Bayesian analysis of parent-specific transmission ratio distortion in seven Spanish beef cattle breeds.

Transmission ratio distortion (TRD) is the departure from the expected Mendelian ratio in offspring, a poorly investigated biological phenomenon in livestock species. Given the current availability of specific parametric methods for the analysis of segregation data, this study focused on the screening of TRD in 602 402 single nucleotide polymorphisms covering all autosomal chromosomes in seven Spanish beef cattle breeds. On average, 0.13% (n = 786) and 0.01% (n = 29) of genetic markers evidenced sire- or dam-specific TRD respectively. There were no single nucleotide polymorphisms accounting for both sire- and dam-specific TRD at the same time, and only one marker (rs43147474) accounted for (sire-specific) TRD in all seven breeds. It must be noted that rs43147474 is located in the fourth intronic region of the GTP-binding protein 10 gene, and this locus has been previously linked to the maintenance of mitochondria and nucleolar architectures. Alternatively, other candidate genes surround this hot-spot for sire-specific TRD in the cattle genome, and they are related to embryonic and postnatal lethality as well as prostate cancer, among others. This research characterized the distribution of TRD in the bovine genome, highlighting heterogeneous results when comparing across breeds.

Canalization analysis of birth weight in Bruna dels Pirineus beef cattle.

Although heteroskedasticity has been a main topic of interest in beef cattle during recent decades, the current availability of canalization models provided new insights for animal breeding programs. Within this context, birth BW (BWT) was analyzed in the Bruna dels Pirineus beef cattle breed by implementing canalization models that accounted for heterogeneous residual variances due to systematic, permanent environmental effects and additive genetic effects. Analyses were performed on BWT data from 8,130 calves born in 12 commercial breeding herds contributing to the yield recording scheme of the Bruna dels Pirineus breed. Analytical models accounted for direct additive genetic, permanent environmental, and 4 systematic effects (i.e., age of the dam, sex of the calf, birth type, and herd-year-season), and the same effects were evaluated as potential sources of variation in the residual term. Their relevance was checked by the deviance information criterion (DIC), and only residual additive genetic, permanent environmental, birth type, and herd-year-season remained in the operational model, all of them originating relevant reductions in the DIC parameter. Bruna dels Pirineus calves showed a moderate heritability of 0.30 (95% high posterior density, 0.19 to 0.40) for BWT; additional additive genetic variability was revealed in the residual term, this being positively correlated with the direct additive genetic component (0.44; 95% high posterior density, 0.37 to 0.54). Genetic trends were evaluated on both sources of additive genetic variance, and relevant patterns were identified in several herds. Although this breed did not evidence a homogeneous genetic trend for the whole population, herd-specific positive and negative trends were revealed, suggesting the plausibility of genetic selection for canalization on BWT in beef cattle breeds. These results must be viewed as a contribution to the canalization research field, providing relevant information for the breeding scheme of the Bruna dels Pirineus breed, as well as important insights about the genetic background of BWT for the beef industry worldwide.

Backfat thickness and longissimus dorsi real-time ultrasound measurements in light lambs.

The aim of this study was to assess the accuracy of ultrasound measurements for predicting carcass traits in 124 Spanish pascual-type lambs (13 to 16 kg carcass weight). Ultrasound images were taken transversal and longitudinal to the vertebral column and at thoracic (TV; between 12th and 13th ribs) and lumbar (LV; between first and second lumbar vertebrae) locations. Skin thickness, subcutaneous backfat thickness (BFT), and depth (DLD), width (WLD), and area (ALD) of longissimus dorsi were obtained with ImageJ 1.42q software. After slaughter, BFT (TV, 2.30 ± 0.06 mm; LV, 2.46 ± 0.06 mm), DLD (TV, 2.47 ± 0.03 cm; LV, 2.48 ± 0.03 cm), WLD (TV, 4.50 ± 0.04 cm; LV, 4.60 ± 0.04 cm), and ALD (TV, 9.96 ± 0.12 cm(2); LV, 10.19 ± 0.13 cm(2)) were directly measured on the lamb carcass. Correlations between ultrasound and direct carcass measurements were greater than 0.61 for DLD, WLD, and ALD (P < 0.05) whereas they fluctuated between 0.32 and 0.60 for BFT (P < 0.05); moreover, correlations were significantly (P < 0.05) greater for transversal than for longitudinal views. In a similar way, linear regression analyses suggested a moderate underestimation for BFT and lumbar DLD when using real-time ultrasound technologies whereas WLD, ALD, and thoracic DLD suffered from under- and overestimation for small and large values of carcass traits, respectively. After decomposing the mean square prediction error (MSPE) for the different ultrasound measurements, we found that the error due to disturbance contributed most to the MSPE followed by the error of central tendency and the error due to regression. The SE of prediction (SEP) was also calculated as an additional precision indicator, obtaining estimates less than that in previous studies with larger lambs. In conclusion, transversal ultrasound measurements at the thoracic and lumbar levels could be a useful tool for predicting DLD, WLD, and ALD in light lambs, perhaps suffering from worse prediction properties when focusing on BFT. This information could be of special relevance for light lamb producers worldwide, with a special emphasis in the Mediterranean basin where this kind of production system accounts for a large percentage of the sheep industry.

