Genotype by environment interactions for behavioral reactivity in sheep.

In sheep, social reactivity and reactivity to humans are relevant behavioral responses that are used to investigate the behavioral adaptation of farm animals to various rearing conditions. Such traits were previously reported as heritable and associated with several QTLs. However, few behavior-related genotype by environment (G × E) interactions have been reported to date. The experiment was performed on 2,989 male and female lambs issued from 30 sires. Every sire had progeny reared under both intensive and extensive conditions. After weaning, all lambs were individually exposed to two standardized behavioral tests. A broad range of behaviors including vocalizations, locomotion, localization, vigilance, and flight distance were assessed. Two complementary statistic approaches, with and without assumptions on the biological significance of behaviors, were performed to investigate social reactivity and reactivity to humans. G × E interactions were investigated based on the genetic correlations estimated for each factor or trait between farming conditions; those significantly different from 1 indicating a G × E. Environmental effects showed that social reactivity and reactivity to humans were higher in intensively reared lambs. The heritability of factors or traits used to measure social reactivity and reactivity to humans was similar in both rearing conditions. Estimated heritabilities were high for vocalizations in response to social isolation, moderate for locomotion and vigilance in response to social isolation, and low for both flight distance to an approaching human and proximity to a motionless human. No significant G × E interaction was found for vocalizations. G × E interactions were found for locomotion, vigilance and flight distance. Genetic correlations between both environments were low to moderate for vigilance, locomotion and flight distance. Vocalization in response to social isolation with or without human presence was identified as a robust trait and could be used to improve sheep sociability, independently of the environment. A G × E interaction was observed for behavioral reactivity to humans. Although moderate, the genetic correlation for this trait between intensive and extensive conditions could be used to select sires in the same environment by taking into account the G × E and to produce in different environments progenies that are less reactive to humans.

Importance of birthcoat for lamb survival and growth in the Romane sheep breed extensively managed on rangelands.

The Romane sheep breed proved to be adapted to harsh conditions with high prolificacy and lamb survival in outdoor farming, even under bad climate conditions. This breed shows large variability in its fleece type at birth and it has been suggested that lamb survival could be related to birthcoat type. The aim of the present study was to: i) characterize the coat of the lamb at birth and quantify lamb survival in relation to the birthcoat type and its protective properties concerning heat loss in the Romane breed raised under permanent exposure outdoors from birth, and ii) estimate genetic parameters of birthcoat type in relation to lamb survival and live body weight. A total of 7,880 lambs from 104 sires and 1,664 dams were used in a 14-yr experiment. The pedigree file included 9,625 individuals over 15 generations. Birthcoat type, coat surface temperature, coat depth, lamb survival, and growth were measured from birth to weaning. Weather data (temperature, wind speed, relative humidity, and precipitation) were recorded daily during lambing time. Two types of coats were observed at birth: hairy coat (62.9% of lambs) with a long coat depth (average 23.3 mm) or woolly 1 (37.1% of lambs) with a short coat depth (average 8.3 mm). Birthcoat type was an important factor affecting lamb survival and growth from birth in the Romane breed. Total mortality rate was significantly less in hairy-bearing coat lambs than in short-woolly coat ones: 7.0% vs. 9.6%, 11.6 % vs. 14.8%, and 15.7 % vs. 20.1 % at 2, 10, and 50 d, respectively, and the relative risks of death increased by 37%, 67%, and 46 % at 2, 10, and 50 d of age, respectively, in short-woolly lambs. At birth, a significant lower coat surface temperature, indicating less heat loss, was observed in long-hairy coat lambs compared with others (21.1°C vs. 26.1°C). Heavier body weights and better growth performances up to the age of 50 d were observed in long-hairy-bearing coat lambs. Lamb survival was positively correlated to weather conditions at lambing time in short-woolly lambs, whereas no relation was observed in hairy-coat lambs. Heritability estimate of birthcoat type (hairy or woolly) is high (0.88). Genetic correlations between birthcoat type, lamb survival, and live body weight at birth were positive and moderate, suggesting that it would be beneficial to take birthcoat into account as an adaptive trait in any genetic strategies for sheep production in harsh conditions on rangelands.

A genome scan for QTL affecting resistance to Haemonchus contortus in sheep.

