Morphological and molecular characterization of developing vertebral fusions using a teleost model

<p>Abstract</p> <p>Background</p> <p>Spinal disorders are a major cause of disability for humans and an important health problem for intensively farmed animals. Experiments have shown that vertebral deformities present a complex but comparable etiology across species. However, the underlying molecular mechanisms involved in bone deformities are still far from understood. To further explicate the mechanisms involved, we have examined the fundamental aspects of bone metabolism and pathogenesis of vertebral fusions in Atlantic salmon (<it>Salmo salar</it>).</p> <p>Results</p> <p>Experimentally, juvenile salmon were subjected to hyperthermic conditions where more than 28% developed fused vertebral bodies. To characterize the fusion process we analyzed an intermediate and a terminal stage of the pathology by using x-ray, histology, immunohistochemistry, real-time quantitative PCR and <it>in situ </it>hybridization. At early stage in the fusion process, disorganized and proliferating osteoblasts were prominent at the growth zones of the vertebral body endplates. PCNA positive cells further extended along the rims of fusing vertebral bodies. During the developing pathology, the marked border between the osteoblast growth zones and the chondrocytic areas connected to the arches became less distinct, as proliferating cells and chondrocytes blended through an intermediate zone. This cell proliferation appeared to be closely linked to fusion of opposing arch centra. During the fusion process a metaplastic shift appeared in the arch centra where cells in the intermediate zone between osteoblasts and chondrocytes co-expressed mixed signals of chondrogenic and osteogenic markers. A similar shift also occurred in the notochord where proliferating chordoblasts changed transcription profile from chondrogenic to also include osteogenic marker genes. In progressed fusions, arch centra and intervertebral space mineralized.</p> <p>Conclusion</p> <p>Loss of cell integrity through cell proliferation and metaplastic shifts seem to be key events in the fusion process. The fusion process involves molecular regulation and cellular changes similar to those found in mammalian deformities, indicating that salmon is suitable for studying general bone development and to be a comparative model for spinal deformities.</p

Molecular pathology of vertebral deformities in hyperthermic Atlantic salmon (Salmo salar)

<p>Abstract</p> <p>Background</p> <p>Hyperthermia has been shown in a number of organisms to induce developmental defects as a result of changes in cell proliferation, differentiation and gene expression. In spite of this, salmon aquaculture commonly uses high water temperature to speed up developmental rate in intensive production systems, resulting in an increased frequency of skeletal deformities. In order to study the molecular pathology of vertebral deformities, Atlantic salmon was subjected to hyperthermic conditions from fertilization until after the juvenile stage.</p> <p>Results</p> <p>Fish exposed to the high temperature regime showed a markedly higher growth rate and a significant higher percentage of deformities in the spinal column than fish reared at low temperatures. By analyzing phenotypically normal spinal columns from the two temperature regimes, we found that the increased risk of developing vertebral deformities was linked to an altered gene transcription. In particular, down-regulation of extracellular matrix (ECM) genes such as <it>col1a1</it>, <it>osteocalcin</it>, <it>osteonectin </it>and <it>decorin</it>, indicated that maturation and mineralization of osteoblasts were restrained. Moreover, histological staining and <it>in situ </it>hybridization visualized areas with distorted chondrocytes and an increased population of hypertrophic cells. These findings were further confirmed by an up-regulation of <it>mef2c </it>and <it>col10a</it>, genes involved in chondrocyte hypertrophy.</p> <p>Conclusion</p> <p>The presented data strongly indicates that temperature induced fast growth is severely affecting gene transcription in osteoblasts and chondrocytes; hence change in the vertebral tissue structure and composition. A disrupted bone and cartilage production was detected, which most likely is involved in the higher rate of deformities developed in the high intensive group. Our results are of basic interest for bone metabolism and contribute to the understanding of the mechanisms involved in development of temperature induced vertebral pathology. The findings may further conduce to future molecular tools for assessing fish welfare in practical farming.</p

Photoperiod in recirculation aquaculture systems and timing of seawater transfer affect seawater growth performance of Atlantic salmon (Salmo salar)