Sources of sire-specific genetic variance for birth and weaning weight in Bruna dels Pirineus beef calves.

This research investigated two sources of sire-specific genetic effects on the birth weight (BWT) and weaning weight (WWT) of Bruna dels Pirineus beef calves. More specifically, we focused on the influence of genes located in the non-autosomal region of the Y chromosome and the contribution of paternal imprinting. Our analyses were performed on 8130 BWT and 1245 WWT records from 12 and 2 purebred herds, respectively, they being collected between years 1986 and 2010. All animals included in the study were registered in the Yield Recording Scheme of the Bruna dels Pirineus breed. Both BWT and WWT were analyzed using a univariate linear animal model, and the relevance of paternal imprinting and Y chromosome-linked effects were checked by the deviance information criterion (DIC). In addition to sire-specific and direct genetic effects, our model accounted for random permanent effects (dam and herd-year-season) and three systematic sources of variation, that is, sex of the calf (male or female), age of the dam at calving (six levels) and birth type (single or twin). Both weight traits evidenced remarkable effects from the Y chromosome, whereas paternal imprinting was only revealed in WWT. Note that differences in DIC between the preferred model and the remaining ones exceed 39 000 and 2 800 000 DIC units for BWT and WWT, respectively. It is important to highlight that Y chromosome accounted for ∼2% and ∼6% of the total phenotypic variance for BWT and WWT, respectively, and paternal imprinting accounted for ∼13% of the phenotypic variance for WWT. These results revealed two relevant sources of sire-specific genetic variability with potential contributions to the current breeding scheme of the Bruna dels Pirineus beef cattle breed; moreover, these sire-specific effects could be included in other beef cattle breeding programs or, at least, they must be considered and appropriately analyzed.

A phase II trial of short course fludarabine, mitoxantrone, rituximab followed by 9°Y-ibritumomab tiuxetan in untreated intermediate/high-risk follicular lymphoma.

BACKGROUND: A prospective, single-arm, open-label, multicenter, nonrandomised phase II trial to evaluate efficacy and safety of short fludarabine, mitoxantrone, and rituximab (FMR) induction followed by radioimmunotherapy, in untreated, intermediate/high-risk follicular non-Hodgkin's lymphoma (NHL) patients. PATIENTS AND METHODS: Fifty-five patients were treated using a sequential treatment schedule of four induction cycles of FMR chemoimmunotherapy, and a subsequent consolidating single administration of (90)Y-ibritumomab tiuxetan ((90)Y-IT), 8-14 weeks later. Patients were eligible for radioimmunotherapy if at least in partial response (PR) after induction, with normal platelet and granulocyte counts and a bone marrow infiltration ≤ 25%. Primary study end points were response rate and hematologic toxic effects; secondary end points were overall survival (OS) and progression-free survival (PFS). RESULTS: All the 55 patients received four induction cycles with an overall response rate of 96% (38 complete responses [CR] and 15 PR). Fifty-one patients (38 in CR and 13 in PR) received (90)Y-IT. By the end of the treatment, 49/55 patients achieved a CR. With a median follow-up of 21 months, the estimated 3-year PFS was 81% and the 3-year OS 100%. CONCLUSIONS: This study has established feasibility, tolerability, and efficacy of a regimen composed by short FMR induction with (90)Y-IT consolidation in untreated intermediate/high-risk follicular NHL patients.

Parametric bootstrap for testing model fitting of threshold and grouped data models: an application to the analysis of calving ease of Bruna dels Pirineus beef cattle.