Gastrointestinal nematodes are one of the main health issues in sheep breeding. To identify loci affecting the resistance to Haemonchus contortus, a genome scan was carried out using 1,275 Romane × Martinik Black Belly backcross lambs. The entire population was challenged with Haemonchus contortus in 2 consecutive experimental infections, and fecal egg counts (FEC) and packed cell volumes were measured. A subgroup of 332 lambs with extreme FEC was necropsied to determine the total worm burden, length of female worms, sex ratio in the worm population, abomasal pH, and serum and mucosal G immunoglobulins (IgG) responses. Pepsinogen concentration was measured in another subset of 229 lambs. For QTL detection, 160 microsatellite markers were used as well as the Illumina OvineSNP50 BeadChip that provided 42,469 SNP markers after quality control. Linkage, association, and joint linkage and association analyses were performed with the QTLMAP software. Linkage disequilibrium (LD) was estimated within each pure breed, and association analyses were carried out either considering or not the breed origin of the haplotypes. Four QTL regions on sheep chromosomes (OAR)5, 12, 13, and 21 were identified as key players among many other QTL with small to moderate effects. A QTL on OAR21 affecting pepsinogen concentration exactly matched the pepsinogen (PGA5) locus. A 10-Mbp region affecting FEC after the 1st and 2nd infections was found on OAR12. The SNP markers outperformed microsatellites in the linkage analysis. Taking advantage of the LD helped to refine the locations of the QTL mapped on OAR5 and 13.

Quantitative trait loci for resistance to infection in sheep using a live Salmonella Abortusovis vaccine.

Quantitative trait loci (QTL) mapping for susceptibility to a Salmonella Abortusovis vaccinal strain was performed using an experimental design involving 30 Romane sheep sire families (1216 progenies). Nine QTL corresponding to bacterial load, weight variations and antibody response criteria were mapped on eight chromosomes, including the major histocompatibility complex area on chromosome 20. Surprisingly, none was found to be significant in the SLC11A1 region (formerly NRAMP1) that has been shown to influence Salmonella susceptibility in other species.

Immune response to Haemonchus contortus infection in susceptible (INRA 401) and resistant (Barbados Black Belly) breeds of lambs.

The immune responses to Haemonchus contortus were compared in studies in resistant Barbados Black Belly (BBB) and susceptible INRA 401 (INRA) breeds of lambs. The cytokine patterns indicated a Th2-biased response in both breeds. A more persistent and elevated Th2 cytokine mRNA transcription and blood eosinophilia were noted in the BBB lambs. However, at days 4 and 30 post-infection, abomasal recruitment of eosinophils and mast cells were similar between the two breeds. Following primary infections, the BBB demonstrated a substantially lower faecal egg count compared to the INRA lambs. Furthermore, worm counts at 4 and 30 days post-infection, and adult female worm size and in utero egg counts 30 days after the first infection were significantly lower in the BBB than in the INRA breed. In the INRA breed, re-infection caused a significant reduction in most parasitological parameters compared with those observed after the primary infection. A similar response was not observed in the BBB sheep. In conclusion, while the major driving force in the response to H. contortus infection is a Th2-biased immunity in which the BBB showed its maximal performance during the primary infection, the INRA breed performed better after re-infection compared to its response to first exposure.

Proteomic analysis of ovine muscle hypertrophy.

Two-dimensional electrophoresis was used to investigate the effects of a QTL for muscle hypertrophy on sarcoplasmic protein expression in ovine muscles. In the Belgian Texel breed, the QTL for muscle hypertrophy is localized in the myostatin-encoding gene. Based on microsatellite markers flanking the myostatin gene, we compared the hypertrophied genotype with the normal genotype. The average age of the sheep was 3 mo. Among the 4 muscles studied, in the hypertrophied genotype only the vastus medialis was normal, whereas the semimembranosus, tensor fasciae latae, and LM were hypertrophied. In the hypertrophied genotype, these muscles showed upregulation of enzymes involved in glycolytic metabolism together with oxidative metabolism in LM. Certain chaperone proteins, including glutathione S-transferase-Pi, heat shock protein-27, and heat shock cognate-70, were also more highly expressed, probably due to increased use of energetic pathways. Expression of the iron transport protein transferrin was increased. Alpha-1-antitrypsin was the only protein showing a similar pattern of expression (i.e., less expressed) in all 4 muscles of the hypertrophied genotype. It is suggested that transferrin and alpha-1-antitrypsin may interact to reinforce myogenic proliferative signaling.

