21.12.2022

The preparation of spermathecae with the help of peptidases

The spermathecae of spiders consist of chitin and often have a different shape from species to species and therefore play an essential role in the taxonomic determination of spiders. In spiders, they are located ventrally/basally in the opisthosoma, between the anterior booklung behind a trapezoidal plate consisting of slightly thickened chitin, the so-called genital plate.

The problem with spermathecae that are to be prepared from a spider preserved in alcohol is the fact that the spermathecae have fleshy components attached to them that are not easily detached. This can happen, for example, through years of improper storage of the spider in incorrectly diluted alcohol. If the alcohol content is too high or the spider has been dry for a long time, this can lead to the tissue around the spermatheca drying out and sticking to it. If one now tries to release the spermatheca mechanically, e.g. by scraping off the fleshy parts with dissecting needles, considerable damage to the spermatheca can occur very quickly. In 2006, we described a quick and effective way of safely preparing a spermatheca from a spider preserved in alcohol using maceration with heated lactic acid (von Wirth 2006). We had also briefly explained this method on our WEBSITE.

However, other methods for the preparation of spermathecae from spiders preserved in alcohol are also described in spider systematics. Often this is done with the help of a digestive enzyme, a so-called peptidase. In the following we would like to present this method, because the preparation of spermathecae with the help of maceration with lactic acid is not always welcome, because it produces slightly irritant vapours and, especially in modern natural history museums which are equipped with smoke detectors, the curators are often afraid that the smoke detectors will go off because light vapours are produced when the lactic acid is heated. The preparation of the spermatheca with the help of a peptidase takes longer, but takes place without heating and vapour development. In tarantula taxonomy, especially among Chinese spider taxonomists, the peptidase trypsin is often used ( see Lin et al. 2022a; Lin et al. 2022b). However, we have used the peptidase pepsin for our following description, as this digestive enzyme is somewhat easier to obtain here.

Pepsin is primarily used in entomology and arachnology to soften dried arthropods so that they can be better prepared and fixed. In entomology shops, you can often get a ready-made pepsin solution consisting of 5 grams of pepsin powder, 15 ml hydrochloric acid (HCI) and 250 ml distilled water (Kless 1986). This is also suitable for the preparation of the spermatheca. The pepsin solution should be stored permanently in a cool place. Do not reuse used pepsin solution, but dispose of it.
Proceed as follows for the preparation:

After the genital plate with the spermatheca and the fleshy parts adhering to it has been prepared from the preserved spider (see von Wirth 2006 or HERE) (Pic 1), it is placed in a block bowl, the pepsin solution is removed with a pipette (Pic 2) and the block bowl is now filled with the pepsin solution (Pic 3). Finally, the block bowl with the pepsin solution and the genital preparation is covered with a block bowl cover plate made of glass (Pic 4) and stored at room temperature or in an incubator at 37 degrees. Now the pepsin solution "digests" the organic-meaty components that adhere to the spermatheca. After 24 hours the spermatheca is almost free (Pic 5), but the milky substrate shows that the "digestion process" is not yet complete. After 48 hours, however, the main part of the fleshy components has been removed (Pic 6) and the spermatheca can now be dissected out of the genital plate, photographed (Pic 7) or drawn and then added to the preserved spider in a genital tube as described by von Wirth 2006 and HERE.

Resumè:

The preparation of spermathecae from a spider preserved in alcohol using maceration with heated lactic acid is much faster than the method of preparation with pepsidase pepsin presented here, but there are situations in which this type of spermathecae preparation should not be used or cannot be used for safety reasons. In these cases, the presented preparation with the help of a pepsidase is suitable for freeing the spermatheca of a spider from disturbing fleshy components.

Reference:

KLESS, J. (1986). Ein neues Verfahren zum Aufweichen unpräparierter Käfer. Entomologische Blätter 82 (Kleine Mitteilungen Nr. 2051): 120-121.

LIN, Y. J., YAN, X. Y. & LI, S. Q. (2022a). Two new species of the genus Chilobrachys (Araneae, Theraphosidae) from China. ZooKeys 1081: 99-109

LIN, Y. J., YAN, X. Y. & LI, S. Q. (2022b). A new species of the tarantula genus Haplocosmia (Araneae, Theraphosidae) from Tibet, China. Biodiversity Data Journal 10(e82682): 1-6.

VON WIRTH, V. (2006): Präparationstechniken von (Vogel-)Spinnen-Spermatheken. ARACHNE 11 (6), pp. 16 – 28