The aim of this study was to compare the goodness of fit of the threshold models with homoscedasticity or heteroscedasticity and the grouped data model for the analysis of calving ease in beef cattle by using a parametric bootstrap procedure. Field data included 8,205 records of the Bruna dels Pirineus beef cattle breed in the Pyrenean mountain areas of Catalonia (Spain). The actual distribution was 81.81% of calvings without assistance, 11.02% slightly assisted by the farmer, 5.12% strongly assisted by the farmer, 0.89% assisted by the veterinarian, and 1.16% cesarean, but these percentages were very different in the different herds. This can be explained partially by the different subjective way of scoring of each farmer. Primiparous cows had a greater (P < 0.001) difficulty calving than cows with 5 or more parities (11.74 vs. 4.49% of calvings strongly assisted by the farmer or the veterinarian and 2.8 vs. 0.65% cesarean). Male calves caused greater (P < 0.001) calving difficulty than females (7.71% of male calvings strongly assisted by the farmer or the veterinarian vs. 4.25% of females and 1.83% cesarean in males vs. 0.47% in females). The month and year of calving also had a strong influence on calving ease. These data were analyzed using 3 different models: the threshold models with homoscedasticity or heteroscedasticity and the grouped data model. The bootstrap comparison among models suggested that the threshold models, even allowing for heteroscedasticity, did not fit the herd effects well. In contrast, fitting deficiencies were not observed for the grouped data model in any factor. The variance of direct effect of the calf was estimated using the 3 models, and the heritability estimate ranged from 0.165 for the grouped data model to 0.185 for the hereroscedastic threshold model. This heritability was moderate, but it would justify the inclusion of direct effects of the calf on calving ease in the breeding objective. Overall, results highlighted the flexibility of the grouped data model for the analysis of discrete traits, like calving ease of beef calves.

MACOP-B regimen in the treatment of adult Langerhans cell histiocytosis: experience on seven patients.

BACKGROUND: Adult Langerhans cell histiocytosis (LCH) is a rare disease. The combination of vinblastine and prednisone, given in a 6-month course, is the standard of care but prospective randomized trials are lacking. PATIENTS AND METHODS: We report our monocentric experience in the treatment of seven adult patients with multisystem (MS) LCH (n = 3) or single-system multifocal (SS-m) LCH (n = 4) with the short-course intensive chemotherapy regimen methotrexate, doxorubicin, cyclophosphamide, vincristine, prednisone and bleomicin (MACOP-B). RESULTS: The overall response rate was 100% [five complete response (CR), two partial response (PR)]. After a median follow-up of 6.5 years, four patients are in first continuous CR and three patients relapsed after 5, 8 and 62 months, respectively. Four patients were evaluated with positron emission tomography (PET) scan: all three PET-negative patients at the end of treatment had a long-lasting response with only one patient relapsing after 5 years. PET scan detected additional bone lesions at diagnosis in two of four patients, changing the treatment program in one of them. CONCLUSIONS: MACOP-B regimen seems to be very active in the treatment of adult MS or SS-m LCH, with long-lasting responses in five of seven patients. PET scan merits further evaluation in the initial staging and in the evaluation of the response to chemotherapy.

Gemcitabine as single agent in pretreated T-cell lymphoma patients: evaluation of the long-term outcome.

BACKGROUND: Peripheral T-cell lymphoma unspecified (PTCLU) and mycosis fungoides (MF) often show resistance to conventional chemotherapy. Gemcitabine should be considered a suitable option. We report the long-term update of 39 pretreated T-cell lymphoma patients treated with gemcitabine. PATIENTS AND METHODS: From May 1997 to September 2007, 39 pretreated MF and PTCLU patients received gemcitabine. Inclusion criteria were as follows: histologic diagnosis of MF or PTCLU; relapsed/refractory disease; age > or =18 years; and World Health Organization performance status of two or less. Nineteen patients had MF and 20 PTCLU. All patients with MF had a T3-T4, N0, and M0 disease and patients with PTCLU had stage III-IV disease. Gemcitabine was given on days 1, 8, and 15 on a 28-day schedule (1200 mg/m(2)/day) for a total of three to six cycles. RESULTS: Overall response rate was 51% (20 of 39 patients); complete response (CR) and partial response (PR) rates were 23% (9 of 39 patients) and 28% (11 of 39 patients), respectively. Patients with MF had a CR rate of 16% and a PR rate of 32% compared with a CR rate of 30% and a PR rate of 25% of PTCLU patients. Among the CR patients, 7 of 9 are in continuous complete response with a variable disease-free interval (15-120 months). CONCLUSION: In our experience, gemcitabine proved to be effective in pretreated MF and PTCLU patients, even in the long term.