Production of Atlantic salmon smolts in recirculation aquaculture systems (RAS) is growing, and novel production protocols using continuous light in RAS are being implemented in the industry. In the present study, Atlantic Salmon parr were exposed to either a traditional protocol (short-day winter signal [12:12 L:D] for 6 weeks) or to continuous light. Both photoperiods were applied in freshwater (FW) and brackish water RAS. Salmon from all treatments were transferred to seawater pens at 200 and 600 g and grown until slaughter size. A control group was smoltified with a 6-week short-day winter signal and kept in FW until sea transfer at 100 g. Continuous light gave a higher growth rate in RAS but reduced feed intake and growth and increased feed conversion ratio during the first 8 weeks in seawater. However, at slaughter, fish exposed to continuous light was bigger than fish given a winter signal because of the higher growth rate in RAS. Slaughter weight was lowest in fish transferred to sea at 600 g, despite having the highest day-degree sum during their life span. The best performing group was the control group transferred at 100 g. All treatments handled transfer to seawater and survival and maturation were not affected by the treatments in RAS. The immune status was examined with a multigene expression assay on BioMark HD platform from parr stage to 5–7 months after seawater transfer. Overall, there was no significant effect of photoperiod or salinity on the expression of the selected immune genes. In sum, the results from this study indicate that using continuous light in RAS may have negative effects on performance shortly after transfer in fish transferred to sea at 200 g, whereas at 600 g, all treatments had reduced growth after transfer irrespective of treatment in RAS.publishedVersio

Cardiac responses to elevated seawater temperature in Atlantic salmon

publishedVersio

Association between swimming performance, cardiorespiratory morphometry, and thermal tolerance in Atlantic salmon (Salmo salar L.)

This experiment tested the hypothesis that swimming performance in Atlantic salmon (Salmo salar) parr is connected to cardiorespiratory performance and morphology, as well as maximum heart rate (fHmax) related measures of thermal tolerance. Moreover, it was hypothesized that the cardiorespiratory differences between poor and strong swimmers will be retained in a later life stage, i.e., 15 weeks post-smoltification and seawater transfer. This experiment screened a population of 3200 parr (11.2 ± 0.25 g) for their swimming performance, classifying them as poor and good swimmers based on their critical swimming speeds (4.4 ± 0.1 body length s−1 and > 6.8 ± 0.1 body length s−1, respectively). Compared with poor performing parr, good swimmers had a significantly thicker compact myocardium (by 23.7%) and taller gill secondary lamellae (by 16.2%). In contrast, there was no significant difference in maximum oxygen consumption between the two groups as assessed using a “chase” protocol, and the relationship between heart rate specific measures of thermal tolerance and swim performance was variable. For example, three measures did not differ between the two groups, whereas the Arrhenius breakpoint temperature for fHmax and the highest fHmax value were lower and higher, respectively, in the poor swimmers. Importantly, the identified morphological and difference in the highest fHmax value at the parr stage persisted after 15 weeks of common garden rearing in seawater, and they were associated with an increase in relative ventricular mass and a small, but significant, improvement in growth rate. Therefore, it seems that an early assessment of swimming performance can effectively screen for morphological capacities related to oxygen supply and growth rate, but less so for heart rate related measures of thermal tolerance.</p

ANALISIS KEMAMPUAN BERFIKIR ALJABAR DI KELAS VII SMP NEGERI 1 KRANGKENG KABUPATEN INDRAMAYU (Studi Survei di Kelas VII Tahun Akademik 2012/2013)

Setiati Rahayu, 2013. Analisis Kemampuan Berfikir Aljabar di Kelas VII di SMP Negeri 1 Krangkeng Kab. Indramayu (Studi Survei di Kelas VII Tahun Akademik 2012/2013) Untuk mengukur kemampuan berfikir aljabar seorang siswa diperlukan beberapa indikator. Terdapat beberapa indikator untuk memahami aljabar diantaranya memahami pengertian koefisien variabel suku sejenis, kemampuan melakuakan operasi hitung, kemampuan mengggunakan simbol matematika, kemampuan menggunakan bahasa sehari-hari, kemampuan menyederhanakan operasi aljabar, kemampuan menyatakan berbagai hubungan, kemampuan menggunakan diagram alur dan memahami konsep aljabar invers. Tujuan dalam penelitian ini untuk menganalisis kemampuan berfikir aljabar di kelas VII yang ditinjau dari operasi aljabar, penggunaan aljabar, masalah aljabar, dan pemahaman konsep aljabar saat menjawab soal-soal aljabar. Kemampuan siswa dalam memahami aljabar merupakan suatu kondisi yang terdapat dalam sekolah SMP tersebut. Aljabar adalah suatu cabang ilmu matematika yang menggunakan tandatanda dan huruf-huruf yang mewakili angka-angka. Banyaknya indikator yang menjadi indikator kemampuan aljabar, maka akan ada beberapa indikator yang akan menjadi komponen utama dalam kemampuan aljabar. Untuk mengetahui kemampuan siswa dalam memahami aljabar, tidak perlu semua indikator pada\ud materi aljabar diujikan dalam tes. Mengetahui sangat pentingnya komponen utama dalam materi aljabar untuk mengukur kemampuan berfikir aljabar seorang siswa maka peneliti mencoba menganalisis kemampuan berfikir aljabar siswa kelas VII di SMPN 1 Krangkeng. Teknik pengumpulan data yang digunakan dalam penelitian ini adalah tes. Populasi dalam penelitian ini adalah Kelas VII yang berjumlah (160 siswa). Sampel diambil dari kelas VIIB cluster random sampling. Untuk uji coba dilakukan di kelas VIIA yang berjumlah 32. Dari hasil penelitian diperoleh hasil sebagai berikut: 1) Aljabar di SMP N 1 Krangkeng cukup baik hal ini ditunjukan dengan nilai rata-rata 75. 2) Indikator kemampuan berfikir aljabar secara mekanisme dan indikator Kemampuan terbesar yang dicapai oleh siswa kelas VII SMP Negeri 1 Krangkeng tahun ajaran 2012/2013 dalam memahami aljabar adalah pada pengertian koefisien, variabel, konstanta, faktor, suku sejenis adalah sebesar 89.58% dan kemampuan yang kurang dikuasai oleh siswa kelas VII SMP Negeri 1 Krangkeng tahun ajaran 2012/2013 dalam memahami aljabar adalah memahami invers adalah sebesar 85.94%. 3) Dari delapan indicator kemampuan berfikir aljabar ada tiga komponen yang dominan yaitu menggunakan diagram alur, pengertian koefisien variable konstanta, faktor suku sejenis, dan menyederhanakan operasi aljaba

