Early Development and Tissue Distribution of Pseudoloma neurophilia in the Zebrafish,Danio rerio

The early proliferative stages of the microsporidian parasite, Pseudoloma neurophilia were visualized in larval zebrafish, Danio rerio, using histological sections with a combination of an in situ hybridization probe specific to the P. neurophilia small‐subunit ribosomal RNA gene, standard hematoxylin‐eosin stain, and the Luna stain to visualize spores. Beginning at 5 d post fertilization, fish were exposed to P. neurophilia and examined at 12, 24, 36, 48, 72, 96, and 120 h post exposure (hpe). At 12 hpe, intact spores in the intestinal lumen and proliferative stages developing in the epithelial cells of the anterior intestine and the pharynx and within hepatocytes were observed. Proliferative stages were visualized in the pancreas and kidney at 36–48 hpe and in the spinal cord, eye, and skeletal muscle beginning at 72 hpe. The first spore stages of P. neurophilia were observed at 96 hpe in the pharyngeal epithelium, liver, spinal cord, and skeletal muscle. The parasite was only observed in the brain of larval fish at 120 hpe. The distribution of the early stages of P. neurophilia and the lack of mature spores until 96 hpe indicates that the parasite gains access to organs distant from the initial site of entry, likely by penetrating the intestinal wall with the polar tube.     

Identification of quantitative trait loci affecting gastrointestinal parasite resistance in an experimental Angus population

DNA markers associated with quantitative trait loci (QTL) affecting host tolerance to gastrointestinal (GI) parasite infection are ideal targets for marker‐assisted selection. However, few studies in cattle have attempted to identify this type of QTL due to the difficulty of generating accurate phenotypic data from a resource population with adequate statistical power for detection. For this effort, we amassed fecal egg count (FEC) measures from annual natural field challenges with GI nematodes that spanned 12 different contemporary groups of Angus calves (1992–2000) derived from a closed breeding population. FEC and blood pepsinogen measures were taken weekly over a 26‐week period post‐weaning, and the FEC data were Box‐Cox transformed to normalize the distribution of phenotypes. These 305 test animals and more than 100 founding animals from the extended pedigree were genotyped across 190 microsatellites markers. The genome‐wide analyses identified a suggestive genome‐wide QTL on bovine chromosome (Chr) 8 (P < 0.002) and nominal QTL on Chr 4, 12 and 17 (P < 0.05). These findings were unique for cattle, and some corresponded to previously identified QTL locations for parasite‐related traits in sheep to provide genome locations for further fine mapping of parasite resistance/susceptibility in Angus cattle.     

Zebrafish transgenic Enhancer TRAP line database (ZETRAP)

Background  

The zebrafish, Danio rerio, is used as a model organism to study vertebrate genetics and development. An effective enhancer trap (ET) in zebrafish using the Tol2 transposon has been demonstrated. This approach could be used to study embryogenesis of a vertebrate species in real time and with high resolution.     

Booster vaccination with live attenuated Vibrio anguillarum elicits strong protection despite weak specific antibody response in zebrafish

A live attenuated Vibrio anguillarum vaccine was recently established in the laboratory that induced immunoprotection against vibriosis in zebrafish, Danio rerio. To improve immunogenicity, the effects of different booster vaccination regimens were investigated using bath‐vaccination in a zebrafish model. Zebrafish receiving booster doses at 2 weeks or at both 2 and 4 weeks after primary vaccination were better protected in comparison to fish that received a single vaccination. In addition, the booster vaccination induced a prolonged specific antibody response. No correlation between a weak specific antibody response and a strong protection was observed, indicating that the booster vaccination could enhance the affinity of Immunoglobulin M (IgM) rather than the amount. Moreover, changes in the immune‐related gene expression of the booster‐vaccinated group suggested that the booster enhanced the adaptive immune responses.     

Systematic analysis of mRNA 5' coding sequence incompleteness in Danio rerio: an automated EST-based approach

Background  

All standard methods for cDNA cloning are affected by a potential inability to effectively clone the 5' region of mRNA. The aim of this work was to estimate mRNA open reading frame (ORF) 5' region sequence completeness in the model organism Danio rerio (zebrafish).     