A phase II trial of CHOP chemotherapy followed by yttrium 90 ibritumomab tiuxetan (Zevalin) for previously untreated elderly diffuse large B-cell lymphoma patients.

BACKGROUND: A prospective, single-arm, open-label, nonrandomized phase II combination chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP) plus radioimmunotherapy trial was conducted to evaluate the efficacy and safety in untreated elderly diffuse large B-cell lymphoma (DLBCL) patients. PATIENTS AND METHODS: From February 2005 to April 2006, in our institute we treated 20 eligible elderly (age > or =60 years) patients with previously untreated DLBCL using a novel regimen consisting of six cycles of CHOP chemotherapy followed 6-10 weeks later by (90)Y ibritumomab tiuxetan. RESULTS: The overall response rate to the entire treatment regimen was 100%, including 95% complete remission (CR) and 5% partial remission. Four (80%) of the five patients who achieved less than a CR with CHOP improved their remission status after radioimmunotherapy. With a median follow-up of 15 months, the 2-year progression-free survival was estimated to be 75%, with a 2-year overall survival of 95%. The (90)Y ibritumomab tiuxetan toxicity included grade > or =3 hematologic toxicity in 12 of 20 patients; the most common grade > or =3 toxic effects were neutropenia (12 patients) and thrombocytopenia (7 patients). Transfusions of red blood cells and/or platelets were given to one patient. CONCLUSION: This study has established the feasibility, tolerability, and efficacy of this regimen for elderly patients with DLBCL.

Early positron emission tomography (PET) restaging: a predictive final response in Hodgkin's disease patients.

BACKGROUND: It is important to distinguish between responders to standard treatment and non-responders Hodgkin's disease (HD) patients. PATIENTS AND METHODS: Between June 2003-September 2004, in our institute, 40 newly-diagnosed patients with advanced stage HD were consecutively treated with ABVD chemotherapy for six cycles. All these patients underwent staging/restaging: computed tomography (CT) and positron emission tomography (PET) at time 0, PET after two cycles, CT and PET after four and six cycles. RESULTS: After two cycles (PET-2), the PET was negative in 28/40 (70%), positive in 8/40 (20%), and minimal residual uptake (MRU) was present in the remaining four (10%) patients. After treatment, among eight patients who were PET-2+, seven showed refractory disease and one had relapse after 3 months. All four patients with MRU at the PET-2 became PET- during the further four cycles and, after treatment, three were in complete response (CR) and one relapsed after 5 months. All 28 PET negative patients at the PET-2 remained PET negative and all of them were in CR after treatment. CONCLUSIONS: The PET use for early (after two cycles) response assessment in HD patients is a significant step forward and has the potential to help physicians make crucial decisions about further treatment.

A canine model of gastroesophageal reflux disease (GERD).

BACKGROUND: Although a variety of antireflux procedures and medications are used to treat gastroesophageal reflux disease (GERD), reliable large-animal models of GERD that can be used to objectively compare the efficacy of these treatments are lacking. METHODS: Esophageal manometry and 24-h gastroesophageal pH monitoring with event data were performed in 18 mongrel dogs with a cervical esophagopexy. We then calculated a modified DeMeester score: The Duke Canine reflux score (DCR). Thereafter, the animals underwent a 4-cm anterior distal esophageal myotomy, incision of the left diaphragmatic crus, and intrathoracic gastric cardiopexy. Postoperative 24-h pH and manometry were obtained 2 weeks later. RESULTS: The postoperative 24-h pH results showed a significant increase in the mean DCR score (5.9 +/- 4.5 vs 84.9 +/- 56.1, p < 0.0002), and manometry indicated a significant decrease in mean lower esophageal sphincter (LES) pressure (7.1 +/- 2.9 vs 3.2 +/- 2.5 mmHg, p < 0.0001). CONCLUSION: This technique reliably creates a canine model of GERD.

Histologic evaluation of fetal brains following maternal pneumoperitoneum.

