Amending the Description of Intraproboscis sanghae Amin, Heckmann, Sist, Basso, 2021 (Acanthocephala: Gigantorhynchidae) from a New Host, Phataginus tricuspis Rafinesque in Central Africa

Collections

The present study is based on 2 pseudo-segmented acanthocephalans collected from one white-bellied pangolin, P. tricuspis. The worms were extracted and preserved in 70% ethanol. These 2 worms were originally obtained from New Jersey vets working on an illegal shipment of pangolins from an unrecorded geographical location in central Africa. These worms ultimately made their way to Dr. Heather Walden at the University of Florida College of Veterinary Medicine at Gainesville, who provided them to us in December, 2020 at the Institute of Parasitic Diseases, Arizona for diagnosis and investigation. The specimens were routinely processed for microscopy and studied. We have subsequently transferred the custody of the 2 specimens to the University of Nebraska’s State Museum’s Harold W. Manter Laboratory (HWML), Lincoln collection for final deposit. We have attempted to obtain males of I. sanghae from P. tetradactyla and P. tricuspis in different locations in Central Africa but have been unsuccessful. Many specimens and other intestinal material collected from vomit and feces of these 2 host species in 2018 and 2020 were provided to us courtesy of Drs. Tamar Cassidy and Marine Montblanc of the Sangha Pangolin Project at Bayanga in the Central African Republic but they included no acanthocephalans.

Methods for Microscopical Studies

The worms were punctured with a fine needle and subsequently stained in Mayer’s acid carmine, destained in 4% hydrochloric acid in 70% ethanol, dehydrated in ascending concentrations of ethanol (24 hr each), and cleared in 100% xylene then in 50% Canada balsam and 50% xylene (24 hr each). Worms were then whole mounted in Canada balsam. Measurements are in micrometers or in mm as noted. The range is followed by mean values between parentheses. Width measurements represent maximum width. Trunk length does not include proboscis.

Line Drawing

Line drawings were created by using a Ken-A-Vision micro-projector (Ward’s Biological Supply Co., Rochester, N.Y.) which uses cool quartz iodine 150W illumination. Color-coded objectives, 10X, 20X, and 43X lenses, are used. Images of stained whole mounted specimens were projected vertically on 300 series Bristol draft paper (Starthmore, Westfield, Massachusetts), then traced and inked with India ink. Projected images were identical to the actual specimens being projected. The completed line drawings were subsequently scanned at 600 pixels, transferred on a USB and subsequently downloaded on a computer.

Optical Microscope Images

Optical microscope images were acquired using a BH2 light Olympus microscope (Olympus Optical Co., Osachi- shibamiya, Okaya, Nagano, Japan) attached to an AmScope 1000 video camera (United Scope LLC, dba AmScope, Irvine, California), linked to an ASUS laptop equipped with HDMI high-definition multimedia interface system (Taiwan- USA, Fremont, California). Images from the microscope are transferred from the laptop to a USB and stored for subsequent processing on a computer.

Deposits

The allotype male and paratype female were deposited in the University of Nebraska’s State Museum’s Harold W. Manter Laboratory (HWML) collection in Lincoln, Nebraska.

The Female

The female specimen was 81.25 mm long and 1.37 mm at its widest point. All other measurements fell within the range of those of the other females described by Amin, et al. [2] and noted in Table 1. This one specimen was, however, unique in having a partially retracted proboscis (Figure 1) which provided an opportunity to assess the operation of the receptacle which otherwise is totally embedded within the proboscis when normally extended as described in Amin, et al. [2].

The Male

This perfect specimen was incomplete posteriorly extending from the fully evaginated proboscis to the partially wrinkled posterior testis (Figure 2). The long lemnisci, Taenia-like pseudo-segmentation of trunk, and the imposing shape and size of the anterior testis were clearly demonstrable. Measurements of this male specimen are compared with those of the females of the same species as reported by Amin, et al. [2] (Table 1). Measurements of all structures in the male were smaller than those of the females but the lemnisci were slightly longer. Being the only male specimen of I. sanghae available, it is herein designated as the Allotype.

Taxonomic Summary

Host: Phataginus tricuspis Rafinesque. (Manidae) Type host: Phataginus tetradactyla Linn. (Manidae) Locality: Undetermined location in Central Africa. Type locality: SW Central African Republic at the Dzanga Sangha Protected Area (3°6′N 16°20′E). Site of infection:

Intestine.

Specimens: University of Nebraska’s State Museum’s Harold W. Manter Laboratory (HWML), Lincoln coll. no. 216916 (allotype male on one slide; Figure 2) and no. 216917 (paratype female on two slides; Figure 1). The HWML accession form included the following statements.

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Line drawings of a female and male specimens of Intraproboscis sanghae collected from Phataginus tricuspis in Central Africa. Figure 1: The anterior part of the female specimen showing the retracted anterior proboscis, the posterior displacement of the proboscis receptacle, and insertion of a lemniscus and the receptacle. Figure 2: The posteriorly incomplete male specimen showing the characteristic pseudo-segmentation, everted proboscis, long lemnisci, and complete anterior testis. “Additional notes: specimens were collected in New Jersey while working on an illegal shipment of pangolins.

The specimens were sent to Dr. Heather D. S. Walden (University of Florida College of Veterinary Medicine, Department of Comparative, Diagnostic and Population Medicine) who sent them to Omar Amin for further processing, identification and study. No detailed records are available on where and when the pangolins were originally collected except that they are from somewhere Central Africa.”

The Female

Structural details of the female specimen agreed closely with those of the other 4 females in the original description and its measurements fell within the range of reported measurements [2]. The proboscis was, however, partially retracted. The type of proboscis characteristic of these worms apparently only retracts but is not equipped to invaginate (Figure 1). Upon retraction, the receptacle appeared to get displaced more posteriorly extending well beyond the posterior end of the proboscis. The insertion and detail of the anteriorly nucleated lemnisci are newly documented. Compare the line drawing (Fig. 1) with the line drawing of the anterior part of a female specimen with totally everted proboscis in Amin, et al. [2].

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The Male

Although the posterior-most part of the male specimen was missing, the morphology and measurements of the anterior larger part of the specimen including the proboscis, hooks, receptacle and lemnisci were conclusive of its identity as I. sanghae (Fig. 2) and justifying its assignment as the allotype. This statement is more readily documented when considering the similarities of this male specimens with those of the females in Amin, et al. [2] (Figures 1-4 of anterior trunk, proboscis, hooks, receptacle, and lemnisci). Practically, all male measurements were comparable to those of female I. sanghae except for being somewhat smaller than those of the females (Table 1) depicting the usual sexual differentiation between male and female acanthocephalans. The only exception is the size of lemnisci that appeared to be slightly longer in the male specimen compared to the females. Sample size of 1 may be a factor. In addition to the size and shape of characters in common with the females (Table 1), the size, shape, position of the anterior testis and its distance from anterior trunk, and the fact that testes are not contiguous are important diagnostic features of males of I. sanghae. The generic diagnosis is herein emended to reflect this additional information of the male anatomy (Fig. 2).

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Addendum to the Diagnosis of Intraproboscis

Shared structures of males morphologically similar to and relatively smaller than those of females. Testes large, oblong, not contiguous. Anterior testis distant from anterior end of trunk.