Warner-Bratzler shear force (WBSF), slice shear force (SSF), or being fed to trained sensory panelists. Increasing dose and potency of ß-agonist increased WBSF by 4 to 17% and SSF by 5 to 24% (P < 0.05). Steaks from steers fed ZH had higher WBSF and SSF values compared with all other treatments (P < 0.05), whereas steaks from controls and steers fed RH 200 were not different (P > 0.05). Probability of steaks failing to meet shear force standards to be certified tender (WBSF <4.4 kg, SSF < 20 kg) was increased from an initial probability of <0.06 in steaks from steers in the control treatment to 0.10 to 0.20 in steers fed RH 400 or ZH (P < 0.05). No difference was detected in panel ratings for overall tenderness of steaks from steers fed RH 200 compared with controls (P > 0.05). Steaks from steers fed RH 300 and RH 400 were comparable for all sensory attributes; however, both RH 300 and RH 400 were rated lower for overall tenderness than controls (P < 0.05). Panelists failed to detect differences in overall tenderness of steaks from steers fed RH 400 and ZH (P < 0.05). Panelists detected no difference in flavor profile or juiciness among treatments (P > 0.05). Results from this study indicated ß-agonists negatively affected beef tenderness and these effects may be more noticeable in steers supplemented with ZH and higher doses of RH.

North American Beef Tenderness Survey 2011-2012: benchmarking tenderness and sample shipping procedures.

Fifty-four stores in 30 U.S. cities were sampled from June 2011 through May 2012 to benchmark beef tenderness at retail, as assessed by Warner-Bratzler shear force (WBSF). Top loin (Longissimus dorsi; n = 980) and sirloin (Gluteus medius and Biceps femoris; n = 860) steaks were collected at random (Quality Grade and brand) and shipped via overnight delivery to Colorado State University. From June 2011 through November 2011, North American Beef Tenderness Survey (NABTS) Period 1, samples were shipped fresh and then frozen. Mean WBSF values during Period 1 were 2.9 and 3.5 kg for top loin and sirloin steaks, respectively. Frequencies of steaks classified as tough (WBSF ≥ 4.4 kg) were 8.6% and 17.7% for top loin and sirloin steaks, respectively. When shipped fresh, a disproportionately high frequency (16.9%) of top loin steaks had WBSF ≤ 2.0 kg, representing a deviation from previous works. Two trials were conducted to assess the effect of freezing, retail display, and shipping on WBSF and slice shear force (SSF) of beef top loin steaks. Freezing, retail display, and shipment reduced WBSF by 0.4, 0.3, and 0.0 kg during Trial 1, and by 0.4, 0.3, and 0.1 kg during Trial 2. Slice shear force was lower (P < 0.05) in steaks exposed to shipping conditions during Trial 1; however, this difference was not observed in Trial 2. Shipping decreased the frequency of steaks categorized as tough (SSF ≥ 20.0 kg) from 11.1 to 5.7% and from 30.5 to 28.6%, during Trial 1 and 2, respectively. During Trial 1, WBSF indicated that shipping increased incidence of tough samples from 0.0 to 3.8%, but this trend was reversed during Trial 2 when shipping reduced incidence of tough samples from 13.0 to 5.6%. Coefficients of variation for treatment effects suggested variance remained unchanged (± 2.0%), with respect to shear force values. However, mean values were reduced as a result of shipping conditions. These findings dictated a change in NABTS protocol from December 2011 through May 2012 (Period 2), during which time samples were shipped frozen. Mean WBSF values were 3.4 and 4.0 kg for top loin and sirloin samples, respectively. Frequencies of steaks classified as tough were 14% and 23.5% for top loin and sirloin steaks, respectively. These findings suggest that freezing samples before shipment may influence shear force of steaks collected at the retail level. These data should be considered when evaluating beef tenderness surveys and in the design of future works.

Novel properties from experimental charge densities: an application to the zwitterionic neurotransmitter taurine.

