Equipment and  
Materials  

Field and Laboratory  
Research 

Other Research Questions

Students work in groups of four to five throughout the four to six laboratory periods that this exercise is taught.  A typical module involving four laboratory periods is briefly described below, session by session.  This module is self-sustaining, so each session begins with a presentation by the instructor, giving students basic background on relevant terms, concepts, and/or methods.  As the module progresses, students learn about the basic ecology of freshwater springs, methods of sampling and environmental measurement, research design and statistical analysis, and ways of presenting results in the form of a scientific paper suitable for hypothetical submission to the journal Ecology.  In this way, several important facets of the research process are introduced to students. 

Quizzes based on assigned readings (from Halliday, 1980; Ward, 1988; and Wiley et al., 1991) are also given at the start of each session to encourage students to come prepared. 

First Session:   

Students travel to a nearby spring where they: 

• Take several environmental measurements:

Physical: temperature, depth, width, flow rate, discharge, substrate types 
Chemical: pH, conductivity, dissolved O₂ 
Biotic: types and coverage of macrophytes (aquatic plants) and springside vegetation 
These measurements are used in the study-site section of the students’ research report (see also below). 
 
• Collect representative specimens of the fauna for later identification and phylogenetic analysis.  Animals are collected by dipnets and sieves, and by picking them by hand from the substrate and macrophytes.

• Use dipnets to randomly collect 50 amplexed pairs, 50 single mature males, 50 single mature females, and 50 brooding females (which may double-count with females in the previous categories) of the amphipod Gammarus minus.  Males have larger gnathopods (forelegs) than females, and only mature females have oostegites (marsupial plates) on the inner surface of the coxal plates of some of their legs (usually 2nd to 5th).  Brooding females can be easily spotted because the young in their marsupium (brood pouch) often appear as a dark ventral patch.

(Note: the above tasks can be divided up among the assigned student groups so that each group collects only 10-15 pairs/individuals of each category).

The students continue to identify the specimens that they have collected, at least to genus or family level.  They also identify at least four characters to be used in their assessment of the evolutionary relationships of at least five taxa.  Each character is assigned a set of two or more states (e.g., presence vs absence, large vs. small, number of legs or other appendages, etc.). 

During this session, students also test their four hypotheses concerning sexual selection in the amphipod G. minus.  Body size is estimated by body length, measured from the base of the antenna to the tip of the tail (uropod) to the nearest millimeter using a metric ruler.  To do this, each animal must be anesthetized in carbonated water and then fully stretched, as they are normally “hump-shaped”.  The stretching is best done with micro-pins using the edge of a metric ruler for support.  (Note: it is essential that the test amphipods have been randomly chosen.  Otherwise the students will select the larger, more conspicuous amphipods for study, which would invalidate the intended statistical tests).  To complete their tests, the students must also remove the eggs/embryos from the brood pouches of 50 anesthetized females.  This is done by pinning each female to a wax sheet, and then using a pipette of spring water to carefully flush the eggs/embryos from the brood pouch. 
 
Third Session: 

In this session, the students use the computer program PAUP 4.0 to estimate the evolutionary relatedness of their chosen taxa.  The students are asked to turn in a brief written report on their results by the end of the laboratory period.  The report should include: 

• The input data matrix, including names of taxa and characters

• Pairwise distances between the taxa 

• One or more possible evolutionary trees, including a consensus tree if necessary 

• A proposed classification 

where i is selection intensity, X_a is the mean body length of amplexing males, X_t  is the mean body length of both amplexing and non-amplexing males, and s_t is the standard deviation of X_t (see Ward, 1988). 

Fourth Session: 

During this session, the students learn how to present the results of their sexual selection research in the form of graphs or tables.  Each group also constructs an outline of their sexual selection research report.  This outline includes a study-site description based on the environmental measurements taken during the first laboratory session.  The final report is due a week after the last laboratory session.  The students are asked to follow the “Instructions to Authors” of the journal Ecology.  They are also given extensive background information on writing scientific papers (e.g., Ambrose & Ambrose, 1987). 
 
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