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Morphology and Ultrastructure of Brachymystax Lenok Tsinlingensis

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G Model YTICE-1010; No. of Pages 7 ARTICLE IN PRESS Tissue and Cell xxx (2016) xxx–xxx Contents lists available at ScienceDirect Tissue and Cell journal homepage: www.elsevier.com/locate/tice Morphology and ultrastructure of Brachymystax lenok tsinlingensis spermatozoa by scanning and transmission electron microscopy a,b,c c,d c,e c a,c,d,f,∗ Wei Guo , Jian Shao , Ping Li , Jinming Wu , Qiwei Wei a Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China b University of Chinese Academy of Sciences, Beijing 100039, China c Key Laboratory of Freshwater Biodiversity Conservation, Ministry of Agriculture of China, Yangtze River Fisheries Research Institute, Chinese Academy of Fishery Sciences, No. 8, 1st Wudayuan Road, Donghu Hi-tech Development Zone, Wuhan 430223, China d College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China e University of South Bohemia in Ceskéˇ Budejovice,ˇ Faculty of Fisheries and Protection of Waters, South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses, Research Institute of Fish Culture and Hydrobiology, Zátisíˇ 728/II, 389 25 Vodnany,ˇ Czech Republic f Freshwater Fisheries Research Center, Chinese Academy of Fisheries Science, Wuxi 214081, China a r t i c l e i n f o a b s t r a c t Article history: This study was conducted to investigate Brachymystax lenok tsinlingensis spermatozoa cell morphology Received 6 March 2016 and ultrastructure through scanning and transmission electron microscopy. Findings revealed that the Received in revised form 28 May 2016 spermatozoa can be differentiated into three major parts: a spherical head without an acrosome, a short Accepted 28 May 2016 mid-piece, and a long, cylindrical flagellum. The mean length of the spermatozoa was 36.11 ± 2.84 ␮m, Available online xxx with a spherical head length of 2.78 ± 0.31 ␮m. The mean anterior and posterior head widths were ± 2.20 0.42 ␮m and 2.55 ± 0.53 ␮m, respectively. The nuclear fossa was positioned at the base of the Keywords: nucleus that contained the anterior portion of flagellum and a centriolar complex (proximal and distal Morphology Ultrastructure centrioles). The short mid-piece was located laterally to the nucleus and possessed just one spherical mitochondrion with a mean diameter of 0.65 ± 0.14 ␮m. The spermatozoa flagellum was long and cylin- Brachymystax lenok tsinlingensis Spermatozoa drical, and could be separated into two parts: a long main-piece and a short end-piece. The main piece of Scanning electron microscopy the flagellum had short irregular side-fins. The axoneme composed the typical ‘9 + 2’ microtubular dou- Transmission electron microscopy blet structure and was enclosed by the cell membran e. This study confirmed that B. lenok tsinlingensis spermatozoa can be categorized as teleostean “Type I” spermatozoa; ‘primitive’ or ‘ect-aquasperm type’ spermatozoa. To the best of the authers knowledge, this was the first study conducted on the morphology and ultrastructure of B. lenok tsinlingensis spermatozoa. © 2016 Elsevier Ltd. All rights reserved. 1. Introduction the spermatozoa structure has revealed a high diversity, predom- inately between systematic families. This diversity is reflected in Spermatozoa transport the male haploid chromosome set into the head shape, in the number, shape and location of mitochondra, the oocyte. For this purpose, spermatozoa compose four differ- and in the number, length and structure of the flagellum. The sper- ent compartments: the acrosome, the head, the mid-piece, and matozoa structure among Osteichthyes is diverse. Spermatozoa of the tail or flagellum (Callard and Callard, 1999; Knobil and Neill, Actinistia, Dipnoi, and Chondrostei spp. contain an acrosome, have 1999). Spermatozoa structure in teleost fish is influenced by both an elongated head, and a nucleus pervaded by a channel along its the mode of reproduction and systematic position of the species longitudinal axis. The mid-piece is small in species with external (Knobil and Neill, 1999). In the teleost species studied to date, fertilization (Dipnoi and Chondrostei spp.), but is well developed and of comparable size to those of elasmobranch fish in species with internal fertilization (Actinistia spp.) (Jamieson, 1991; Mattei, ∗ 1991). The head and mid-piece of the spermatozoa in Neopterygii Corresponding author at: Key Laboratory of Freshwater Biodiversity Conser- spp. is very small (≤10 ␮m), the latter contains mitochondrion and vation, Ministry of Agriculture of China, Yangtze River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Institute of Hydrobiology, No. 8, 1st Wudayuan is pervaded by a cytoplasmic channel. The structure of the flagel- Road, Donghu Hi-tech Development Zone, Wuhan 430223, China. lum typically exhibits a ‘9 + 2’ microtubule pattern and may include E-mail addresses: [email protected] (W. Guo), lateral side-fins that can be considered as paddles that enhance the [email protected] (J. Shao), liping@yfi.ac.cn (P. Li), jinming@yfi.ac.cn (J. Wu), swimming ability of the flagellum (Lahnsteiner and Patzner, 2008). weiqw@yfi.ac.cn (Q. Wei). http://dx.doi.org/10.1016/j.tice.2016.05.009 0040-8166/© 2016 Elsevier Ltd. All rights reserved. Please cite this article in press as: Guo, W., et al., Morphology and ultrastructure of Brachymystax lenok tsinlingensis spermatozoa by scanning and transmission electron microscopy. Tissue Cell (2016), http://dx.doi.org/10.1016/j.tice.2016.05.009 G Model YTICE-1010; No. of Pages 7 ARTICLE IN PRESS 2 W. Guo et al. / Tissue and Cell xxx (2016) xxx–xxx Table 1 Data of the broodfish and the spermatozoa. Samples Total Body Weight (g) Density of the length (cm) length (cm) spermatozoa (1010/ml) Male 1 22.9 19.4 116.7 2.65 ± 0.072 Male 2 23.9 19.8 135.8 3.07 ± 0.043 Male 3 21.9 18.7 93.4 2.55 ± 0.025 Male 4 28.7 25.1 247.2 3.15 ± 0.110 Fig. 1. Ultrastructure of Brachymystax lenok tsinlingensis spermatozoa: head (H); mid-piece (MP); mitochondrion (M); flagellum (F). Scale bar: 10 ␮m. Teleost spermatozoa exhibit a diverse range of structural fea- tures that makes it difficult to depict a common sperm type (Mattei, 1991). The structure of fish spermatozoa varies between families: 2.2. Ultrastructural study from aflagellate to biflagellae, while shape, size, and structure can vary significantly according to whether a species adopts internal Electron microscopy studies were carried out at the Laboratory or external fertilization (Jones and Butler, 1988). Spermiogenesis is of Electron Microscopy of the Institute of Hydrobiology, Chinese broadly categorized into two types (I and II); however, in teleosts it Academy of Sciences. shows a wide variety of patterns (Mattei, 1991). In Type I, rotation The samples were fixed in 0.1 M cacodylate buffer (pH 7.5) con- ◦ of the nucleus occurs, the diplosome enters the nuclear fossa, and taining 2.5% glutaraldehyde (Karnovsky, 1965) for 2 h at 4.0 C. the flagellum is symmetrically located, while for in Type II, there is The samples were then washed in cacodylate buffer for 1 h, and no nuclear rotation, the diplosome remains outside the fossa, and post-fixed in 1% cacodylate-buffered osmium tetroxide for 1 h at ◦ the flagellum is asymmetrically located (Mattei, 1970). 4.0 C. Following double fixation, the spermatozoa pellets were pro- B. lenok tsinlingensis (Qinling lenok) is only found in the cold- cessed for SEM and TEM. For SEM, the spermatozoa samples were water mountain streams and rivers of Qinling Mountains, Shaanxi, dehydrated through an ascending series of ethanol (30%, 50%, 70%, China. The fish is listed as a second class state-protected wild ani- 80%, 100%, 10mins for every concentration and doubled for 100%), mal in the China Red Data Book of Endangered Animal (1988). The and the dehydrated samples were critical-point dried, mounted on fine structures of the testis in B. lenok tsinlingensis have been stud- specimen holders, and sputter-coated with 20 nm gold palladium. ied by transmission and scanning electron microscopy (TEM and Preparations were examined using a SEM (Hitachi S-4800, Tokyo, SEM, respectively), testicular structure is a lobular type, containing Japan) at 20 kV (Greven and Schmahl, 2006). For TEM, the sam- numerous cysts surrounded by a layer of Sertoli cells (Zhang and ples were dehydrated as above, and embedded in epoxide resin. Wang, 1992). Despite the importance of this species, very little is Ultra-thin sections, 60–100 nm thick, were collected using glass known about its reproductive biology. Reports on the breeding of knives. The sections were then placed on copper grids, stained with this fish in captivity are lacking (Liu et al., 2013) and hatchery pro- uranyl and lead citrate, and screened under a TEM (Hitachi HT-700) duction of this species is yet to be developed for large-scale farming. (Ravaglia and Maggese, 2003). Considering the importance of protecting this fish, it is essential All SEM and TEM measurements were evaluated using MicroIm- to understand its reproductive biology. Consequently, the present age software (version 4.0.1 for Windows, Olympus Optical Co., study aimed to investigate the ultrastructure of B. lenok tsinlingensis Hamburg, Germany). Spermatozoa morphological characteristics ± spermatozoa using SEM and TEM. were assessed and expressed as mean standard deviation, num- ber, and range. The experiment of SEM and TEM has been repeated four times. 2. Materials and methods 3. Results 2.1. Broodfish sperm collection Morphologically, the spermatozoa of B. lenok tsinlingensis com- prised three major parts: a head without an acrosome, a mid-piece, Fish were collected from the wild environment in Taibai River, and a single flagellum (Fig. 1). Morphological measurements of the located in the Baoji of Shaanxi province, China. They were placed in spermatozoa are shown in Table 2. The mean total length of the broodfish tanks in a hatchery for 10 days prior to spawning induc- spermatozoa was 36.11 ± 2.84 ␮m (n = 29).
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  • Complete Mitochondrial Genomes of the Cherskii's Sculpin Cottus