The charge distribution of taurine (2-aminoethane-sulfonic acid) is revisited by using an orbital-based method that describes the density in a fixed molecular orbital basis with variable orbital occupation numbers. A new neutron data set is also employed to explore whether this improves the deconvolution of thermal motion and charge density. A range of molecular properties that are novel for experimentally determined charge densities are computed, including Weinhold population analysis, Mayer bond orders, and local kinetic energy densities, in addition to charge topological analysis and quantum theory of atoms-in-molecules (QTAIM) integrated properties. The ease with which a distributed multipole analysis can be performed on the fitted density matrix makes it straightforward to compute molecular moments, the lattice energy, and the electrostatic interaction energies of molecules removed from the crystal. Results are compared with high-level (QCISD) gas-phase calculations and band structure calculations employing density functional theory. Finally, the avenues available for extending the range of molecular properties that can be calculated from experimental charge densities still further using this approach are discussed.

The rapidly growing mycobacteria: saprophytes and parasites.

Rapidly growing mycobacteria are widespread saprophytes, but approximately one-third of identified species are also opportunistic pathogens in humans and animals, associated with skin, soft tissue, bone, and pulmonary infections as well as disseminated disease. Clinical and experimental evidence indicates a major role for the cell-mediated immune response in the pathogenesis of infection.

A 43-nucleotide RNA cis-acting element governs the site-specific formation of the 3' end of a poxvirus late mRNA.

The 3' ends of late mRNAs of the ati gene, encoding the major component of the A-type inclusions, are generated by endoribonucleolytic cleavage at a specific site in the primary transcript [Antczak et al., (1992), Proc. Natl. Acad. Sci. USA 89, 12033-12037]. In this study, sequence analysis of cDNAs of the 3' ends of ati mRNAs showed these mRNAs are 3' polyadenylated at the RNA cleavage site. This suggests that ati mRNA 3' end formation involves cleavage of a late transcript, with subsequent 3' polyadenylation of the 5' cleavage product. The RNA cis-acting element, the AX element, directing orientation-dependent formation of these mRNA 3' ends, was mapped to a 345-bp AluI-XbaI fragment. Deletion analyses of this fragment showed that the boundaries of the AX element are within -5 and +38 of the RNA cleavage site. Scanning mutagenesis showed that the AX element contains at least two subelements: subelement I, 5'-UUUAU downward arrowCCGAUAAUUC-3', containing the cleavage site ( downward arrow), separated from the downstream subelement II, 5'-AAUUUCGGAUUUGAAUGC-3', by a 10-nucleotide region, whose composition may be altered without effect on RNA 3' end formation. These features, which differ from those of other elements controlling RNA processing, suggest that the AX element is a component of a novel mechanism of RNA 3' end formation.

Absence of the genetic marker IS6110 from a strain of Mycobacterium tuberculosis isolated in Ontario.

A 35-year-old female patient from Waterloo, Ontario was diagnosed with pulmonary tuberculosis in June 1995. Records indicated that the patient had emigrated from Laos circa 1990. A culture grown from a bronchoalveolar lavage specimen was identified as Mycobacterium tuberculosis by standard biochemical methods. Drug-susceptibility testing indicated the strain was resistant to pyrazinamide (PZA), and a mutation was detected within pncA, a gene associated with PZA resistance. Sequence data from the 16S rRNA gene and the 16S/23S rRNA gene spacer confirmed that the strain was a member of the M tuberculosis complex, and analysis of the mpcA and pncA genes supported the identification of the strain as M tuberculosis rather than Mycobacterium bovis. However, the insertion element IS6110, which is used for epidemiological tracing of M tuberculosis, was not detected in this strain by either restriction fragment length polymorphism analysis or by polymerase chain reaction. Two other genetic markers associated with the M tuberculosis complex, IS1081 and the direct repeat element, were present. The arrival of immigrants with tuberculosis from southeast Asia, where most strains of M tuberculosis lacking IS6110 have been traced, has important implications for epidemiological studies of tuberculosis in North America.

