www.nature.com/scientificreports OPEN Species composition of the largest shark fn retail‑market in mainland China Diego Cardeñosa1,2*, Andrew T. Fields1, Elizabeth A. Babcock3, Stanley K. H. Shea4, Kevin A. Feldheim5 & Demian D. Chapman6 Species‑specifc monitoring through large shark fn market surveys has been a valuable data source to estimate global catches and international shark fn trade dynamics. Hong Kong and Guangzhou, mainland China, are the largest shark fn markets and consumption centers in the world. We used molecular identifcation protocols on randomly collected processed fn trimmings (n = 2000) and non‑ parametric species estimators to investigate the species composition of the Guangzhou retail market and compare the species diversity between the Guangzhou and Hong Kong shark fn retail markets. Species diversity was similar between both trade hubs with a small subset of species dominating the composition. The blue shark (Prionace glauca) was the most common species overall followed by the CITES‑listed silky shark (Carcharhinus falciformis), scalloped hammerhead shark (Sphyrna lewini), smooth hammerhead shark (S. zygaena) and shortfn mako shark (Isurus oxyrinchus). Our results support previous indications of high connectivity between the shark fn markets of Hong Kong and mainland China and suggest that systematic studies of other fn trade hubs within Mainland China and stronger law‑enforcement protocols and capacity building are needed. Many shark populations have declined in the last four decades, mainly due to overexploitation to supply the demand for their fns in Asia and meat in many other countries 1–4. Mainland China was historically the world’s second largest importer of shark fns and foremost consumer of shark fn soup, yet very little is known about the species composition of shark fns in this trade hub2. Most global shark catch and trade data are aggregated, unreported, or misidentifed at the species level, hampering species-specifc management and product trace- ability throughout supply chains5,6. Species-specifc monitoring of the shark trade has become a priority for most countries, in part because of international treaty obligations under the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES) where several shark species traded in large volumes have been listed on Appendix II7,8. One key source of species-specifc information on the international trade of shark fns has been the systematic studies of the dried fn market of Hong Kong Special Administrative Region of the People’s Republic of China (hereafer referred to as Hong Kong)1,7,9. Hong Kong is arguably the world’s largest and most consistent importer and re-exporter of shark fns, a small-scale processor (i.e., removing extraneous tissue and preparing fns for the retail market), and a major consumer of shark fn soup 2,10. However, despite its consistency it is unwise to assume that the species composition of Hong Kong is representative of all of the international fn trade because there are other hubs in Asia, each with their own internal dynamics, supply chains, and customer preferences 2. Te fn trade in Mainland China, for example, difers from Hong Kong in at least two major respects: it is also a shark fn producer through its distant water fshing feet2 and Guangdong province in southern China hosts a substantial fn processing industry, where fns landed or imported into China (including many from Hong Kong) are dried, soaked in water, bleached and trimmed of extraneous tissue (e.g., muscle, skin, cartilage) to isolate the ceratotrichia that are the primary soup ingredient 2,11. Shark fns in the city of Guangzhou are obtained from processing plants in Guangdong and then sold to local costumers, and restaurants and wholesalers in Beijing, 1School of Marine and Atmospheric Science, Stony Brook University, Stony Brook, NY 11794, USA. 2Fundación Colombia Azul, Bogotá, Colombia. 3Department of Marine Biology and Ecology, Rosenstiel School of Marine and Atmospheric Science, University of Miami, Miami, FL 33149, USA. 4BLOOM Association, c/o, ADMCF, Suite 2405, Queen’s Road Central, Hong Kong, China. 5Pritzker Laboratory for Molecular Systematics and Evolution, The Field Museum, Chicago, IL 60605, USA. 6Department of Biological Sciences, Florida International University, 3000 NE 151st Street, North Miami, FL 33181, USA. *email: [email protected] SCIENTIFIC REPORTS | (2020) 10:12914 | https://doi.org/10.1038/s41598-020-69555-1 1 Vol.:(0123456789) www.nature.com/scientificreports/ Shanghai and other cities 11. Interview surveys with local traders in Guangzhou suggested that shark fns in this market include tiger sharks (Galeocerdo cuvier), silky sharks (Carcharhinus falciformis), blue sharks (Prionace glauca) and oceanic whitetip sharks (C. longimanus)11, although no species-specifc survey has ever been con- ducted in this market, hampering a direct comparison with other shark