Variation in volatile organic compounds in Atlantic salmon mucus is associated with resistance to salmon lice infection.

Salmon lice are ectoparasites that threaten wild and farmed salmonids. Artificial selection of salmon for resistance to the infectious copepodid lice stage currently relies on in vivo challenge trials on thousands of salmon a year. We challenged 5750 salmon with salmon lice (Lepeophtheirus salmonis) from two distinct farmed strains of salmon in two separate trials. We found that volatile organic compounds (VOC), 1-penten-3-ol, 1-octen-3-ol and 6-methyl-5-hepten-2-one in the mucus of the salmon host after salmon lice infection, were significantly associated with lice infection numbers across a range of water temperatures (5 °C, 10 °C, 17 °C). Some VOCs (benzene, 1-octen-3-ol and 3,5,5-trimethyl-2-hexene) were significantly different between lines divergently selected for salmon lice resistance. In a combined population assessment, selected VOCs varied between families in the range of 47- 59% indicating a genetic component and were positively correlated to the salmon hosts estimated breeding values 0.59-0.74. Mucosal VOC phenotypes could supplement current breeding practices and have the potential to be a more direct and ethical proxy for salmon lice resistance provided they can be measured prior to lice infestation.

Quantitative trait loci and genes associated with salmonid alphavirus load in Atlantic salmon: implications for pancreas disease resistance and tolerance.

Salmonid alphavirus infection results in pancreas disease causing severe economic losses for Atlantic salmon aquaculture. Knowledge about genes and pathways contributing to resistance is limited. A 54 K SNP panel was used to genotype 10 full-sibling families each consisting of ~ 110 offspring challenged with salmonid alphavirus subtype 3. Relative heart viral load was assessed at 4- and 10-weeks post-infection using quantitative PCR. A moderate genomic heritability of viral load at 4 weeks (0.15-0.21) and a high positive correlation with survival (0.91-0.98) were detected. Positions of QTL detected on chromosome 3 matched those for survival detected by other studies. The SNP of highest significance occurred in the 3' untranslated region of gig1, a fish-specific antiviral effector. Locus B of immunoglobulin heavy chain mapped to an area containing multiple SNPs with genome-wide association. Heart mRNA-seq comparing parr from families with high- versus low-genomic breeding value, and matching sample genotypes for SNPs, identified two eQTL for salmonid alphavirus load. Immune genes associated with trans-eQTL were numerous and spread throughout the genome. QTL regions contained several genes with known or predicted immune functions, some differentially expressed. The putative functional genes and variants identified could help improve marker-based selection for pancreas disease resistance.

Response of the Salmon Heart Transcriptome to Pancreas Disease: Differences Between High- and Low-Ranking Families for Resistance.

Pancreas disease caused by salmonid alphaviruses leads to severe losses in Atlantic salmon aquaculture. The aim of our study was to gain a better understanding of the biological differences between salmon with high and low genomic breeding values (H-gEBV and L-gEBV respectively) for pancreas disease resistance. Fish from H- and L-gEBV families were challenged by intraperitoneal injection of salmonid alphavirus or co-habitation with infected fish. Mortality was higher with co-habitation than injection, and for L- than H-gEBV. Heart for RNA-seq and histopathology was collected before challenge and at four- and ten-weeks post-challenge. Heart damage was less severe in injection-challenged H- than L-gEBV fish at week 4. Viral load was lower in H- than L-gEBV salmon after co-habitant challenge. Gene expression differences between H- and L-gEBV manifested before challenge, peaked at week 4, and moderated by week 10. At week 4, H-gEBV salmon showed lower expression of innate antiviral defence genes, stimulation of B- and T-cell immune function, and weaker stress responses. Retarded resolution of the disease explains the higher expression of immune genes in L-gEBV at week 10. Results suggest earlier mobilization of acquired immunity better protects H-gEBV salmon by accelerating clearance of the virus and resolution of the disease.

Mapping and validation of a major QTL affecting resistance to pancreas disease (salmonid alphavirus) in Atlantic salmon (Salmo salar).