Improved genome assembly provides new insights into genome evolution in a desert poplar (Populus euphratica)

Populus euphratica is well adapted to extreme desert environments and is an important model species for elucidating the mechanisms of abiotic stress resistance in trees. The current assembly of P. euphratica genome is highly fragmented with many gaps and errors, thereby impeding downstream applications. Here, we report an improved chromosome‐level reference genome of P. euphratica (v2.0) using single‐molecule sequencing and chromosome conformation capture (Hi‐C) technologies. Relative to the previous reference genome, our assembly represents a nearly 60‐fold improvement in contiguity, with a scaffold N50 size of 28.59 Mb. Using this genome, we have found that extensive expansion of Gypsy elements in P. euphratica led to its rapid increase in genome size compared to any other Salicaceae species studied to date, and potentially contributed to adaptive divergence driven by insertions near genes involved in stress tolerance. We also detected a wide range of unique structural rearrangements in P. euphratica, including 2,549 translocations, 454 inversions, 121 tandem and 14 segmental duplications. Several key genes likely to be involved in tolerance to abiotic stress were identified within these regions. This high‐quality genome represents a valuable resource for poplar breeding and genetic improvement in the future, as well as comparative genomic analysis with other Salicaceae species.     

The zebrafish as a model organism for the study of apoptosis

Apoptosis plays important roles in embryogenesis, tissue homeostasis, and immune system regulation. The zebrafish (Danio rerio) is a powerful vertebrate model organism that has been extensively used to study apoptotic cell death during normal development and under conditions of cellular stress. In the past 5 years, a detailed picture has begun to emerge of the molecular underpinnings of the cell-intrinsic and the cell-extrinsic apoptosis signaling pathways in zebrafish. We begin this review with an introduction to the techniques and experimental approaches that are used to study apoptosis in zebrafish. We follow with a general overview of developmental apoptosis during zebrafish embryogenesis. Finally, we present a comprehensive review of the intrinsic and extrinsic apoptosis pathways in zebrafish, focusing on the high degree of conservation with humans and other mammals. Recent publications that draw upon the unique advantages of the zebrafish system to study novel aspects of apoptosis regulation and function are highlighted throughout.     

The genome of the marine medaka Oryzias melastigma

Marine medaka (Oryzias melastigma) is considered to be a useful fish model for marine and estuarine ecotoxicology studies and has good potential for field‐based population genomics because of its geographical distribution in Asian estuarine and coastal areas. In this study, we present the first whole‐genome draft of O. melastigma. The genome assembly consists of 8,602 scaffolds (N50 = 23.737 Mb) and a total genome length of 779.4 Mb. A total of 23,528 genes were predicted, and 12,670 gene families shared with three teleost species (Japanese medaka, mangrove killifish and zebrafish) were identified. Genome analyses revealed that the O. melastigma genome is highly heterozygous and contains a large number of repeat sequences. This assembly represents a useful genomic resource for fish scientists.     

Gelatinases, endonuclease and Vascular Endothelial Growth Factor during development and regression of swine luteal tissue

Background  

Zebrafish (D. rerio) has become a powerful and widely used model system for the analysis of vertebrate embryogenesis and organ development. While genetic methods are readily available in zebrafish, protocols for two dimensional (2D) gel electrophoresis and proteomics have yet to be developed.     

Chromosome‐level genome assembly of a cyprinid fish Onychostoma macrolepis by integration of nanopore sequencing,Bionano and Hi‐C technology

