W&M ScholarWorks Reports 8-1-1977 Histopathological Studies on Twelve Benthic Invertebrates from the Middle Atlantic Outer Continental Shelf Craig L. Ruddell Virginia Institute of Marine Science Follow this and additional works at: https://scholarworks.wm.edu/reports Part of the Marine Biology Commons Recommended Citation Ruddell, C. L. (1977) Histopathological Studies on Twelve Benthic Invertebrates from the Middle Atlantic Outer Continental Shelf. Special Reports in Applied Marine Science and Ocean Engineering (SRAMSOE) No. 179. Virginia Institute of Marine Science, College of William and Mary. https://doi.org/10.21220/ V5C452 This Report is brought to you for free and open access by W&M ScholarWorks. It has been accepted for inclusion in Reports by an authorized administrator of W&M ScholarWorks. For more information, please contact [email protected]. HISTOPATHOLOGICAL STUDIES ON TWELVE BENTHIC INVERTEBRATES FROM THE MIDDLE ATLANTIC OlITER CONTINENTAL SHELF Craig L. Ruddell Special Report in Applied Marine Scie:nce and Ocean Engineering No. 179 of the Virginia Institute of Marine Science Gloucester Point, Virginia 23062 This report was extracted from "Middle Atlantic Outer Continental Shelf Environmental Studies: Volume II. Chemical and Biological Benchmark Studies", the final report to the Bureau of Land Management, U. S. Department of Interior, pre­ pared by the Virginia Institute of Marine: Science under Contract No. 08550-CT-5-42. References to other chapters and appendices of the complete report have been left intact in this extract. August 1977 CHAPTER 10 TABLE OF CONTENTS INTRODUCTION ..... 10-1 METHODS AND MATERIALS . 10-1 Overview ..... 10-1 Organisms Chosen for Analysis ... 10-1 Processing of Samples on Shipboard .... 10-2 Organs and Tissues Sampled. 10-2 Fixation ............ 10-2 Numbers of Animals Sampled 10-3 Data Collection .... 10-3 Mensuration .. 10-3 Laboratory Processing .. 10-4 Preparation of Tissues for Sectioning 10-4 Material Preserved in Dietrich's Fixative . 10-4 Material Preserved in Aldehyde Mixtures 10-4 Sectioning ..... 10-4 Paraffin Blocks .. 10-4 Plastic Blocks . 10-4 Staining ...... .. 10-4 Paraffin Blocks ... 10-4 Glycol Methacrylate Sections . ... 10-7 Araldite Sections . 10-7 Storage of Sections and Blocks . 10-7 Data Recording . 10-7 Field Data . 10-7 Laboratory Data . 10-8 RESULTS ... 10-8 Crustacea .... ... 10-8 Dichelopandalus leptoceras .... 10-8 Distribution and Length-Frequency Data ... 10-12 Gonads and Gonadal Maturation . 10-12 Symbionts . 10-12 Crangon septemspinosa ......... 10-17 Distribution and Length-Frequency Data . 10-17 Gonads and Gonadal Maturation . 10-17 Symbionts . 10-19 Pontophilus brevirostris .......... 10-19 Distribution and Length-Frequency Data. 10-19 Gonads and Gonadal Maturation . 10-20 Symbionts . ...... 10-20 Cancer irroratus ............ 10-20 Distribution and Length-Frequency Data . 10-22 Gonads and Gonadal Maturation . 10-22 Symbionts and Lesions ....... 10-22 Cancer borealis ............. 10-24 Distribution and Basic Length-Frequency Data .. 10-24 Gonads and Gonadal Maturation . 10-24 Symbionts . 10-26 Molluscs . 10-26 Astarte casta:nea. 10-26 Distribution. 10-26 Gonads and Gonadal Maturity . 10-26 Symbionts 10-26 Astarte undata . ... .•... 10-28 Distribution ....... ..... 10-28 Gonads and Gonadal Maturity . 10-28 Symbionts . .. 10-28 PZacopecten magellanicus ... • 10-28 Distribution ........ ..... 10-29 Gonads and Gonadal Maturity 10-29 Symbionts . 10-29 Echinoderms .... ..... 10-29 Echinarachnius parma 10-29 Distribution . 10-29 Gonads and Gonadal Maturity .... .... 10-29 Symbionts .... .... 10-32 Astropecten americanus .... 10-32 Distribution and Size ... 10-32 Gonads and Gonadal Maturity .... 10-32 Symbionts . ... .... 10-34 Asterias forbesi . ........... .... 10-34 Distribution ........... 10-34 Gonads and Gonadal Maturity .... 10-34 Symbionts ........ 10-34 Asterias vulgaris .... 10-34 Distribution. ....... 10-34 Gonads and Gonadal Maturation . 10-34 Symbionts .... 10-34 DISCUSSION . 10-36 Summary of Significant Findings . 10-42 ACKNOWLEDGEMENTS .. 10-43 LITERATURE CITED . 