Data from: Alternative mating tactics in brown widow spiders: mating with or without male self-sacrifice does not affect the copulatory mechanisms

Scans obtained by in-copula freezing of mating males and females of the spider Latrodectus geometricus (for detailed methods on scan aquisition see below). The scans were used to inspect and compare copulatory mechanism of normal mating (adult male with adult female) and immature mating (adult male with subadult female).

Adult and late subadult females of Latrodectus geometricus were transferred to clean experimental boxes (10cm x 10 cm x 6 cm). After two days, males were introduced into the box and the pairs were observed under a dissecting microscope. When copulation occurred, we fixed the pair in copula by pouring liquid nitrogen (-196°C) over them. The frozen couples were transferred to cold 80% ethanol and stored at -40°C for three weeks to ensure stable fixation. We successfully froze and inspected 17 couples during adult mating (17 in somersault position and 6 prior to the somersault) and 11 couples during immature mating.

The samples were dehydrated in a graded ethanol series (three times in 80%, 90%, 96%, and 99% ethanol for 1 hr each) and contrasted for 48 hours using a 1% iodine solution (in pure ethanol). For scanning, the samples were critical point dried with a BAL-TEC CPD 030 and mounted on insect pins using super glue. Scans were performed in an Xradia Micro XCT-200 (Carl Zeiss Microscopy GmbH) using the macro lens (4x) for overviews of copulating pairs (scan parameters: 40 kV, 200 μA, exposure time 0.75 - 1.5, pixel size 3.98 - 5.56 µm) and the 10x lens for close ups (scan parameters: 40 kV, 200 μA, exposure time 3.0 - 4.5 s/frame, pixel size 2.21-2.34 µm) on the interlocked copulatory organs. The male and female copulatory organs in resting position were scanned using a 20x lens (scan parameters: 40 kV, 200 μA, exposure time 8-9 s/frame, pixel size 1.01 - 1.13 µm). Image stacks were created using XMReconstructor software (Carl Zeiss Microscopy GmbH). Data were visualized and processed using the 3D analysis software AMIRA 5.6 (Visualization Science Group, FEI).