Onychostoma macrolepis is an emerging commercial cyprinid fish species. It is a model system for studies of sexual dimorphism and genome evolution. Here, we report the chromosome‐level assembly of the O.macrolepis genome obtained from the integration of nanopore long‐read sequencing with physical maps produced using Bionano and Hi‐C technology. A total of 87.9 Gb of nanopore sequence provided approximately 100‐fold coverage of the genome. The preliminary genome assembly was 883.2 Mb in size with a contig N50 size of 11.2 Mb. The 969 corrected contigs obtained from Bionano optical mapping were assembled into 853 scaffolds and produced an assembly of 886.5 Mb with a scaffold N50 of 16.5 Mb. Finally, using the Hi‐C data, 881.3 Mb (99.4% of genome) in 526 scaffolds were anchored and oriented in 25 chromosomes ranging in size from 25.27 to 56.49 Mb. In total, 24,770 protein‐coding genes were predicted in the genome, and ~96.85% of the genes were functionally annotated. The annotated assembly contains 93.3% complete genes from the BUSCO reference set. In addition, we identified 409 Mb (46.23% of the genome) of repetitive sequence, and 11,213 non‐coding RNAs, in the genome. Evolutionary analysis revealed that O. macrolepis diverged from common carp approximately 24.25 million years ago. The chromosomes of O. macrolepis showed an unambiguous correspondence to the chromosomes of zebrafish. The high‐quality genome assembled in this work provides a valuable genomic resource for further biological and evolutionary studies of O. macrolepis.     

Chromosome‐level genome assembly of the predator Propylea japonica to understand its tolerance to insecticides and high temperatures

The ladybird beetle Propylea japonica is an important natural enemy in agro‐ecological systems. Studies on the strong tolerance of P. japonica to high temperatures and insecticides, and its population and phenotype diversity have recently increased. However, abundant genome resources for obtaining insights into stress‐resistance mechanisms and genetic intra‐species diversity for P. japonica are lacking. Here, we constructed the P. japonica genome maps using Pacific Bioscience (PacBio) and Illumina sequencing technologies. The genome size was 850.90 Mb with a contig N50 of 813.13 kb. The Hi‐C sequence data were used to upgrade draft genome assemblies; 4,777 contigs were assembled to 10 chromosomes; and the final draft genome assembly was 803.93 Mb with a contig N50 of 813.98 kb and a scaffold N50 of 100.34 Mb. Approximately 495.38 Mb of repeated sequences was annotated. The 18,018 protein‐coding genes were predicted, of which 95.78% were functionally annotated, and 1,407 genes were species‐specific. The phylogenetic analysis showed that P. japonica diverged from the ancestor of Anoplophora glabripennis and Tribolium castaneum ~ 236.21 million years ago. We detected that some important gene families involved in detoxification of pesticides and tolerance to heat stress were expanded in P. japonica, especially cytochrome P450 and Hsp70 genes. Overall, the high‐quality draft genome sequence of P. japonica will provide invaluable resource for understanding the molecular mechanisms of stress resistance and will facilitate the research on population genetics, evolution and phylogeny of Coccinellidae. This genome will also provide new avenues for conserving the diversity of predator insects.     

Genomic evidence for population‐specific responses to co‐evolving parasites in a New Zealand freshwater snail

Reciprocal co‐evolving interactions between hosts and parasites are a primary source of strong selection that can promote rapid and often population‐ or genotype‐specific evolutionary change. These host–parasite interactions are also a major source of disease. Despite their importance, very little is known about the genomic basis of co‐evolving host–parasite interactions in natural populations, especially in animals. Here, we use gene expression and sequence evolution approaches to take critical steps towards characterizing the genomic basis of interactions between the freshwater snail Potamopyrgus antipodarum and its co‐evolving sterilizing trematode parasite, Microphallus sp., a textbook example of natural coevolution. We found that Microphallus‐infected P. antipodarum exhibit systematic downregulation of genes relative to uninfected P. antipodarum. The specific genes involved in parasite response differ markedly across lakes, consistent with a scenario where population‐level co‐evolution is leading to population‐specific host–parasite interactions and evolutionary trajectories. We also used an F_ST‐based approach to identify a set of loci that represent promising candidates for targets of parasite‐mediated selection across lakes as well as within each lake population. These results constitute the first genomic evidence for population‐specific responses to co‐evolving infection in the P. antipodarum‐Microphallus interaction and provide new insights into the genomic basis of co‐evolutionary interactions in nature.     

