Spider Inventories General Data

Estación de Biología Chamela, IB-UNAM
Estación de Biología los Tuxtlas, IB-UNAM
Parque Nacional Volcán Pico de Orizaba
Parque Eco-Turístico Las Pozas, Xilitla

Measuring Spider Diversity

Collecting every spider species in a large area is impossible, so richness estimators and other statistical tools are needed to measure community parameters. Standardized protocols are essential for comparing inventories among localities. Most arachnologists have used these protocols with minor variations.

The main objective is to make direct collecting effort explicit, usually within one-hectare plots. Following these two rules makes inventories easier to compare and more useful for future studies.

1. Make collecting effort explicit. For Araneomorphae, 1 hour/person/method is the standard unit.
2. Use areas equal to or larger than 2,500 m² (50 × 50 m plots). This allows species/area extrapolations with other studies. Transects are NOT recommended because they cover much less area and prevent meaningful extrapolations.

Tracing a Plot with Simple Tools

A sampling plot can be traced with a compass, a long measuring tape, and two people. Choose an orientation, for example North–South, and project a straight line toward a visible landmark while the second person extends the tape. Add 90° turns at the corners until the plot is closed. Irregular polygons can be measured with simple geometry, a ruler, and field notes. If done correctly, you should end near the starting point.

Microscopy Methods

This section summarizes methods to obtain high-quality digital images with relatively inexpensive equipment. Most techniques were shared by other arachnologists; some were developed in our laboratory. Improvements and suggestions are welcome.

Regular optical microscopes with 10X and 20X PLAN-APO objectives produce sharper images than dissecting microscopes at the same magnification, but they have very short focal length and almost no working distance. Dissecting microscopes improve working distance and focus using many lenses, although resolution is reduced because light passes through more glass. High-quality optics optimize this balance but are usually expensive.

The shallow focal length of optical microscopes is solved by combining images taken at different focus levels. We have used Helicon Focus for many years and highly recommend it. Most combined images require little or no editing, which is important for projects involving thousands of photographs. We usually combine about 35 manually focused images per final picture; automated systems can obtain similar or better results faster.

Preparation Setup

The Petri dish with sand. This traditional method is useful when hundreds of specimens must be sorted and photographed. Partially cover the specimen with sand until the desired position is stable, then remove alcohol until the surface is close to the specimen. Alcohol-based gels are not recommended because they are difficult to remove, produce bubbles, and create more optical distortion than liquids. If necessary, sand can be removed from the image with ImageMagick or AI software.

Technique for 100X. This setup uses the small dish shown at the bottom right of the image, glued over a glass slide. It is useful for non-cleared structures or specimens smaller than 1 mm. A plastic lid or small well also works if it is wide enough for the objective lens and tall enough to contain alcohol. Our plastic lid has an internal diameter of 1.53 cm, an external diameter of 1.66 cm, and a height of 0.9 cm. Illumination is provided with an LED lamp.

Mounting the specimen. Under a dissecting microscope, place a drop of petroleum jelly at the bottom of the plastic lid. The surface must be dry or the jelly will not stick. Fill the dish with alcohol, shape the jelly to hold the specimen, and attach it. Transfer the preparation to the optical microscope. Petroleum jelly is removed from the specimen by placing it for 15 seconds in a vial with chloroform or xylene.

Technique for 200X. Repeat the 100X procedure but use an excavated glass slide (1 or 2 mm deep). Cut the excavated slide to fit a small Petri dish, fill the dish with alcohol, and introduce the slide. Seal the cover with petroleum jelly so it can hold alcohol for several minutes before bubbles appear. All procedures are done under alcohol inside the Petri dish. Illumination is provided from above with an LED lamp. For background, place white or black paper under the preparation.

Technique for cleared structures. This uses the temporary slide shown at the top right, with either clove oil or methyl salicylate. Jonathan Coddington described this preparation in a difficult-to-find book chapter, so a brief description is included here. Paste several coverslips on a microscope slide; the stack must be slightly taller than the structure to be observed. Cut a narrow glass strip from a long coverslip and attach it with petroleum jelly to the top of the stack. Add a drop of clove oil or methyl salicylate to join the glass pieces and cover the specimen.

Important. Never mix methyl salicylate with water because it will damage the specimen. First transfer the structure to 96% alcohol for a few minutes, then transfer it to the clearing agent. Under a dissecting microscope, transfer the specimen with a pipette and place it beneath the tallest part of the narrow glass strip. Adjust the structure until the desired view is stable, then move the preparation to the optical microscope. Minor adjustments can be made by gently tapping the coverslip, but major changes should always be done under the dissecting microscope. Check that the glass strip protects the objective from contact with the clove oil.

Image Composition Rules

These rules keep image composition constant within and among inventories.

• Remove all legs and the pedipalp from the left side of the specimen. Dissect cephalothorax appendages at the membrane between the coxa and trochanter, and remove cuticle or muscle fragments that may spoil the image. Specimen orientation is flexible but should remain consistent.
• Maximize information content with few high-resolution images instead of several small images of the same structure.
• Do not crop images; keep the original width and height from the camera. If the structure looks too small, use an instrument with higher magnification.
• Sixteen standard views are enough to document the anatomy of a species with both sexes and provide data for reliable identifications: eight images for overall anatomy (habitus and prosoma), four for epigyna (two attached and two dissected/cleared), and four for the male left pedipalp. We recommend an upper limit of 20 images per species.

Workstations

All images were taken with Nikon equipment. The first workstation uses two digital cameras: a DS-Fi1 connected to an SMZ1270 dissecting microscope and a DS-Fi2 connected to an E200 optical microscope, both controlled with a Dell Inspiron 660s. The second workstation has SMZ1000 and E200 microscopes sharing a DS-Fi3 connected to an HP Slimline 270-a0xx. Illumination is provided with custom lamps made of two 1.5-watt LEDs mounted on flexible tubes with aluminum heads to diffuse heat.