title: Mesoscale morphology at nanoscale resolution: serial block-face scanning electron microscopy reveals fine 3D detail of a novel silk spinneret system in a tube-building tanaid crustacean.
abstract: BACKGROUND
  The study of morphology is experiencing a renaissance due to rapid improvements in technologies for 3D visualization of complex internal and external structures. But 3D visualization of the internal structure of mesoscale objects - those in the 10-1000 μm range - remains problematic. They are too small for microCT, many lack suitable specific fluorescent markers for confocal microscopy, or they require labor-intensive stacking and smoothing of individual TEM images. Here we illustrate the first comprehensive morphological description of a complete mesoscale biological system at nanoscopic resolution using ultra-modern technology for 3D visualization - serial block-face scanning electron microscopy (SBF-SEM). The SBF-SEM machine combines an in-chamber ultramicrotome, which creates a serial array of exposed surfaces, with an SEM that images each surface as it is exposed. The serial images are then stacked automatically by 3D reconstruction software. We used SBF-SEM to study the spinneret (thread-producing) system of a small, tube-dwelling crustacean that weaves tubes of silk. Thread-producing ability is critical for the survival of many small-bodied animals but the basic morphology of these systems remains mysterious due to the limits of traditional microscopy.
  
  
  RESULTS
  SBF-SEM allowed us to describe - in full 3D - well-resolved components (glands, ducts, pores, and associated nerves and muscles) of the spinneret system in the thoracic legs and body segments of Sinelobus sp. (Crustacea, Peracarida, Tanaidacea), a tube-building tanaid only 2 mm in body length. The 3D reconstruction by SBF-SEM revealed at nanoscale resolution a unique structure to the gland and duct systems: In each of three thread-producing thoracic segments, two separate ducts, derived from two separate glands located in the body, run through the entire leg and merge at the leg tip just before the spinneret pore opening. We also resolved nerves connecting to individual setae, spines and pores on the walking legs, and individual muscles within each leg segment.
  
  
  CONCLUSIONS
  Our results significantly expand our understanding of the diversity of spinneret systems in the Crustacea by providing the first well-resolved view of spinneret components in the peracarid crustacean order, Tanaidacea. More significantly, our results reveal the great power of SBF-SEM technology for comprehensive studies of the morphology of microscopic animals.
authors.affiliation: Systematics and Evolution Group, Department of Biological Sciences, University of Alberta, Edmonton, AB T6G 2E9 Canada ; Allgemeine & Spezielle Zoologie, Institut fuer Biowissenschaften, Universität Rostock, Rostock, 18055 Germany.
authors.forename: Tomonari
authors.initials: T
authors.lastname: Kaji
authors.affiliation: Department of Biological Sciences, Faculty of Science, Hokkaido University, Sapporo, 060-0810 Japan.
authors.forename: Keiichi
authors.initials: K
authors.lastname: Kakui
authors.affiliation: National Institute for Physiological Sciences, Okazaki, Aichi 444-8585 Japan.
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Miyazaki
authors.affiliation: National Institute for Physiological Sciences, Okazaki, Aichi 444-8585 Japan.
authors.forename: Kazuyoshi
authors.initials: K
authors.lastname: Murata
authors.affiliation: Systematics and Evolution Group, Department of Biological Sciences, University of Alberta, Edmonton, AB T6G 2E9 Canada.
authors.forename: A Richard
authors.initials: AR
authors.lastname: Palmer
keywords: Arthropoda; Comparative morphology; High-resolution imaging; Renaissance of morphology; Serial block-face scanning electron microscope; Silk gland morphology; Tanaidacea; Thread secretion; Three-dimensional imaging