fn markets. Te objectives of this study were to (i) investigate for the frst time the species composition of the Guangzhou dried fn market and (ii) compare the species composition of the Guangzhou and neighboring Hong Kong retail markets in terms of species diversity and most commonly traded species. From 2014 onwards, fn market surveys in Hong Kong have used fn-trimmings, an inexpensively sold byproduct of fn processing that is composed of pieces of fn with cartilage that have been cut away from the ceratotrichia, as an afordable market proxy 7,9. Tis same proxy was used in Guangzhou. Methods Guangzhou, the capital city of the Guangdong Province, is the largest shark fn trade hub in mainland China 11 and lies 129 km from Hong Kong. Its retail market is more centralized than Hong Kong’s, comprising a mall that includes mixed wholesale-retail stalls and shops with serial numbers, where shark fns and other highly-priced traditional Chinese medicine products are sold11. Te shark fn retail market of Guangzhou was sampled every 2–3 months from June 2015 to August 2017 for a total of ten sampling events. We generated a list of all vendors based on their serial numbers, and ten random vendors were randomly selected from the complete shop list each sampling event. Sampling events consisted of purchasing two bags of processed shark fn trimmings from each randomly selected vendor, yielding a total of 20 bags of trimmings per sampling, similar to the sampling method described by Fields et al.9 and Cardeñosa et al.7 for Hong Kong. Te contents of each bag were counted, numbered, and ten trimmings were randomly selected for species identifcation. Genomic DNA was extracted following the protocols used by Refs.12,13. Briefy, a small piece of tissue (processed fn trimming) of approximately 2 mm2 was cut and placed in a PCR tube with 200 μl of 10% Chelex Resin (BioRad). Processed fn samples were agitated under water before extraction to reduce potential contamination. Once in Chelex, samples were heated at 60 °C for 20 min and then at 99 °C for 25 min, followed by a brief centrifugation. Each 25 μl PCR included 0.5 μl of extracted DNA, 12.5 μl of GoTaq Hot Start Green Master Mix (Promega) and fve mini-barcoding primers for the cytochrome oxidase I (COI) locus as described by Ref.14. Tese mini-barcoding primers yield up to three fragments for each template (650 bp, 150 bp, 200 bp) but typically only one or both of the smaller fragments due to low quality gDNA present in these processed fns. Te multiplex PCR was amplifed with the following conditions: an initial denaturation at 94 °C for 2 min, followed by 35 cycles at 94 °C for 1 min, 52 °C for 1 min, and 72 °C for 1 min, with a fnal extension of 72 °C for 10 min. Multiplex PCRs were checked on a 3% agarose gel and all products were cleaned using ExoSAP-IT (Afymetrix, Inc., Santa Clara, CA, USA). All products were sequenced twice using the Big Dye Terminator v3.1 cycle sequencing kit (Applied Biosystems, Foster City, CA, USA). Sequencing was performed on an ABI 3730 DNA Analyzer (Applied Biosystems) using the M13 forward primer and the M13 reverse primer. All forward and reverse sequences were checked by eye and priming sites were trimmed using Geneious Pro v. 3.6.1 (https:// www.genei ous.com). Trimmed sequences were compared to BOLD (FISH-BOL) and BLAST (GenBank) data- bases to identify them to the lowest taxonomic category possible (e.g. genus and/or species). Fin trimmings were assigned a species level identifcation when (i) the closest matching subject sequence(s) from only one species exhibited either an exact match to the query or a maximum of 2 bp diferences afer a BLAST search and (ii) when BOLD also returned the same unambiguous species level identifcation. Fin samples that did not fulfll both criteria were only assigned to genus. Tere is also one global species complex (the ‘blacktip sharks’) that includes Carcharhinus limbatus, C. leiodon, C. tilstoni, and C. amblyrhynchoides that exhibit identical sequences for the smaller mini-barcoding fragments; all trimmings that exhibited these sequences were assigned to the blacktip complex. Identical sampling and identifcation protocols have been implemented fortnightly (Febru- ary 2014–January 2015) or monthly (February 2015–present) in Hong Kong 7,9. Terefore, we used trimmings collected and analyzed from the same sampling months in Guangzhou and Hong Kong for comparative species diversity analysis (see below; i.e. we used the same number of trimmings and sampling events per location, even though we had many more from Hong Kong). A rarefaction curve was generated using iNEXT Online15 to estimate the total number of species (i.e., spe- cies diversity) at a given number of samples (i.e., abundance data) and sampling units (i.e., incidence data) for both locations, based on the unifed rarefaction and extrapolation sampling curves of Hill numbers for q = 0, 1 and 216.