Pancreas disease (PD), caused by a salmonid alphavirus (SAV), has a large negative economic and animal welfare impact on Atlantic salmon aquaculture. Evidence for genetic variation in host resistance to this disease has been reported, suggesting that selective breeding may potentially form an important component of disease control. The aim of this study was to explore the genetic architecture of resistance to PD, using survival data collected from two unrelated populations of Atlantic salmon; one challenged with SAV as fry in freshwater (POP 1) and one challenged with SAV as post-smolts in sea water (POP 2). Analyses of the binary survival data revealed a moderate-to-high heritability for host resistance to PD in both populations (fry POP 1 h(2)~0.5; post-smolt POP 2 h(2)~0.4). Subsets of both populations were genotyped for single nucleotide polymorphism markers, and six putative resistance quantitative trait loci (QTL) were identified. One of these QTL was mapped to the same location on chromosome 3 in both populations, reaching chromosome-wide significance in both the sire- and dam-based analyses in POP 1, and genome-wide significance in a combined analysis in POP 2. This independently verified QTL explains a significant proportion of host genetic variation in resistance to PD in both populations, suggesting a common underlying mechanism for genetic resistance across lifecycle stages. Markers associated with this QTL are being incorporated into selective breeding programs to improve PD resistance.

Sampling strategies for whole genome association studies in aquaculture and outcrossing plant species.

A number of farmed species are characterized by breeding populations of large full-sib families, including aquaculture species and outcrossing plant species. Whole genome association studies in such species must account for stratification arising from the full-sib family structure to avoid high rates of false discovery. Here, we demonstrate the value of selective genotyping strategies which balance the contribution of families across high and low phenotypes to greatly reduce rates of false discovery with a minimal effect on power.

Detection of QTL for growth rate in the blacklip abalone (Haliotis rubra Leach) using selective DNA pooling.

The objective of this study was to identify QTL for growth rate in the blacklip abalone Haliotis rubra using selective DNA pooling. Three full-sibling families of H. rubra derived from crosses of wild broodstock were used. DNA was extracted from the largest and smallest 10% of progeny and combined into two pools for each phenotypic tail. The DNA pools were typed with 139 microsatellites, and markers showing significant differences between the peak height ratios of alleles inherited from the parents were individually genotyped and analysed by interval mapping. A strong correlation (r = 0.94, P < 0.001) was found between the t-values from the analysis of pools and the t-values from the analysis of individual genotypes. Based on the interval mapping analysis, QTL were detected on nine linkage groups at a chromosome-wide P < 0.01 and one linkage group at a chromosome-wide P < 0.05. The study demonstrated that selective DNA pooling is efficient and effective as a first-pass screen for the discovery of QTL in an aquaculture species.

A microsatellite linkage map of the blacklip abalone, Haliotis rubra.

There is considerable scope for genetic improvement of cultured blacklip abalone Haliotis rubra in Australia using molecular marker-assisted, selective-breeding practices. Such improvement is dependent on the availability of primary genetic resources, such as a genetic linkage map. This study presents a first-generation linkage map of H. rubra, containing 122 microsatellite markers typed in a single full-sib family. These loci mapped to 17 and 20 linkage groups for the male and female respectively, and when aligned, the consensus map represented 18 linkage groups. The male linkage map contained 102 markers (one unlinked) covering 621 cM with an average intermarker spacing of 7.3 cM, and the female map contained 98 markers (eight unlinked) covering 766 cM with an average intermarker spacing of 9.8 cM. Analysis of markers informative in both parents showed a significantly higher recombination rate in the female parent, with an average male-to-female recombination ratio of 1:1.45 between linked pairs of markers. This linkage map represents a significant advancement in the genetic resource available for H. rubra and provides a framework for future quantitative trait loci mapping and eventual implementation of marker-assisted selection.

Effect of crystal freezing and small-molecule binding on internal cavity size in a large protein: X-ray and docking studies of lipoxygenase at ambient and low temperature at 2.0 A resolution.