BACKGROUND: The purpose of this study was to determine if maternal pneumoperitoneum with carbon dioxide (CO2) produces evidence of central nervous system (CNS) injury in preterm fetal guinea pigs. METHODS: Thirty pregnant guinea pigs at gestational day (GD) 45 were assigned at random to one of three treatment groups: anesthesia only, CO2 pneumoperitoneum (5 mmHg), or laparotomy. Dams were killed 3 or 5 days postprocedure and fetal brains (83 total) harvested and fixed for subsequent histopathologic evaluation. For comparative purposes, histologic features of fetal guinea pig brain injury were defined from examination of fetal brains harvested from an additional dam that underwent laparotomy with 20 min of uterine arterial occlusion. RESULTS: Carbon dioxide pneumoperitoneum did not increase maternal/fetal morbidity. No evidence of brain injury was found in fetuses from any of the treatment groups. CONCLUSION: Carbon dioxide pneumoperitoneum at 5 mmHg for 40 min in the pregnant guinea pig does not produce evidence of fetal brain injury.

Intraperitoneal immunity and pneumoperitoneum.

BACKGROUND: Carbon dioxide (CO(2)) pneumoperitoneum has been implicated as a possible factor in depressed intraperitoneal immunity. Using in vitro functional assays, CO(2) has been shown to decrease the function of peritoneal macrophages harvested from insufflated mice. However, an effective in vivo assessment is lacking. Listeria monocytogenes (LM), an intracellular pathogen, has served as a well-established in vivo model to study cell-mediated immune responses in mice. This study examines the immune competence of mice based on their ability to clear intraperitoneally administered LM following CO(2) vs helium (He) insufflation. METHODS: Eighty-five mice (C57Bl/6, males, 4-6 weeks old) were divided between the following four treatment groups: CO(2) insufflation, He insufflation, abdominal laparotomy (Lap), and control (anesthesia only). Immediately postoperatively, each group was inoculated percutaneously and intraperitoneally with a sublethal dose (.015 x 10(6) org) of virulent LM (EGD strain). Half of the animals were killed on postoperative day 3 and half on day 5. Spleens and livers (sites of bacterial predilection) were harvested, homogenized, and plated on TSB agar. The amount of bacteria (1 x 10(6) LM/spleen and liver) from each group was then compared. Statistical significance was set at p

Cardioprotective effects of 70-kDa heat shock protein in transgenic mice.

Heat shock proteins are proposed to limit injury resulting from diverse environmental stresses, but direct metabolic evidence for such a cytoprotective function in vertebrates has been largely limited to studies of cultured cells. We generated lines of transgenic mice to express human 70-kDa heat shock protein constitutively in the myocardium. Hearts isolated from these animals demonstrated enhanced recovery of high energy phosphate stores and correction of metabolic acidosis following brief periods of global ischemia sufficient to induce sustained abnormalities of these variables in hearts from nontransgenic littermates. These data demonstrate a direct cardioprotective effect of 70-kDa heat shock protein to enhance postischemic recovery of the intact heart.

Expression of mRNAs encoding alpha and beta subunit isoforms of Na,K-ATPase in the vestibular labyrinth and endolymphatic sac of the rat.

The distribution of mRNAs coding for three different isoforms of the alpha and two of the beta subunit of Na,K-ATPase was studied in the rat vestibular system using in situ mRNA hybridization. The dark cells of the utricular macula and of the ampullae of the semicircular canals expressed high levels of mRNA encoding the alpha 1 and beta 2 isoforms of the Na,K-ATPase, a composition that in the cochlea has been uniquely found in the stria vascularis. However, in the dark cells it was coupled with a weak expression of beta 1. The sensory epithelia of the vestibular system showed alpha 1 and beta 1 expression at much higher levels than in the cochlear sensory epithelium. Weak expression limited to the alpha 1, beta 1, and beta 2 isoforms was observed in the endolymphatic sac, contrasting previous cytochemical results which suggested extensive Na,K-ATPase activity to the sac. The results support the widely held hypothesis that the vestibular dark cells play a role similar to that of the stria vascularis in endolymph production. They indicate that the ion transport requirements of the vestibular sensory epithelia may be different than those in the cochlea. They also suggest that the endolymphatic sac may not be a major site of inner ear ion exchange.

Direct application of basic fibroblast growth factor improves tympanic membrane perforation healing.