Polymorphic microRNA-target interactions: a novel source of phenotypic variation.

Studying the muscular hypertrophy of Texel sheep by forward genetics, we have identified an A-to-G transition in the 3'UTR of the GDF8 gene that reveals an illegitimate target site for microRNAs miR-1 and miR-206 that are highly expressed in skeletal muscle. This causes the down-regulation of this muscle-specific chalone and hence contributes to the muscular hypertrophy of Texel sheep. We demonstrate that polymorphisms which alter the content of putative miRNA target sites are common in human and mice, and provide evidence that both conserved and nonconserved target sites are selectively constrained. We speculate that these polymorphisms might be important mediators of phenotypic variation including disease. To facilitate studies along those lines, we have constructed a database (www.patrocles.org) listing putative polymorphic microRNA-target interactions.

Effects of a quantitative trait locus for muscle hypertrophy from Belgian Texel sheep on carcass conformation and muscularity.

A QTL for muscle hypertrophy has been identified in the Belgian Texel breed. A population of F2 and backcross lambs created from crosses of Belgian Texel rams with Romanov ewes was studied. Effects on carcass traits and muscle development of the Belgian Texel breed polygenes and Belgian Texel single QTL were compared. In both cases, carcass conformation and muscularity were improved. The Texel polygenic environment improved conformation mainly through changes in skeletal frame shape. Segments were shorter and bone weight lower. Muscles were more compact, shorter, and thicker. The single QTL affected muscle development. Thickness and weight of muscles were increased. Composition in myosin changed toward an increase of fast contractile type. The relative contribution of hind limb joint to carcass weight was increased. Differences in skeletal frame morphology among the three genotypes of the single QTL were small. Conformation scoring was mainly influenced by leg muscularity. Back and shoulder muscle development, which largely contributed to variability of muscularity, were less involved in the conformation scoring. Lastly, the QTL explains a small part of differences between these Belgian Texel and Romanov breeds for conformation or muscle development. A large part of genetic variability remains to be explored.

High genetic correlation between resistance to Haemonchus contortus and to Trichostrongylus colubriformis in INRA 401 sheep.

To compare the protection acquired by infection with Haemonchus contortus or Trichostrongylus colubriformis, successive cross infections with the two species were conducted in two genetically similar groups of 150 ewe lambs from 30 sires of INRA 401 breed. At 6 and 7.5 months of age, each group received two doses of 10000 L3 separated by a treatment of one of the species, followed by a similar infection with the second species at 9 and 11.5 months. Faecal egg counts (FECs) were done 28 and 35 days after each dose. There were very high genetic correlation within and between species suggesting that the environmental conditions were very similar. The heritability of FEC of H. contortus ranged from 0.39 to 0.48 and the genetic correlations between FEC after the first and second infections with the same or different species were all near 1. Selection against this species could be done on the response to only one dose. For T. colubriformis, only the response to challenge infection had a similar heritability (0.47); the genetic correlations within and among species were also near one. These results will assist the selection of sheep resistant to nematode parasites.

Impact of alcohol education and training on general practitioners' diagnostic and management skills: findings from a World Health Organization collaborative study.

OBJECTIVE: The potential of general practitioners (GPs) to reduce the prevalence of alcohol-related problems via alcohol intervention contrasts sharply with actual practice. One explanation for GPs' limited involvement in alcohol intervention is that they have had inadequate training or continuing medical education (CME) on alcohol-related issues. This study examined GPs' experience of alcohol-related CME and its possible relationship with attitudes and practice behavior regarding alcohol-related problems. METHOD: A questionnaire-based survey was returned by 2,139 GPs from 13 countries across Western and Eastern Europe. North America and Australasia. Diagnostic and management skills were assessed by responses to standardized case vignettes. RESULTS: The survey response rate was 54%. Approximately one third of GPs (32%) reported receiving no alcohol-related CME, while 8% could not remember whether or not they had received any such training or education. Of the remaining GPs (n = 1,217), 23% reported less than 4 hours (low levels) and 37% reported 4 or more hours (high levels) of alcohol-related CME. GPs who reported higher levels of alcohol-related CME were more likely to obtain information about alcohol, were more prepared to counsel problem drinkers and managed more patients for alcohol issues than did colleagues reporting lower levels of CME. Those with greater CME experience were also more confident about their ability to alleviate alcohol-related problems and reported more appropriate management strategies than did GPs with less CME experience. CONCLUSIONS: Greater exposure to alcohol-related CME appears to result in better diagnosis and more appropriate management of alcohol-related problems by GPs. Whether this is directly due to the CME itself or because GPs with greater interest in alcohol issues seek out such experience cannot be ascertained from the current study and requires further investigation.