Flash-freezing is a technique that is commonly used nowadays to collect diffraction data for X-ray structural analysis. It can affect both the crystal and molecular structure and the molecule's surface, as well as the internal cavities. X-ray structural data often serve as a template for the protein receptor in docking calculations. Thus, the size and shape of the binding site determines which small molecules could be found as potential ligands in silico, especially during high-throughput rigid docking. Data were analyzed for wild soybean lipoxygenase-3 (MW 97 kDa) at 293 and 93 K and compared with the results from studies of its molecular complexes with known inhibitors, structures published by others for a derivative of the same enzyme (98 K) or a topologically close isozyme lipoxygenase-1 (at ambient temperature and 100 K). Analysis of these data allows the following conclusions. (i) Very small changes in the relative orientation of the molecules in the crystal can cause major changes in the crystal reciprocal lattice. (ii) The volume of the internal cavities can ;shrink' by several percent upon freezing even when the unit-cell and the protein molecular volume show changes of only 1-2%. (iii) Using a receptor structure determined based on cryogenic data as a target for computational screening requires flexible docking to enable the expansion of the binding-site cavity and sampling of the alternative conformations of the crucial residues.

Sfrp-1 and sfrp-2 are expressed in overlapping and distinct domains during chick development.

Secreted frizzled related proteins (Sfrps) are thought to bind and regulate Wnt activity through a cysteine rich domain that is highly similar to that of Frizzled receptors. To investigate possible roles for Sfrps in chick development, we have isolated partial cDNAs encoding Sfrp-1 and Sfrp-2 and have thoroughly characterized the expression patterns of both genes. Both sfrp-1 and sfrp-2 are expressed at all stages of development analyzed, ranging from Hamburger and Hamilton stage 4 through stage 32. Expression of both sfrp-1 and sfrp-2 is observed in mesodermal and ectodermal derivatives, while sfrp-1 is also found in endodermal lineages.

The dynamic expression pattern of frzb-1 suggests multiple roles in chick development.

The Wnt family of secreted proteins has been shown to have multiple roles in embryonic development. Wnt signals are thought to be propagated by binding to the cysteine-rich extracellular domain (CRD) of Frizzled, a seven-transmembrane-domain cell surface receptor. Secreted Frizzled-related proteins (generally denoted Frzb or Sfrp) possess a domain with a high degree of sequence identity and structural similarity with the CRD of Frizzled. Current data indicate that the cysteine-rich domain of secreted Frzb proteins can bind Wnt proteins, suggesting the possibility that Frzbs compete with membrane-bound Frizzled for Wnt binding and consequently act as competitive inhibitors of Wnt signaling. In order to gain a better understanding of the potential roles of Frzb-1 in chick development, we utilized the polymerase chain reaction to isolate a partial cDNA of the chick orthologue of frzb-1, cfrzb-1, and compared its expression pattern to that of Wnt-1, Wnt-3a, Wnt-5a, Wnt-7a, and Wnt-8c. Whole-mount in situ hybridizations have revealed three major phases of expression for cfrzb-1 in the developing chick. The earliest expression of cfrzb-1 is in cells fated to become neural ectoderm in streak-stage embryos. Expression of cfrzb-1 in the neural ectoderm continues up through stage 8. After stage 8, cfrzb-1 expression is gradually attenuated in the closing neural tube of the trunk and is concomitantly up-regulated in neural crest cells. Finally, cfrzb-1 appears in the condensing mesenchyme of the bones in both the limb and the trunk in stage 25+ embryos. Comparative analysis of the cfrzb-1 and the Wnt gene expression patterns suggests possible interactions between cFrzb-1 and all of the Wnt family members examined.

Frozen and radiation-sterilized bone allografts in the treatment of post-traumatic malformation of bones.

Post-traumatical malformations of bone are often reconstructed with the use of preserved bone allografts. At the Institute of Traumatology, Orthopaedics and Neurosurgery 495 patients were treated with use of preserved (frozen and radiation-sterilized) bone allografts, following trauma, between 1981 and 1995. Non union of bone and osteomyelitis were main reasons for allotransplantation of bone. Remodelling of bone allografts has been observed during 2-8 years after surgery. Substitution of allografts and good result of treatment were found in 80% of all cases. In paper the analysis of results of treatment is presented.

Fusion of spine in children scoliosis with frozen & radiation--sterilized bone allograft.

102 children have been treated at the Institute of Traumatology, Orthopaedics & Neurosurgery a result of scoliosis. In all of these multi-step treatment has been applied. Initially a telescopic rod was implanted into the spine. Allografts were introduced around the rod hook, after 6 or 8 months, when angle of scoliosis increased, the rod was exchanged for a longer one. The final step was performed when conditions permitted and the scoliosis was markedly corrected and the fusion of the spine with the solid allograft was accomplished. The result of treatment was evaluated 1 to 7 years after surgery. Bone allografts were rebuilt within 6 month. Correction of scoliosis of 50% to 70% was achieved in all cases.