10-43 CHAPTER 10 HISTOPATHOLOGICAL STUDIES Craig Rud de 11 INTRO DUCT ION A number of investigators have shown that acute and chronic pollution of the marine environment with crude or waste oils resulted in the appear­ ance of histologically demonstrable lesions in fish and shellfish (Blanton and Robinson 1973; Barry 1974; Barry and Yevich 1975; Gardner 1975; Gardner et al. 1975; Yevich 1975a, b). This work suggested that profound physiological changes and dysfunctions may have occurred in animals sub­ jected to these compounds. The work presented below was prompted by these studies an<l was undertaken in an effort to define the normal or baseline histology of a number of common benthic marine invertebrates found in the vicinity of the Middle Atlantic oil drilling lease areas. It is expected that this work will be useful in evaluating the effects of environmental contamination associated with future gas and oil production in these areas. The material presented below will review some of the significant findings of the histopathological effort; more specifically, this report will focus on (1) the distribution of the organisms chosen for study, (2) details relating to the production of gametes by these organisms, and (3) the symbionts associated with these organisms. A histological atlas depicting the major organ systems, parasites, and pathologies of these invertebrates will be presented at a later time. METHODS AND MATERIALS Overview A number of benthic marine invertebrates were collected from the dredge and trawl stations during each of the four cruises (see Chapter 2 for locations and general framework). Portions of these animals were then pre­ pared for histological examination. Sections of these animals were ex­ amined in an effort to deter~ine baseline histological data. Details of these procedures are presented below. Organisms Chosen for Analysis Twelve benthic marine invertebrates were chosen for histopathological analysis. These organisms were selected because: (1) they were representative of the fauna found in a given area; (2) they could be captured by the gear available to us; (3) they occurred in numbers sufficient to satisfy the needs not only of the histopathology effort but also of the chemists; (4) they could 10-1 be captured and brought to the surface in a relatively untraumatized condition; (5) they represented diverse feeding types; and (6) some of the organisms, i.e. Placopecten megallanicus, represented species of commercial importance. The organisms chosen for analysis included: Molluscs: Astarte undata, A. castanea, and Placopecten magellanicus Shrimp: DicheZopandaZus leptoceras, PontophiZus brevirostris, andCrangon septemspinosa Crabs: Cancer irroratus and C. borealie Echinoderms: Echinarachnius parma, Asterias forbesi, A. vuZgaris and Astropecten americanus Processing of Samples on Shipboard Organs and Tissues Sampled For routine work, the smaller crustacea, specifically the shrimp and small crabs, were fixed whole without any attempt to dissect out discrete organ systems. Similarly, the echinoderms and small molluscs were reduced or dissected to yield only broad anatomical units such as the arms or disk area in the echinoderms or slabs of gut, mantle, and foot in the smaller molluscs. The larger crabs and the giant scallop, P. mageZZanicus, were dissected to provide portions of gill, digestive diverticula, stomach (crabs only), gonad, muscle, heart, mantle (scallops only), or kidney (scallops only). Fine dissections of other animals were occasionally performed when it was desired to obtain material for plastic embedding (see below). Fixation. Samples to be processed in the "routine" manner were preserved in Dietrich's fluid (9000 ml distilled water, 4500 ml 95% ethanol, 1500 ml 40% formalin, 300 ml glacial acetic acid). This fixative was recommended by Barozcz and Yevich (1975) as the fixative of choice for field work and ship­ board use because animals can be stored in this fixative for several months without undue hardening of tissues. In addition, Dietrich's fixative is easily prepared, penetrates tissues quickly, remains usable for long periods, even when maintained at room temperature, is inexpensive to prepare, and does not leave explosive or toxic residues on spilling. (Many of the classical histological fixatives do leave dangerous residues.) During the first (fall) cruise, as recommended by Barozcz and Yevich (1975), samples were placed in perforated "zip-lock" plastic bags, appropriately labeled, and immersed in large containers of fixative. This procedure proved to be somewhat tedious and wasteful of fixative, and resulted in a number of poorly preserved specimens. On all subsequent cruises, therefore, specimens were placed in small, 100 ml polyethylene screw-top bottles which had been filled with fixative one week prior to the cruise. 10-2 Although Dietrich's fluid was an adequate histological fixative, it did not preserve the fine structural elements of tissues and appeared to wash out glycogen from tissues. In order to appreciate the finer morphological details, we preserved selected portions of the animals referred to above in aqueous fixatives employing a variety of aldehydes as the primary fixative species. These fixatives included a phosphate-buffered, acrolein-formaldehyde mixture (PAF) and a two-step seawater acrolein-glutaraldehyde (SAG) formulation. PAF consisted of: 90 ml distilled water, 10 ml formalin (40% formaldehyde),