Simulation analysis to test the influence of model adequacy and data structure on the estimation of genetic parameters for traits with direct and maternal effects.

Simulations were used to study the influence of model adequacy and data structure on the estimation of genetic parameters for traits governed by direct and maternal effects. To test model adequacy, several data sets were simulated according to different underlying genetic assumptions and analysed by comparing the correct and incorrect models. Results showed that omission of one of the random effects leads to an incorrect decomposition of the other components. If maternal genetic effects exist but are neglected, direct heritability is overestimated, and sometimes more than double. The bias depends on the value of the genetic correlation between direct and maternal effects. To study the influence of data structure on the estimation of genetic parameters, several populations were simulated, with different degrees of known paternity and different levels of genetic connectedness between flocks. Results showed that the lack of connectedness affects estimates when flocks have different genetic means because no distinction can be made between genetic and environmental differences between flocks. In this case, direct and maternal heritabilities are under-estimated, whereas maternal environmental effects are overestimated. The insufficiency of pedigree leads to biased estimates of genetic parameters.

Assessment of genetic variability of resistance to gastrointestinal nematode parasites in Creole goats in the humid tropics.

Goat production is widespread in the tropics. Goats are very susceptible to gastrointestinal nematode infection, but there is less evidence of their genetic resistance. Genetic resistance of Creole goats to gastrointestinal nematodes has been studied at Guadeloupe in the French West Indies since 1995. The objective of this research was to investigate genetic variation for resistance to gastrointestinal nematode infection, in order to introduce this trait into breeding schemes. Genetic variability was assessed within a Creole experimental flock. Forty-nine sire groups were characterized at weaning and 55 during fattening after weaning. Kids were naturally infected, mainly by Haemonchus contortus and Trichostrongylus colubriformis. Fecal egg counts were determined once at weaning and every 6 and 7 wk after drenching during fattening. Blood samples were collected every 7 wk during fattening for determination of packed cell volume. Live weights were recorded at weaning and at the beginning and middle of every infection period during fattening. Genetic parameters were estimated using the REML for multivariate animal models. The heritability estimate for transformed fecal egg count was 0.37+/-0.06 at weaning. During fattening, it increased from 0.14+/-0.05 at 4 mo to 0.33+/-0.06 at 10 mo. Heritabilities of packed cell volume ranged from 0.10 to 0.33. At weaning, maternal heritability of fecal egg count reached 0.26 and direct heritability 0.20. After 6 mo of age, maternal effects were found to be unimportant for fecal egg count and packed cell volume. Live weights presented significant genetic variability. Genetic relationships between fecal egg counts and live weight in infected pastures were never significant. Genetic correlations between packed cell volume and live weight decreased from 0.47 to 0.10 from weaning to 10 mo of age. These results demonstrated the feasibility of breeding for improved resistance to nematodes in Creole kids.

Genetic resistance to gastrointestinal nematode parasites in Polish long-wool sheep.

A study was undertaken from 1991 to 1994 on a farm in southern Poland to evaluate the genetic parameters of resistance to gastrointestinal nematodes. The predominant species were Teladorsagia circumcincta and Haemonchus contortus. A total of 32 sires were evaluated, around 15 per year. Faecal egg counts were measured twice during the 4-month grazing season for lambs (total 659 lambs) and three times for their mothers (total 327 ewes). Infection levels were high during the first 2 years and low during the last 2 years. Using an animal model, the heritability of log10(epg+25) increased from 0.20 in August to 0.33 in September for lambs, and from 0.18 in May to 0.25 in September for ewes. The repeatability of ewe faecal egg count between years was 0.25. A genetic correlation of 0.58 was found between faecal egg count in ewes and in 6-7-month-old lambs. A negative genetic correlation (-0.61) was estimated between faecal egg count in September and daily weight gain of lambs from 70 days of age to the end of grazing season (7 months of age). The results confirm the feasibility of genetic selection of sheep for resistance to nematode parasites in an environment where T. circumcincta and H. contortus are the dominant species.