Fine‐scale population epigenetic structure in relation to gastrointestinal parasite load in red grouse (Lagopus lagopus scotica)

Epigenetic modification of cytosine methylation states can be elicited by environmental stresses and may be a key process affecting phenotypic plasticity and adaptation. Parasites are potent stressors with profound physiological and ecological effects on their host, but there is little understanding in how parasites may influence host methylation states. Here, we estimate epigenetic diversity and differentiation among 21 populations of red grouse (Lagopus lagopus scotica) in north‐east Scotland and test for association of gastrointestinal parasite load (caecal nematode Trichostrongylus tenuis) with hepatic genome‐wide and locus‐specific methylation states. Following methylation‐sensitive AFLP (MSAP), 129 bands, representing 73 methylation‐susceptible and 56 nonmethylated epiloci, were scored across 234 individuals. The populations differed significantly in genome‐wide methylation levels and were also significantly epigenetically (F_SC = 0.0227; P < 0.001) and genetically (F_SC = 0.0058; P < 0.001) differentiated. Parasite load was not associated with either genome‐wide methylation levels or epigenetic differentiation. Instead, we found eight disproportionately differentiated epilocus‐specific methylation states (F_ST outliers) using bayescan software and significant positive and negative association of 35 methylation states with parasite load from bespoke generalized estimating equations (GEE), simple logistic regression (sam ) and Bayesian environmental analysis (bayenv 2). Following Sanger sequencing, genome mapping and geneontology (go ) annotation, some of these epiloci were linked to genes involved in regulation of cell cycle, signalling, metabolism, immune system and notably rRNA methylation, histone acetylation and small RNAs. These findings demonstrate an epigenetic signature of parasite load in populations of a wild bird and suggest intriguing physiological effects of parasite‐associated cytosine methylation.     

Development of mandibular,hyoid and hypobranchial muscles in the zebrafish: homologies and evolution of these muscles within bony fishes and tetrapods

Background  

During vertebrate head evolution, muscle changes accompanied radical modification of the skeleton. Recent studies have suggested that muscles and their innervation evolve less rapidly than cartilage. The freshwater teleostean zebrafish (Danio rerio) is the most studied actinopterygian model organism, and is sometimes taken to represent osteichthyans as a whole, which include bony fishes and tetrapods. Most work concerning zebrafish cranial muscles has focused on larval stages. We set out to describe the later development of zebrafish head muscles and compare muscle homologies across the Osteichthyes.     

Profiling of Histidine Phosphoproteome in Danio rerio by TiO2 Enrichment

Histidine phosphorylation is a reversible post‐translational modification that is known to regulate signal transduction in prokaryotes. However, functional studies in eukaryotes have been largely neglected due to the labile nature of N‐linked phosphorylated amino acids. In an effort to help elucidate the heretofore hidden vertebrate phosphoproteome, this report presents a global phosphorylation analysis of Danio rerio (zebrafish) larvae. Phosphopeptide enrichment is performed using a TiO₂ affinity technique. A total of 68 unique phosphohistidine sites are detected on 63 proteins among 1076 unique phosphosites on 708 proteins. Data are available via ProteomeXchange with identifier PXD012735. This report provides the first phosphohistidine dataset obtained from zebrafish.     

A novel approach to parasite population genetics: Experimental infection reveals geographic differentiation,recombination and host‐mediated population structure in Pasteuria ramosa,a bacterial parasite of Daphnia

The population structure of parasites is central to the ecology and evolution of host‐parasite systems. Here, we investigate the population genetics of Pasteuria ramosa, a bacterial parasite of Daphnia. We used natural P. ramosa spore banks from the sediments of two geographically well‐separated ponds to experimentally infect a panel of Daphnia magna host clones whose resistance phenotypes were previously known. In this way, we were able to assess the population structure of P. ramosa based on geography, host resistance phenotype and host genotype. Overall, genetic diversity of P. ramosa was high, and nearly all infected D. magna hosted more than one parasite haplotype. On the basis of the observation of recombinant haplotypes and relatively low levels of linkage disequilibrium, we conclude that P. ramosa engages in substantial recombination. Isolates were strongly differentiated by pond, indicating that gene flow is spatially restricted. Pasteuria ramosa isolates within one pond were segregated completely based on the resistance phenotype of the host—a result that, to our knowledge, has not been previously reported for a nonhuman parasite. To assess the comparability of experimental infections with natural P. ramosa isolates, we examined the population structure of naturally infected D. magna native to one of the two source ponds. We found that experimental and natural infections of the same host resistance phenotype from the same source pond were indistinguishable, indicating that experimental infections provide a means to representatively sample the diversity of P. ramosa while reducing the sampling bias often associated with studies of parasite epidemics. These results expand our knowledge of this model parasite, provide important context for the large existing body of research on this system and will guide the design of future studies of this host‐parasite system.     