Fractures of thoracic and lumbar spine; treatment and follow up.

The number of posttraumatic spinal fractures is constantly growing. About 15-20% of patients injury need surgery after injury, as result of spinal instability or canal stenosis often leading to neurological complications. In 1987-97 56 patients (6 female and 50 male) were treated following serious spine trauma with fractures. The age of the patients was 19-61 years. Car accidents were the most common cause of trauma affecting 30 patients (53%), falls from heights affecting 26 patients (47%). Multiple injuries were diagnosed in 24 patients (43%). More than one vertebra was fractured in 15 patients (27%). In 9 cases (16%) surgery was performed within 24 hours of the accident, in 22 (39%) within 72 hours and in 25 patients (45%) after 72 hours, in some cases even several weeks after the trauma. 20 patients (36%) required spinal decompression after the accident. The fracture in one case was located in the thoracic section, in 17 patients in Th11-L2 and in 2 patients in L3-L5 section. In 4 cases reconstruction of the meninx was performed. 48 patients were treated by stabilisation using Harrington rods; in 5 patients Kluger stabilisers were used, in the remaining 3 patients stabilization was carried out by wire loop. In all cases frozen and radiation-sterilized bone allografts were applied. Results were evaluated 2-8 years after surgery using X-rays, CT-pictures and direct examination. In 25 patients (45%) results were found to be good, in 23 patients (41%) mediocre and in 8 patients (14%) results were unsatisfactory.

A study of preventable trauma mortality in rural Michigan.

OBJECTIVE: To determine the preventable death rate (PDR) and the frequency and types of inappropriate medical care in a large, rural region of Michigan. DESIGN: A prospective study of all deaths caused by injury during a 1-year period. METHODS: Preventability of death and appropriateness of care were determined using a structured implicit review process and expert panel. A second panel was convened to confirm the reliability of the review process. MAIN RESULTS: One hundred fifty-five injury-related deaths underwent panel review. Four deaths (2.6%) were found to be definitely preventable and 16 (10.3%) possibly preventable, for a combined preventable death rate of 12.9%. Sixty-five deaths (41.9%) occurred in the emergency department or hospital; 18 of these (27.7%) were judged to be definitely preventable or possibly preventable. Forty-three episodes of inappropriate care were identified in 27 (17.4%) of the 155 cases reviewed. These occurred primarily in the emergency department and hospital rather than during prehospital care or transfer. CONCLUSIONS: A relatively small percentage of trauma fatalities in rural Michigan could have been prevented by more appropriate or timely medical care. Efforts to improve the care of injured persons in rural Michigan should be directed primarily at the emergency department and inpatient phases of trauma system care.

Treatment of dermal cysticercosis with praziquantel. A new cestocidal agent.

Vinte pacientes adultos, com cisticercose subcutanea, sem associacao com envolvimento ocular ou cerebral, foram tratados com praziquantel. Os primeiros 11 casos receberam 60 mg/kg/dia e os ultimos nove 30 mg/kg/dia. Em ambos os grupos a dose oral diaria foi dividida em tres tomadas, a intervalos de quatro a seis horas e a administracao do medicamento estendeu-se por seis dias consecutivos. O segundo grupo de pacientes tomou tambem dexametasona, 3 mg diarios, desde um dia antes ate quatro dias depois do periodo de tratamento com praziquantel. A droga mostrou-se 100% eficaz como evidenciado histopatologicamente, pela morte da larva da Taenia solium (Cysticercus cellulosae) em biopsias seriadas, realizadas a partir da segunda semana apos a terapeutica, bem como clinicamente, pelo paulatino desaparecimento dos nodulos cutaneos durante os seis meses seguintes ao tratamento. A tolerancia ao praziquantel foi boa, tendo em vista a pouca relevancia da incidencia e da intensidade dos efeitos colaterais. A seguranca do medicamento foi confirmada atraves de provas laboratoriais que nao revelaram quaisquer anormalidades relativas as funcoes hematopoietica, hepatica e renal