Mineral nutrition and bone health in salmonids

In the recent years, challenges faced in salmonid farming with regard to deformities have helped to improve our knowledge on skeletal biology and development of bone health in salmonids. Different nutritional, genetic and environmental factors are associated with skeletal deformities in salmonids and other fish species. Minerals are a group of essential nutrients having a vital role to play in skeletal development, growth and remodelling. The knowledge generated thus far on the structural and functional importance of minerals in salmonid bone health is largely restricted to phosphorus. A brief account of dietary phosphorus‐related bone deformities encountered in salmonids, critical life stages for the development of bone deformities, recent developments in the understanding of their aetiology and dietary phosphorus levels required to improve bone health are presented. The effect of increased dietary level of phosphorus in reducing bone health disorders under conditions of improved feed conversion efficiency and use of triploids in salmon farming are illustrated with help of metadata analysis from literature. With regard to the role of other essential minerals (macro‐ and micro‐) impacting bone health in salmonids, data available are extremely limited and hence information on other fish species and mammals is summarized in relation to bone development and incidence of deformities. The need to improve mineral bioavailability, utilization and reduce effluent mineral load is presented in brief with example from Norwegian salmon farming. Refinements in mineral requirement recommendations for salmonids and advanced methodologies for studying aetiology of skeletal anomalies, bone mineral status, skeletal development and deformities are also discussed.publishedVersio

Transcriptome profiling of immune responses to cardiomyopathy syndrome (CMS) in Atlantic salmon

<p>Abstract</p> <p>Background</p> <p>Cardiomyopathy syndrome (CMS) is a disease associated with severe myocarditis primarily in adult farmed Atlantic salmon (<it>Salmo salar </it>L.), caused by a double-stranded RNA virus named piscine myocarditis virus (PMCV) with structural similarities to the <it>Totiviridae </it>family. Here we present the first characterisation of host immune responses to CMS assessed by microarray transcriptome profiling.</p> <p>Results</p> <p>Unvaccinated farmed Atlantic salmon post-smolts were infected by intraperitoneal injection of PMCV and developed cardiac pathology consistent with CMS. From analysis of heart samples at several time points and different tissues at early and clinical stages by oligonucleotide microarrays (SIQ2.0 chip), six gene sets representing a broad range of immune responses were identified, showing significant temporal and spatial regulation. Histopathological examination of cardiac tissue showed myocardial lesions from 6 weeks post infection (wpi) that peaked at 8-9 wpi and was followed by a recovery. Viral RNA was detected in all organs from 4 wpi suggesting a broad tissue tropism. High correlation between viral load and cardiac histopathology score suggested that cytopathic effect of infection was a major determinant of the myocardial changes. Strong and systemic induction of antiviral and IFN-dependent genes from 2 wpi that levelled off during infection, was followed by a biphasic activation of pathways for B cells and MHC antigen presentation, both peaking at clinical pathology. This was preceded by a distinct cardiac activation of complement at 6 wpi, suggesting a complement-dependent activation of humoral Ab-responses. Peak of cardiac pathology and viral load coincided with cardiac-specific upregulation of T cell response genes and splenic induction of complement genes. Preceding the reduction in viral load and pathology, these responses were probably important for viral clearance and recovery.</p> <p>Conclusions</p> <p>By comparative analysis of gene expression, histology and viral load, the temporal and spatial regulation of immune responses were characterised and novel immune genes identified, ultimately leading to a more complete understanding of host-virus responses and pathology and protection in Atlantic salmon during CMS.</p