Analysis of genetic polymorphism in the phospholipase region of Mycobacterium tuberculosis.

mtp40 was originally identified as a short genomic region that was found in strains of Mycobacterium tuberculosis but not in Mycobacterium bovis. Subsequent studies have revealed that the sequence is part of the mpcA gene, which encodes a phospholipase C. To investigate further the distribution of the mtp40 sequence, we analyzed strains of the M. tuberculosis complex by PCR and were able to amplify the mtp40 sequence in 90 of 94 strains of M. tuberculosis and in 2 strains of Mycobacterium microti but not in M. bovis or M. bovis BCG. Based on this, we developed a dot blot assay using genomic DNA which allows M. bovis to be distinguished from the majority of M. tuberculosis strains. We also probed Southern blots of 140 clinical isolates of M. tuberculosis to determine the frequency of strains lacking mtp40. This revealed an unexpected polymorphism in the phospholipase region. Two fragments were detected in 57% of samples. The expected fragment of 0.75 kbp corresponds to the region of mpcA containing mtp40. A 2.1-kbp fragment was observed to belong to a recently discovered second phospholipase gene, mpcB. In addition, some strains appeared to lack both genes, while others showed only the presence of mpcA. A few strains had additional bands, suggesting the existence of other homologs to the two phospholipase genes. We also detected the insertion of IS6110 in the mpcA coding region of one strain. The absence of these genes in some clinical isolates raises questions about their function during infection and in the development of tuberculosis disease in humans.

Genetic identification of Mycobacterium bovis BCG by restriction fragment length polymorphism analysis of the direct-repeat region.

Restriction fragment length polymorphism (RFLP) analysis was performed on the direct repeat (DR) regions of 14 strains of Mycobacterium bovis BCG. With AluI-digested DNA, BCG Japanese, Russian, and Mexican had differing RFLP patterns but 11 strains, including Pasteur, Glaxo, and Tice, had an identical pattern not detected in over 60 strains of the M. tuberculosis complex. DR analysis can aid in confirming the identification of clinical BCG isolates.

Molecular and electronic structures of two (anticonvulsant) diphenylhydantoin derivatives.

1-Benzyl-5,5-diphenyl-2,4-dioxo-3-imidazolidineacetic acid (AC), M(r) = 400.434, triclinic, P1, a = 8.7640 (3), b = 11.112 (1), c = 11.323 (2) A, alpha = 102.10 (2), beta = 95.44 (5), gamma = 109.12 (1)., V = 1002.65 (30) A3, Z = 2, Dx = 1.33 g cm-3, F(000) = 420, mu(Mo K alpha) = 0.852 cm-1, T = 293 K, R = 0.066 for 5551 unique observed reflections. The compound crystallizes from ethanol at room temperature in the form of colourless prism. 3-(2,4-Dichlorobenzyl)-5,5-diphenyl-2,4-dioxo-1-imidazolidineacetic++ + acid (AD), M(r) = 469.323, monoclinic, P2(1), a = 8.0399 (7), b = 9.7237 (6), c = 26.9768 (12) A, beta = 94.281 (4)., V = 2102.92 (96) A3, Z = 4, Dx = 1.48 g cm-3, F(000) = 968, mu(Mo K alpha) = 3.417 cm-1, T = 293 K, R = 0.066 for 5677 unique observed reflections. The compound crystallizes from ethanol at room temperature in the form of colourless prisms. Two approaches are employed in trying to understand the known differences in pharmacological activity: an analysis of the molecular geometries, and electronic structure calculations. A detailed analysis is made of the molecular geometries both from the X-ray diffraction results, and following energy minimization with molecular mechanics. The ab initio calculations employ the energy-minimized conformations. Several electronic properties are intercompared for AC, AD and their common parent molecule diphenylhydantoin (DPH). The analyses of geometry and electronic structure indicate dissimilarities between active and inactive compounds which may be linked to differences in the activity.

Unusual form of endobronchial Aspergillosis in a patient with cystic fibrosis.

The isolation of Aspergillus fumigatus from airway secretions from patients with cystic fibrosis (CF) is common and usually denotes asymptomatic colonization or allergic broncho-pulmonary aspergillosis (ABPA). A 12-year-old boy with CF acutely developed moderately severe symptoms of unremitting cough, fever, dyspnea, weight loss, and cyanosis. Chest radiographs demonstrated widespread unilateral infiltrates and volume loss. By bronchoscopy tenacious mucous plugs were seen occluding the left lower lobe bronchus. Cultures from sputum and sequential bronchoalveolar lavage grew Aspergillus fumigatus, but other significant criteria for diagnosing ABPA were lacking. No improvement was seen with a 3 week course of systemic corticosteroid and antibiotic therapy. Treatment with amphotericin B and short-term mechanical ventilation resulted in rapid resolution of all symptoms. This form of endobronchial aspergillosis has not been described previously.