[Fragile X syndrome is still unrecognized: efficacy of molecular diagnosis in mentally retarded probands]. / Le syndrome X fragile est encore méconnu: efficacité du diagnostic moléculaire chez les proposants avec retard mental.

BACKGROUND: The fragile X mental retardation syndrome is the most common cause of inherited mental retardation. Identification of the unstable mutation responsible for the disease has allowed the design of a fully reliable molecular test for the diagnosis of the disease and for genetic counselling (identification of clinically normal carriers and prenatal diagnosis). We started in July 1991 to search for the mutation in mentally retarded probands, with no known cause for their phenotype. We present the results of a 42-month experience. POPULATION AND METHODS: One thousand and one hundred fourty-nine probands were analysed. In case of a positive diagnosis, an extension of the molecular study to relatives was proposed. DNA samples were studied by Southern blot following EcoRI or EcoRI + EagI digestion. Clinical data were collected from referring clinicians. RESULTS: Seventy-three carriers of a full mutation were identified, belonging to 52 families. The mean age of the fragile X probands was 16 +/- 14 years, which is very surprising for a disease that causes significant manifestations by the age of 2 to 3 years. This indicates an insufficient knowledge about this disease in France. Most of the demands for the test were from clinical geneticists. This diagnosis is of major importance for genetic counselling, as illustrated by the following study of 108 women at risk in these families. CONCLUSIONS: The importance of an early diagnosis followed by an extended family study, for carrier screening and prevention of this severe disease, justifies molecular testing on any child with mental retardation or significant language delay of unknown cause, in the absence of clinical signs formally excluding a fragile X diagnosis.

Expansion and methylation status at FRAXE can be detected on EcoRI blots used for FRAXA diagnosis: analysis of four FRAXE families with mild mental retardation in males.

The original test for the analysis of the CCG expansion at the FRAXE locus involves Southern blot analysis of HindIII digests. We show that, by using a different probe, the FRAXE mutation can be detected easily on the same EcoRI or EagI+EcoRI blots as are used for detection of FRAXA. Unexpectedly, we found that both the expansion and methylation status can be determined on a single EcoRI digest, because of the presence of a methylation-sensitive EcoRI site very close to the CCG repeat. We thus detected in a series of mentally retarded individuals previously tested for FRAXA expansion a FRAXE proband who led to the identification of a large sibship (7 of 10 children carrying a mutation). We also show that two fragile X families without FRAXA mutation that previously have been described by Oberlé et al. have the FRAXE expansion. In another family also ascertained initially by cytogenetic finding of a fragile X site, we performed the combined cytogenetic and molecular prenatal diagnosis of a mutated male fetus. All nine males (>3 years old) in whom we found a methylated mutation had mild mental retardation. Our results suggest that the threshold of repeat length for abnormal methylation and fragile-site expression may be smaller at FRAXE than at FRAXA.

Genetic variability in resistance of Creole goats to natural infection with trichostrongylids in Guadeloupe.

The objective of this study was to show the existence of genetic variability in resistance of Creole goats to natural infection with intestinal nematodes. Four successive cohorts of male and female kids were reared from weaning during 9 months in intensive pangola pastures (stocking rate 1.4 t/ha, regrowth of 21 days, 300 kg N/ ha/year of fertilization). All cohorts considered, 203 offsprings from 13 bucks and 149 goats were used in the experiment. Animals were treated with an oral dose of ivermectin every seven weeks, from 3 to 11 months of age. They were sampled for fecal egg counts (FEC) and packed cell volume (PCV), 6 weeks after drenching, when 6 and 10 months old. Haemonchus contortus and Trichostrongylus colubriformis were the predominant species. Data of one cohort when 6 months of age and another one when 10 months old were not included in calculations because a self-cure occurred during a rainy season. Estimated means of FEC after square-root transformation were 1054 and 711 respectively for 6-month-old and 10-month-old kids. Phenotypic correlations between FEC and PCV were -0.46 (p < 0.001) and -0.29 (p < 0.01) when 6 and 10 months old, respectively. FEC repeatabilities estimated as variance ratio were 0.47 and 0.06 for within-age measures (data recorded at one week interval) and between age measures (6 vs. 10 months) respectively. No sire effect was significant for PCV. Sire effect on FEC was more significant when 6 months old (p < 0.03) than when 10 months old (p < 0.15). Further studies are now being carried out to confirm this genetic determinism of resistance to natural infection with strongyles in Creole goats.