Topical application of basic fibroblast growth factor (b-FGF) on tympanic membrane (TM) perforations was studied in guinea pigs. One-millimeter simple round TM perforations or 2-mm TM perforations with medially flapped borders were performed. Either b-FGF or placebo was instilled in each ear on the day of surgery and daily thereafter. Treatment was applied either directly to the perforation or to a Gelfoam pledget over the defect. When no scaffolding material was interposed, b-FGF induced a faster healing response characterized by a hyperplastic but linear subepidermal connective tissue reaction compared to the control. When Gelfoam was interposed as a scaffold, a voluminous scar protruding into the middle ear cavity and involving the ossicles was observed in both b-FGF and control animals. Gelfoam-induced scars did not decrease after long-term observation, therefore discouraging its use.

Human heat shock protein 70 (hsp70) protects murine cells from injury during metabolic stress.

Expression of heat shock protein 70 (hsp70) is stimulated during ischemia, but its proposed cytoprotective function during metabolic stress has remained conjectural. We introduced a human hsp70 gene into mouse 10T1/2 cells and assessed the susceptibility of these cells to injury in response to conditions that mimic ischemia. Transiently transfected cells, in the absence of stress, expressed human hsp70 to levels equal to or greater than those induced by heat shock, as assessed by RNAse protection, immunoblot, and immunohistochemical analyses. By comparison to cells transfected with a control plasmid, cells expressing the human hsp70 transgene were resistant to injury induced by glucose deprivation and inhibition of mitochondrial respiration. These results provide direct evidence for a cytoprotective function of hsp70 during metabolic stress.

[Genetic markers of susceptibility to myocardial infarct]. / Geneticheskie markery predraspolozhennosti k infarktu miokarda.

The study with the two genetic markers--HLA antigens and the pattern of dermatoglyphics--provides strong evidence for the fact that there is a genetic predisposition to coronary heart disease. The dermatoglyphic pattern has been found to be a more reliable marker of predisposition to this disease than HLA antigens. The decisive rules derived from the study may be used in the assessment of a genetic risk for this cardiovascular abnormality.

Improved healing of tympanic membrane perforations with basic fibroblast growth factor.

We have investigated the effects of basic fibroblast growth factor (FGF) on the healing of tympanic membrane (TM) perforations. In the first series of experiments, a simple, round 1-mm perforation was made in the membrane and the effects of basic FGF examined. In a second series of experiments, basic FGF was tested on 2-mm perforations in which the borders were folded inward in order to delay normal healing. Topical applications of saline or basic FGF were administered onto gelfoam overlays of the TM perforations in 51 guinea pigs by delivering 5 microliters aliquots of PBS or 5 microliters of PBS containing 1 microgram of basic FGF on the day of surgery and daily thereafter. Repair of the lesions was evaluated 3, 5 or 8 days after surgery. The results show that basic FGF mediates faster healing of TM perforations by inducing rapid proliferation of the subepithelial connective tissue layer.

The nucleotide sequence and gene organization of red clover necrotic mosaic virus RNA-2.

Red clover necrotic mosaic virus, a member of the dianthovirus group, is characterized by a genome composed of two nonhomologous single-stranded RNAs of approximately 4.0 (RNA-1) and 1.4 kb (RNA-2). The complete nucleotide sequence of the RNA-2 has been determined. RNA-2 is 1448 nucleotides in length with a 5' terminal m7G cap and no 3' terminal poly-A tail or 5' terminal VPg. An open reading frame beginning at the first initiation codon at nucleotide 80 and ending at nucleotide 1030 has been identified which can encode a polypeptide of 35 kDa. RNA-2 directs the synthesis of a 35 kDa polypeptide in vitro. SP6 and T7 transcripts from full length RNA-2 cDNA clones directed the synthesis of a polypeptide with the same electrophoretic mobility as the polypeptide directed from authentic RNA-2. Clones with various 3' terminal deletions both outside and within the 35 kDa open reading frame were transcribed and translated in vitro to define the limits of the 35 kDa open reading frame. A second, small open reading frame capable of encoding a polypeptide of 4.9 kDa was also indicated from the sequence; however, there was no evidence for a protein product of that size. RNA-2 is presumed to be monocistronic and encode a cell-to-cell movement function. A small but significant amino acid sequence homology was observed with the brome mosaic virus RNA-3a polypeptide.