Freshwater Spring


Freshwater Springs as Model Systems 

Further Background 

Equipment and Materials


Classifying the Diversity of Life  
A basic description of any ecological system depends on the proper identification and classification of the organisms living there.  Therefore, an important component of this exercise is to give students an appreciation of how biologists identify, classify and assess the evolutionary relationships of various kinds of organisms.  The fauna of springs, including various kinds of invertebrates, salamanders and fish, are abundant, easily collected, and possess many readily identifiable, distinguishing characteristics, and thus are well suited for taxonomic and phylogenetic analyses.  Students are asked to imagine that they are the first biologists to have studied the spring(s) visited (in many cases this is actually true!), and their job is to make a biotic inventory, including a list of all of the taxa found using taxonomic keys available in the laboratory (see References page).  In addition, they are asked to determine the evolutionary relationships of five or more of these taxa using characters that they themselves have chosen.  The taxon-character matrix that the students construct is then used to estimate the most parsimonious evolutionary tree(s) for the taxa in hand, by means of the computer program, Phylogenetic Analysis Using Parsimony (or PAUP; Swofford, 1991, 1998).  Although quite sophisticated, PAUP is user-friendly.  Background on the basic concepts and terminology of phylogenetic analysis is provided by Wiley et al. (1991). 

A major lesson that students learn is that biological classifications and evolutionary trees are hypotheses.  They are only as good as the data used to construct them.  They also learn that, as stated by the well-known Harvard biologist, E.O. Wilson (1992), “Systematics is mostly science but also a bit of art.” 

Amphipods are small crustaceans (usually 5-20 mm in body length) common in a variety of marine and freshwater habitats.  They are sometimes called “scuds” or “side-swimmers” because of their habit of rolling over on their sides or backs when they swim.  Laymen call amphipods “shrimp”, but they are only distantly related to true shrimp (= decapods).  Most species are scavenging bottom-dwellers often seen crawling over the substrate or vegetation in search of animal or plant food. 

A freshwater Gammarus species 

Over 50 freshwater species are found in North America, most of which are found in subterranean habitats (e.g., caves; Holsinger, 1976).  Gammarus species are most common in relatively cool springs, streams, ponds, and lakes.  Gammarus minus, the species used in the present exercise, is largely restricted to springs, springbrooks, and cave streams throughout much of the Appalachians and west to Illinois, Missouri, and Arkansas.  Cave populations are morphologically distinct with degenerate eyes, long antennae, and bluish coloration (Culver, Kane & Fong, 1995). 

G. minus only occurs in relatively alkaline hardwater (usually pH > 6.0; Glazier, Horne and Lehman, 1992).  The absence of amphipods from acidic softwater is thought to be related to a deficiency of calcium for carapace formation and to difficulties of maintaining energy and ionic balance (Glazier, 1998).  Often extremely abundant in hardwater springs (~ 600-8,000 m-2 in central Pennsylvania), amphipods are readily collected and can be easily studied and maintained in the laboratory.  In addition, they are sensitive indicators of water chemistry and pollution, and thus are widely used in environmental toxicology studies (Maltby, 1994; Plenet, 1995). 

Sexual Selection  

Animals compete not only for resources (e.g., food and shelter), but also for mates.  Those individuals who do the best in this “competitive game”, produce the most offspring and thus their genotypes are favored and proliferate relative to less successful genotypes.  This is natural selection.  A special form of natural selection is sexual selection, which results from competition for mates.  Sexual selection can be simply defined as “differential mating success” (Halliday, 1980).  Those inherited characteristics that increase mating success (e.g., larger body size, stronger weapons, and more elaborate ornamentation in males) will be favored.  This is because those animals that mate the most will have the most offspring and thus their genetic characteristics will be preferentially passed onto future generations. 

Apparently because of intense competition for mates, precopulatory mate guarding has evolved in many crustaceans, including amphipods (Conlan, 1991; Jormalainen, 1998).  The male grasps a female before she is ready to lay her eggs, and carries her beneath him (= precopula or amplexus) for a few days until she molts.  Immediately after molting the male fertilizes the female’s eggs, which are deposited in her ventral brood pouch (marsupium).  Embryonic development takes several weeks (~ 35-40 days in G. minus).  Around the time that the young leave the brood pouch, the female becomes sexually attractive to males again and is soon amplexed. 

Amplexing pair of Gammarus pulex 
(picture taken by David Hollingworth, Department of Animal and Plant Sciences, University of Sheffield, England)

In aquatic amphipods, including G. minus, males are larger than females.  Because other animals with precopula (e.g., isopods) also have larger males than females (contrary to the more general trend in animals of females being larger than males), it has been suggested that this sexual dimorphism is the result of sexual selection.  Experimental studies of G. pulex have shown that males compete for access to females, and that larger males “win” (i.e., mate with females) more often than smaller males (e.g., Elwood, Gibson & Neil, 1987; Ward, 1988).  Since body size is inherited (i.e., passed onto offspring), this male-male competition inevitably leads to the evolution of larger males, as is found in nature.  Also larger males have an advantage over smaller males in carrying females, especially in fast water currents (Naylor & Adams, 1987).  However, other factors such as predation and structural constraints have probably prevented a never-ending escalation of male body size.  In particular, larger males are more conspicuous to visual predators such as fish. 

Sexual dimorphism in Gammarus minus
(male on top, female in middle, and juvenile at bottom)

In this exercise, students typically test the following four hypotheses: 

  • If larger size is advantageous for mating as discussed above, adult males in precopula should be larger, on average, than adult males not in precopula.
  • Since sexual selection is expected to be acting on males only, there should be no difference in body size between amplexed females vs. non-amplexed females.
  • In most animals larger females produce more offspring than smaller females.  This should be true in G. minus.
  • If the above hypothesis is true, mating should be size-assortative, i.e., larger males should prefer to mate with larger females.
A nice feature of this amphipod-spring system is that the above hypotheses can be tested during any season, because amplexus occurs year-round in spring-dwelling populations of G. minus

Some ideas for the sexual selection exercise were taken from a laboratory practical designed by Dr. T.R. Birkhead in the Department of Animal and Plant Sciences at the University of Sheffield, England. 

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©March 1999