Histologic Findings in Captive American Horseshoe Crabs (Limulus polyphemus)

Abstract

Histopathology of 61 captive American horseshoe crabs (HSCs; Limulus polyphemus) is reviewed. HSC organs evaluated histologically included body wall (chitin, epidermis, dermis, and skeletal muscle), hepatopancreas, gut, gonads, book gills, eyes, heart, brain, and coxal gland. In descending order, lesions were most frequently identified in compound eye, body wall, book gills, hepatopancreas, chitinous gut, nonchitinous gut, heart, and brain; lesions were not observed in coxal gland or gonads. Hemocytes (also called amoebocytes) surrounded infectious agents and occluded ulcers. Large hemocyte aggregates had a central eosinophilic coagulum (ie, hemocyte coagulum). Cutaneous ulceration (34/60 cases), branchitis (29/48 cases), and ophthalmitis (17/20 cases) were common lesions and consistently associated with fungi, which were invasive into subjacent tissues, and/or bacteria, which were usually superficial. Fungal culture was performed in 3 cases and isolated Fusarium spp., although fungal morphology varied and multiple fungal species may have been present. Presumptive green algae were associated with ulceration in 1 case with minimal to no inflammation. Presumptive cyanobacteria were identified within a biofilm overlying the gills in 4 of 48 cases and were not invasive. Multifocal, random hepatopancreatitis was identified in 16 of 57 cases, 10 of which were associated with bacteria. Metacercarial cysts were identified in 25 of 61 cases and associated with minimal to no inflammation. Depleted eosinophilic globules in hepatopancreatic interstitial cells were interpreted as decreased nutritional status in 12 of 57 cases.

Microbiome shifts associated with the introduction of wild Atlantic horseshoe crabs (Limulus polyphemus) into a touch-tank exhibit

“…Vibrio is commonly found in the microbiomes of aquatic organisms and some species have been identified as pathogens, commensals, and mutualists, yet, an understanding of the relationships between many Vibrio species and their hosts requires further research (Thompson et al, 2016). A histological analysis of Limulus identified several bacterial species associated with pathological conditions including Pseudomonas and Aeromonas (LaDouceur et al, 2019) which also appear to be major constituents of what we suspect is a pathological condition in our study, as well. A more comprehensive view of a core horseshoe crab microbiome awaits cultivation-independent surveys of other horseshoe crab species, and more individuals from different geographic and physicochemical environments (e.g., estuarine and marine).…”

“…Once the exoskeleton of the horseshoe crab has been breached (or degraded), it would then be susceptible to secondary infections and given that adults do not molt, the damage accumulated is irreparable. Previous studies have associated infections in captive lab horseshoe crab populations with eukaryotic parasites, algae, fungi, and bacteria (commonly Cyanobacteria and Gram-negative bacteria) (Leibovitz and Lewbart 2004, Smith 2006; Braverman et al, 2012, LaDouceur et al, 2019). We hypothesize that the appearance of these lesions could be due to dysbiosis of the horseshoe crab microbiome, resulting in an overall increase in the abundance of chitinolytic bacteria or the appearance of opportunistic pathogens capable of degrading chitin.…”

“…Additionally, several species of Pseudoalteromonas are widely discussed in the literature as symbionts, given their diverse metabolic potential and their ability to produce a variety of antibacterial, antifungal, algicidal, and antifouling compounds (Atencio et al 2018; Offret et al 2016). Fungi have been identified as possible chitinase containing pathogens that could be involved in this process (Tuxbury et al, 2014; LaDouceur et al, 2019). In our study, we excluded eukaryotic sequences from analysis and therefore were unable to document the fungal communities of our samples.…”

Microbiome Shifts Associated With the Introduction of Wild Atlantic Horseshoe Crabs (Limulus polyphemus) Into a Touch-Tank Exhibit

“…This is in direct opposition to a study on cownose rays in touch tanks, which concluded that the transfer of human-associated taxa to the rays was negligible ( Kearns et al, 2017 ). Cultural analyses of hemolymph samples from captive Limulus resulted in the isolation of several bacterial species associated with pathological conditions, including Shewanella putrefaciens (formerly Pseudomonas putrefaciens ) and Aeromonas hydrophila ( Khashe and Janda, 1998 ; LaDouceur et al, 2019 ). We hypothesize that the unclassified species of Pseudomonas and Aeromonas that dominate our tank-acclimated population are involved somehow in the development of the diseased state in our study.…”

“…Once the exoskeleton of the horseshoe crab has been breached (or degraded), it would then be susceptible to secondary infections and given that adults do not molt, the damage accumulated is irreparable. Previous studies have associated infections in captive lab horseshoe crab populations with eukaryotic parasites, algae, fungi, and bacteria (commonly Cyanobacteria and Gram-negative bacteria) ( Leibovitz and Lewbart, 2003 ; Smith, 2007 ; Braverman et al, 2012 ; LaDouceur et al, 2019 ). We hypothesize that the appearance of these lesions could be due to shifts in the horseshoe crab microbiome, resulting in an overall increase in the abundance of chitinolytic bacteria or the appearance of opportunistic pathogens capable of degrading chitin.…”