Environmental Barcoding in the Costa Rican Tropical Dry Forest: Hajibabaei Lab research in the Area de Conservacion Guanacaste.

dscn1376In the latest example of the Hajibabaei Lab’s ongoing work with the Area de Conservacion Guanacaste (ACG), masters’ student Lisa Ledger and  Steven Blewett, a volunteer research assistant have just returned from a one week field study of the tropical dry forest of Sector Santa Rosa.

Tropical dry forest is among the world’s most endangered tropical forest types, but also contains some of its greatest biodiversity. Shifting between an average monthly rainfall of 172mm in the wet season to a mere 29mm in the dry, its unique ecology is one that features cacti thriving alongside liana vines and hosts species adapted to both dry and wet environments: an estimated 23,000 individual species in total.

The parks of Costa Rica’s Area de Conservacion Guanacaste contain over 160,000 hectares of protected forest, in varying states and ages of succession. While all are visually stunning, the question remains unanswered: is the biodiversity of the regenerated tropical dry forest approaching that of primary forest? If so, is it with the same distribution of species, or have anthropogenic impacts permanently altered patterns of biodiversity?

Lisa’s graduate research sets out to answer this through the use of environmental DNA barcoding. In environmental barcoding, a mixed sample such as the entire contents of a Malaise insect trap or a homogenized soil004 sample has its total DNA extracted and then amplified for a series of barcode regions. This is unlike traditional barcoding, where a sample is taken from a single individual, and requires significantly more sequencing power than is cost-effective using traditional Sanger sequencing. Only with the advent of next-generation sequencing systems, such as the Roche 454 FLX and Illumina MiSeq machines used by the Hajibabaei labs, has environmental barcoding become a viable approach for biodiversity assessment.

In consultation with Dr. Dan Janzen, three plots of land in the ACG were chosen for use as study sites: Bosque Humedo, a rare site of primary tropical dry forest, Bosque San Emilio, a site of late secondary succession following its abandonment as a plantation in the 1920s, and an area of grasslands and early-to-mid secondary succession where a yearly firebreak has been established since the late 1970s, mimicking patterns of agricultural land management seen on the borders of the park.

Memo, a parataxonomist who has worked closely with Dr. Janzen in the past, assisted Lisa as both a local guide and with the assembly of the Malaise traps at each site. He also provided a wealth of local knowledge of plant and animal species within the forest, from the life stages of fungi to the local names of species such as the common agouti (‘guatusa’). Memo is also responsible for the final collection of insects from the trap heads, as the two week run of the Malaise traps exceeds the one week duration of the trip.

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In addition to the nine Malaise traps spread across the three study plots, nine soil samples were taken from each as well, with an eye towards expanding the study to belowground as well as aboveground biodiversity.  Unlike the Malaise traps, the soil sampling was a single day operation, with the following three days spent conducting DNA extraction while still within the park.   Despite the use of a commercial soil DNA extraction kit, many challenges were still involved in conducting molecular biology work while under field conditions.   Thanks to the gracious help and loans of equipment from the researchers of the ACG, particularly Luis Felipe Chavarria, the provision of a workspace within the BioLep research station and nearly an entire bottle of ELIMINase (TM), Lisa and Steven were able to meet those challenges and extract the soil DNA despite the tropical heat and the threat of stray insects attempting to become part of the extraction process.

Although the collection and processing of samples was the main focus of the trip, the stay in the ACG also afforded opportunities for hiking and exploration and a memorable education on both the history of the park and a tour of local wildlife courtesy of Dr. Winnie Hallwachs.  The physical remove between the lab and the field can often lead to a mental remove between the research and the reality of the world it’s drawn from, but being introduced to  the work of Dan and Winnie in the cataloguing and barcodign of tropical lepidoptera and their associated plants, witnessing local school science programs introducing children to the protected spaces and species of the ACG and the simple experience of moving among the flora and fauna of a tropical dry forest makes an indelible point of what a rare and precious resource the tropical dry forest is.