Use of laser scanning confocal microscopy for morphological taxonomy and the potential for digital type specimens (e-types)

Abstract

The use of laser scanning confocal microscopy (LSCM) for creating taxonomic descriptions of copepods is investigated. A new technique is described, which employs a contour filter to process digital LSCM images, allowing taxonomic information to be quickly and accurately distilled into a simple illustration. LSCM allows the imaging of whole specimens, which can be rotated and viewed from any angle -a major benefit over light microscopy. Using this technique, it is suggested that taxonomic descriptions can be rapidly produced in a fraction of the time required to produce similar descriptions using traditional light microscopy and hand drawing techniques. Good staining of specimens is, however, essential to produce accurate descriptions and more research is required in this area. The use of LSCM for morphological taxonomy shows great potential, not only for producing taxonomic descriptions, but also providing a complementary adjunct to traditional type specimens in the form of 3D digital 'e-types' deposited in recognised international databases. KEY WORDS: E-type · Digital taxonomy · Confocal Resale or republication not permitted without written consent of the publisherAquat Biol 14: [165][166][167][168][169][170][171][172][173] 2012 MATERIALS AND METHODSWhiting Merlangius merlangus (L.) and, to a lesser degree, cod Gadus morhua (L.) infected with Lernaeocera branchialis were sampled from the catches of commercial demersal trawlers working within the River Forth Estuary at Kincardine, Scotland (56°02' 53' N, 3°40' 59' W). To culture the juvenile stages of L. branchialis, egg strings were dissected from gravid female parasites collected from infected whiting and cod. These were then maintained under aeration in 500 ml beakers kept in a WTB Binder Labortechnik precision environmental chamber at 10°C. Once the eggs began to hatch they were placed in a beaker of fresh aerated sea water (35 ppt) and after a period of time (20 min to 24 h), the egg strings were transferred to another beaker of fresh sea water, leaving behind a 'batch' of nauplii. These batches were then used for experiments as either nauplii or copepodids, depending on how long they were maintained.Lernaeocera branchialis specimens for traditional light microscopy were prepared by clearing individuals in 85% lactic acid (Sigma L1250) for 30 min, which also softened the connective tissues, and then by dissecting them on a cavity slide using fine mounted needles under a dissecting microscope. Individual appendages from each specimen were mounted in 100% glycerol (Sigma G7757) and then sealed with clear nail varnish once a coverslip had been placed over them. Both whole and dissected specimens were viewed on an Olympus BX51 compound microscope and digital micrographs were taken using a Zeiss AxioCam MRc camera and MRGrab 1.0.0.4 software (Carl Zeiss Vision, 2001), these being used as the basis for initial drawings.Specimens for LSCM were prepared by fixing them in 2.5% glutaraldehyde (>1 wk to enhance autofluorescence), and then rinsing them ...

Description of the free-swimming juvenile stages of Lernaeocera branchialis (Pennellidae), using traditional light and confocal microscopy methods

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Challenges and Advances in the Taxonomy of Deep-Sea Peracarida: From Traditional to Modern Methods

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SEM‐microphotogrammetry, a new take on an old method for generating high‐resolution 3D models from SEM images

“…Confocal microscopy of arthropods, typically through the use of stains to enhance fluorescence (Michels, 2007;Lee et al, 2009;Valdecasas & Abad, 2011;Brooker et al, 2012;Kamanli et al, 2017), is an emerging technique which is proving of great utility for the description of macroinvertebrates. The 3D models, of these specimens, that confocal microscopy yields have good resolution (often submicron) and successfully capture fine-scale structures including setae and finescale surface topography which may be only a few hundreds of nanometers in diameter (Fig.…”