    Complete Mitochondrial Genomes of the Cherskii's Sculpin Cottus

    EVB0010.1177/1176934317726783Evolutionary BioinformaticsBalakirev et al 726783research-article2017 Evolutionary Bioinformatics Complete Mitochondrial Genomes of the Cherskii’s Volume 13: 1–7 © The Author(s) 2017 Sculpin Cottus czerskii and Siberian Taimen Hucho Reprints and permissions: sagepub.co.uk/journalsPermissions.nav taimen Reveal GenBank Entry Errors: Incorrect DOI:https://doi.org/10.1177/1176934317726783 10.1177/1176934317726783 Species Identification and Recombinant Mitochondrial Genome Evgeniy S Balakirev1,2,3, Pavel A Saveliev2 and Francisco J Ayala1 1Department of Ecology and Evolutionary Biology, University of California, Irvine, Irvine, CA, USA. 2A.V. Zhirmunsky Institute of Marine Biology, National Scientific Center of Marine Biology, Far Eastern Branch, Russian Academy of Sciences, Vladivostok, Russia. 3School of Natural Sciences, Far Eastern Federal University, Vladivostok, Russia. ABSTRACT: The complete mitochondrial (mt) genome is sequenced in 2 individuals of the Cherskii’s sculpin Cottus czerskii. A surprisingly high level of sequence divergence (10.3%) has been detected between the 2 genomes of C czerskii studied here and the GenBank mt genome of C czerskii (KJ956027). At the same time, a surprisingly low level of divergence (1.4%) has been detected between the GenBank C czerskii (KJ956027) and the Amur sculpin Cottus szanaga (KX762049, KX762050). We argue that the observed discrepancies are due to incorrect taxonomic identification so that the GenBank accession number KJ956027 represents actually the mt genome of C szanaga erroneously identified as C czerskii. Our results are of consequence concerning the GenBank database quality, highlighting the potential negative consequences of entry errors, which once they are introduced tend to be propagated among databases and subsequent publications.
  • Fish Hosts, Glochidia Features and Life Cycle of the Endemic

    Fish Hosts, Glochidia Features and Life Cycle of the Endemic

    www.nature.com/scientificreports OPEN Fish hosts, glochidia features and life cycle of the endemic freshwater pearl mussel Margaritifera dahurica Received: 12 November 2018 Accepted: 23 May 2019 from the Amur Basin Published: xx xx xxxx Ilya V. Vikhrev 1,2,3, Alexander A. Makhrov 3,4, Valentina S. Artamonova3,4, Alexey V. Ermolenko5, Mikhail Y. Gofarov1,2, Mikhail B. Kabakov2, Alexander V. Kondakov1,2,3, Dmitry G. Chukhchin1, Artem A. Lyubas1,2 & Ivan N. Bolotov 1,2,3 Margaritiferidae is a small freshwater bivalve family with 16 species. In spite of a small number of taxa and long-term history of research, several gaps in our knowledge on the freshwater pearl mussels still exist. Here we present the discovery of host fshes for Margaritifera dahurica, i.e. Lower Amur grayling, sharp-snouted lenok, and blunt-snouted lenok. The host fshes were studied in rivers of the Ussuri Basin. The identifcation of glochidia and fsh hosts was confrmed by DNA analysis. The life cycle of M. dahurica and its glochidia are described for the frst time. The SEM study of glochidia revealed that the rounded, unhooked Margaritifera dahurica larvae are similar to those of the other Margaritiferidae. Margaritifera dahurica is a tachytictic breeder, the larvae of which attach to fsh gills during the Late August – September and fnish the metamorphosis in June. Ancestral host reconstruction and a review of the salmonid - pearl mussel coevolution suggest that the ancestral host of the Margaritiferidae was a non-salmonid fsh, while that of the genus Margaritifera most likely was an early salmonid species or their stem lineage.