Vernal pool habitat has been severely reduced by urban development in southern California. This fenced pool near Mira Mesa Market Center (San Diego County, CA), is the last remaining of an original complex that contained 67 vernal pools.
(Photograph by A.G. Vandergast, USGS)

Fairy shrimp are small crustaceans that inhabit seasonal wetlands such as vernal pools, desert playas and wet meadows. Fairy shrimp are adapted to living in temporary wetlands, and many species can complete the aquatic portion of their life cycles in a matter of days or weeks. During the dry season, embryos encapsulated in hard cysts lay dormant in pool sediments for months or even years. Of the approximately 50 species known in the United States, five that occur in California are federally listed as endangered or threatened, mainly due to loss of habitat. This is especially true in southern California, where over 98% of historical vernal pool habitat has been lost to urban development.

In southern California, the endangered San Diego fairy shrimp (Branchinecta sandiegonensis) and the threatened vernal pool fairy shrimp (Branchinecta lynchi) overlap in distribution with the more common versatile fairy shrimp (Branchinecta lindahli). These three closely related species can only be morphologically distinguished as adults. However, adults are only present when pools have been filled for a sufficient length of time, which, in drought-prone southern California, can happen rarely or in some years not at all.

Developing a method to distinguish Branchinecta fairy shrimp species directly from cysts provides two distinct advantages. First, it allows listed fairy shrimp to be detected over a much wider time frame. Second, sampling cysts from vernal pools during the dormant dry season is less destructive to other co-occurring species, particularly the specialized flora that characterize these sensitive habitats. Cyst identification may allow listed species to be detected on project sites earlier in the planning process and cysts may be a better target for surveys and long-term monitoring at protected and restored sites.

We have developed a genetic assay based on the polymerase chain reaction (PCR) to isolate DNA from fairy shrimp cysts and to screen this DNA to distinguish among species. Based on fixed differences in the DNA sequences of the mitochondrial Cytochrome Oxidase I gene, species-specific primers were designed to amplify fragments of differing length in each of the three Branchinecta species. These primers were combined into a “multiplex PCR” allowing cyst DNA to be identified to one of the target species in a single reaction. This multiplex PCR assay provides a fast and accurate method of identifying Branchinecta cysts from southern California vernal pools. While this assay was strictly designed to distinguish among the three Branchinecta species present in southern California, with the inclusion of additional species-specific primers, this method could be applied to other regional Branchinecta species assemblages.

More generally, multiplex PCR methods may be an appropriate alternative to morphological species identification when species co-occur with closely related congeners and have few distinguishing physical traits, or in cases where these traits are only apparent in particular life stages. For cryptic endangered species, molecular identification can aid in recovery efforts by allowing the presence or absence of listed species to be determined quickly, so that appropriate management actions can be applied in a timely manner, and by aiding in the development of simple and repeatable protocols for monitoring species over time.

The endangered fairy shrimp, Branchinecta sandiegonensis
(Photograph by D. Parsick, USD).