A constitutively expressed vaccinia gene encodes a 42-kDa glycoprotein related to complement control factors that forms part of the extracellular virus envelope.

Nucleotide sequence analysis of a 42-kb region of the vaccinia virus (strain Western Reserve) genome identified a gene with the potential to encode a 35.1-kDa polypeptide with properties of a membrane glycoprotein (Smith et al., J. Gen. Virol. 72, 1349-1376, 1991). The 317 amino acid open reading frame (ORF) has similarity with complement control proteins and a secretory vaccinia virus protein (C28K) which interferes with complement function. The predicted B5R gene product differs from the latter protein in that it contains a C-terminal hydrophobic sequence and may be membrane-associated rather than secretory. Transcriptional mapping by Northern blotting and S1 nuclease protection showed that the gene is transcribed both early and late during infection, with the early RNA start site located 60 bp upstream of the late start site that is present at -9 to -5 bp relative to the ORF. Nevertheless, translation of early and late mRNAs are predicted to produce the same polypeptide. A rabbit antiserum was raised to the predicted external hydrophilic domain of B5R expressed in Escherichia coli and used to immunoprecipitate a M(r) 42 K protein from vaccinia-infected cells. This protein was synthesized throughout infection, with a peak from 6 to 7 hr, and its production was inhibited by tunicamycin but not monensin. Western blotting of proteins from purified extracellular enveloped virus (EEV) or intracellular naked virus with anti-B5R serum showed that this M(r) 42 K protein and two higher molecular weight forms (Mr82 and 87 K) were present only in EEV. Anti-B5R serum inhibited comet formation by the IHD-J strain of virus on RK13 cells. B5R is the third vaccinia gene shown to encode an EEV glycoprotein, the others being the virus hemagglutinin gene, and gene SalL4R which encodes a group of lectin-like glycoproteins of M(r) 22-24 K.

Nucleotide sequence of 42 kbp of vaccinia virus strain WR from near the right inverted terminal repeat.

The nucleotide sequence of 42090 bp of vaccinia virus strain WR is presented. The sequence includes the SalI L, F, G and I fragments and starts near the centre of the HindIII A fragment and extends rightwards towards the genomic terminus, finishing approximately 0.5 kb internal of the inverted terminal repeat (ITR). Translation of this region has identified 65 open reading frames (ORFs) of greater than 65 amino acids in length. Fifty-one of these which do not extensively overlap other larger ORFs have been subjected to further analysis; the other 14 are termed minor ORFs. In the rightmost 28.7 kb, the genes are, with one exception, transcribed towards the genomic terminus, similar to the arrangement of genes at the left end of the virus genome. Internal of this region the genes are expressed off either DNA strand but still predominately rightwards. ORFs are tightly packed with few intergenic non-coding regions of greater than 250 bp. Protein sequence comparisons have established a remarkably high number of homologies with entries in existing protein databases. Of these, DNA ligase, thymidylate kinase, two serine-threonine protein kinases, two serine proteinase inhibitors (serpins), two interleukin-1 receptor homologous and a discontinuous ORF related to tumour necrosis factor receptor have been reported. Other homologies include lectins, profilin, 3 beta-hydroxy steroid dehydrogenase, superoxide dismutase, guanylate kinase, ankyrin and complement factor H. In addition, there are a number of polypeptides with predicted properties of membrane-associated, secretory or glyco-proteins. Twelve gene families are described here and elsewhere. There is considerable similarity between genes from the right and left end of the virus genome that may have arisen by terminal transposition events. Several differences from the corresponding region of vaccinia virus strain Copenhagen sequence are noted. Near the right terminus the sequences diverge completely, and internal of this there are multiple examples of deletion of short sequences (eight to 10 nucleotides) that lie within penta- or hexanucleotide direct repeats.

Vaccinia virus homologues of the Shope fibroma virus inverted terminal repeat proteins and a discontinuous ORF related to the tumor necrosis factor receptor family.