Worm population characteristics and pathological changes in lambs after a single or trickle infection with Teladorsagia circumcincta.

The regulation of the worm population and of its pathological effects was studied after a single or trickle infection with T. circumcincta. One hundred and twenty lambs, 60 Romanov and 60 Mérinos d'Arles, 3 months old, were distributed in four balanced groups: non-infected (G0), infected with 7000 L3 per animal once and slaughtered after 4 weeks (G14) or 8 weeks (G18), and infected 8 times and slaughtered after 8 weeks (G88). Parasitological, histological, haematological parameters and weight gains were recorded on each animal. Female and artificially nursed lambs had lower worm burdens and egg counts (epg) than males and naturally suckled lambs. No difference in parasitological parameters was seen between the two breeds, but Mérinos lambs infected once, had a higher increase in pepsinogen concentrations than Romanov lambs. In the infected animals, a significant proliferation of mast and eosinophil cells was observed in the abomasum wall. Serum pepsinogen concentrations were significantly higher 3 weeks p.i. and the weight gain was depressed during the first month p.i. The worm population was more numerous and younger in group G14 compared with G18 in which 24% of the worms had been expelled during the second month p.i. The female worms in G18 were longer and had more eggs in utero and higher egg output. After the trickle infection (G88) the take was reduced, female worms were longer with more eggs in vagina (pars ejectrix) and there was a higher variability in the number of eggs (compared with G18 data). The pepsinogen rise was smaller but no specific effect was seen on histological and haematological parameters.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of genetic type, lactation and management on helminth infection of ewes in an intensive grazing system on irrigated pasture.

A survey of helminth infection was conducted in a flock of 290 ewes distributed into Romanov (R), Merinos d'Arles (M) and Romanov x Merinos (R x M) genetic types, grazing irrigated pasture in the south of France. Faecal egg and larval counts were done seven times per year from 1981 to 1984 on homogeneous groups of ewes and then individually once to four times every autumn from 1985 to 1988. Helminth fauna was diverse and more abundant during autumn. High levels of strongyle infection occurred in the ewes that remained on the same irrigated pastures during summer. Moving to Alpian pastures during the summer lowered autumnal infection. Significant differences between genotypes in intensity of infection were observed in the order R > R x M > M for strongyles (Teladorsagia circumcincta and/or Trichostrongylus vitrinus, Chabertia ovina and/or Oesophagostomum venulosum, Nematodirus spp.), Moniezia spp. and Dictyocaulus filaria. The hierarchy was reversed for Fasciola hepatica infection and not consistent from one year to another for protostrongylid infections. The effect of lactation intensity on the postparturient rise was studied by equilibrating number of ewes according to reproductive status (zero, one or two lambs in lactation). Merino ewes with two lambs in lactation, as well as primiparous Romanov ewes, had significantly higher strongyle infections than the others. The repeatabilities of the larval and egg counts between the four trial years were 0.24, 0.23 and 0.16, respectively, for protostrongyles, Nematodirus and strongyles, with higher intra-annual values for protostrongyles and inconsistently significant results for strongyles due to the presence of several species.

[Wilson's disease and psychiatry. Apropos of a case]. / Maladie de Wilson et psychiatrie. A propos d'un cas.

This work is based on the case of a 24 year old young man sent to a psychiatry department for behaviour troubles, hysterical demonstrations and a neurological syndrome atypical. The first part develops his familial and medical background, the clinical schemes and, finally, the biological elements leading to the Wilson's disease diagnosis. The second part is a clinical and biological summary. Comparing to our case, we can see that it is a relatively typical form. This work ends with the difficulty of determining such a diagnosis regarding a patient whose personal and familial background allowed for a consistent psychopathological explanation and who had been already treated in this sense, with the risk of neglecting the biological check up. But can this Wilson's disease diagnosis be enough to sum up all the psychiatric pathology of this patient? The question remains to be resolved in the absence of any future studies.