Laser surgery of zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) embryos using femtosecond laser pulses: Optimal parameters for exogenous material delivery,and the laser's effect on short- and long-term development

Background  

Femtosecond (fs) laser pulses have recently received wide interest as an alternative tool for manipulating living biological systems. In various model organisms the excision of cellular components and the intracellular delivery of foreign exogenous materials have been reported. However, the effect of the applied fs laser pulses on cell viability and development has yet to be determined. Using the zebrafish (Danio rerio) as our animal model system, we address both the short- and long-term developmental changes following laser surgery on zebrafish embryonic cells.     

利用Tol2转座子介导的增强子诱捕技术获得血管相关转基因斑马鱼系

心血管系统形成于胚胎发育极早期并为其他器官的发育、维持、修复所必需,血管生长异常可造成多种疾病.然而,由于研究对象所限,胚胎血管的发育机制尚未完全阐明,调控血管发育的基因也所知有限.通过Tol2转座子介导的大规模增强子诱捕筛选到26个血管特异表达绿色荧光蛋白(EGFP)报告基因的转基因斑马鱼系,其中有一些品系在胚胎的某些特异血管结构中表达绿色荧光.通过linker-mediated PCR克隆到22个鱼系中Tol2插入位点附近的斑马鱼基因组序列,其中有17个鱼系的Tol2插入可定位到现有的斑马鱼基因组中的单一位点.通过整体胚胎原位杂交对插入位点附近的基因进行表达谱分析,得到8个表达谱与转基因鱼系一致的基因,涵盖了9个鱼系,其中dusp5基因对应于2个不同的鱼系.这8个基因中包括hhex、ets1a和dusp5等3个功能已知的基因,但是大部分(5个)基因在斑马鱼中尚无功能研究,分别为zvsg1、micall2a、arl8b(1of2)、zgc:73355以及hecw2(1of2).hhex和ets1a基因对血管与血细胞前体的发育具有重要作用,所获得的EGFP报告基因受hhex或ets1a基因增强子控制的转基因斑马鱼(mp378b和mp430c-2)为国际首例,为深入研究这两个基因在血管与血液发育中的作用机制提供了新的机遇.筛选到的功能未知基因可以用来进一步研究其在血管发育中的功能;同时,利用所获得的转基因鱼系,可以实现实时、动态观察成血管细胞的起源、分化与基因表达调控,并可用于高通量小分子药物筛选等重要研究.     

Insight into the evolution and functional characteristics of the pan‐genome assembly from sesame landraces and modern cultivars

Sesame (Sesamum indicum L.) is an important oil crop renowned for its high oil content and quality. Recently, genome assemblies for five sesame varieties including two landraces (S. indicum cv. Baizhima and Mishuozhima) and three modern cultivars (S. indicum var. Zhongzhi13, Yuzhi11 and Swetha), have become available providing a rich resource for comparative genomic analyses and gene discovery. Here, we employed a reference‐assisted assembly approach to improve the draft assemblies of four of the sesame varieties. We then constructed a sesame pan‐genome of 554.05 Mb. The pan‐genome contained 26 472 orthologous gene clusters; 15 409 (58.21%) of them were core (present across all five sesame genomes), whereas the remaining 41.79% (11 063) clusters and the 15 890 variety‐specific genes were dispensable. Comparisons between varieties suggest that modern cultivars from China and India display significant genomic variation. The gene families unique to the sesame modern cultivars contain genes mainly related to yield and quality, while those unique to the landraces contain genes involved in environmental adaptation. Comparative evolutionary analysis indicates that several genes involved in plant‐pathogen interaction and lipid metabolism are under positive selection, which may be associated with sesame environmental adaption and selection for high seed oil content. This study of the sesame pan‐genome provides insights into the evolution and genomic characteristics of this important oilseed and constitutes a resource for further sesame crop improvement.