Nucleotide sequencing data from a region extending 35 kb inward from the right inverted terminal repeat (ITR) of the vaccinia virus (VV) genome established the presence of VV homologues of the Shope fibroma virus (SFV) ITR proteins. The nucleotide sequences, comprising a total of 8.6 kb, and the amino acid translations for nine predicted open reading frames (ORFs) (designated SalF4L, SalF 19R, SalF21R, B4R, B8R, B9R, B10R, and B14R) are presented. Eight of the nine VV genes and all the SFV ORFs are transcribed towards their genomic termini. However, the relative positions of the VV genes (genus Orthopoxvirus) are different than those of the corresponding ORFs in SFV (genus Leporipoxvirus), indicating complex rearrangements of DNA in the genome of one or both of these viruses subsequent to their divergence from a common ancestor. Several other features of the VV ORFs were noted. SalF4L, B7R, B8R, and B9R have hydrophobic amino-terminal signal sequences but lack discernible membrane anchor domains suggesting that the proteins may be secreted. VV ORF SalF19R has a single cysteine-rich region homologous to the multiple domains of nerve growth factor receptor (NGFR), CD40, OX40 (a glycoprotein from the surface of activated murine T lymphocytes), and the recently described tumor necrosis factor receptors. Just downstream of the ORF SalF19R and in a different reading frame, there are another two related cysteine-rich domains, indicating that SalF19R was once a larger gene. B4R has homology to the host range gene of cowpox virus and to related genes near the opposite end of the vaccinia virus genome, and contains regions homologous to the repeat domains of erythrocyte ankyrin. In addition, several of the VV ORFs have homology to ORFs from near the opposite end of the VV genome, thus increasing the number of known VV gene families.

Two early vaccinia virus genes encode polypeptides related to protein kinases.

Vaccinia virus particles contain a protein kinase with an Mr of 62K calculated from sedimentation rate. We have sequenced the SalI G restriction fragment of the vaccinia virus genome near to the right inverted terminal repeat and have identified two genes which share 36% amino acid identity with each other and are related to the family of protein kinase genes. One gene, designated B1R, encodes a 34.2K protein which shares 27% identity with a protein kinase encoded by the herpes simplex virus type 1 US3 gene and contains conserved motifs characteristic of protein kinases of serine/threonine specificity. The second gene, B12R, encodes a protein of 33.3K which is poorly related to known protein kinases and lacks specific amino acids at several highly conserved key positions. The deduced partial amino acid sequence of a gene in the corresponding region of the cowpox virus genome is identical to B12R except for one conservative amino acid substitution. Both of the vaccinia virus genes are transcribed towards the right-hand end of the genome early during infection. It is possible that the product of either or both of these genes associates to form a homo- or heterodimer that represents the 62K virion-associated protein kinase.

Vaccinia virus encodes a family of genes with homology to serine proteinase inhibitors.

Nucleotide sequencing of a region of the vaccinia virus genome proximal to the right inverted terminal repeat (ITR) identified two open reading frames (ORFs) encoding proteins of 39K and 40K with amino acid homology to each other, to another vaccinia virus gene near the opposite end of the virus genome and to the superfamily of serine proteinase inhibitors (serpins). Serpins have now been found in poxviruses from the genera orthopox (cowpox and vaccinia viruses), leporipox (myxoma virus) and avipox (fowlpox virus). One of the vaccinia virus serpins identified here (B13R) shares 92% amino acid identity with the serpin from cowpox virus and 46% and 19% identity with vaccinia serpins B24R and K2L, respectively. The amino acid sequence of B13R reported here differs at 11 positions from a recently reported sequence and contains an additional three internal residues. The serpin genes near the right ITR are separated by 8 kb of DNA. Both genes contain early transcriptional termination signals just downstream of the ORFs and are transcribed in a rightward direction towards the end of the genome. Analysis of mRNAs from virus-infected cells demonstrated that all three vaccinia virus serpin genes are transcribed early during infection. The amino acid sequences at the active sites of these serpins suggest that they may inhibit serine proteinases of differing biochemical specificities. The possible functions of these genes are discussed.