Samantha Isenberg (Presenter)
Centers for Disease Control and Prevention
Bio: Samantha Isenberg is a researcher at the Centers for Disease Control and Prevention in Atlanta, GA. Dr. Isenberg joined the Division of Laboratory Sciences in 2014 as a methods development research scientist on the Special Topics Team in the Emergency Response Branch. In this position, Dr. Isenberg developed a method to assess human exposure to soapberry fruit toxins that have been implicated in illness outbreaks worldwide. This work received the 2015 NCEH/ATSDR Honor Award for Excellence in Emergency Response (International) and was nominated for the 2016 Charles C. Shepard Science Award for Laboratory Science.
Authorship: Samantha L. Isenberg (1), Melissa D. Carter (2), Leigh Ann Graham (1), Thomas P. Mathews (1), Mike A. Mojica (1), Darryl Johnson (3), Shelby R. Hayes (3), Aimee A. Sanford (3), Jerry D. Thomas (2), James L. Pirkle (2), Rudolph C. Johnson (2)
(1) Battelle Memorial Institute at the Centers for Disease Control and Prevention, Atlanta, GA; (2) Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA (3) Oak Ridge Institute for Science and Education Fellow at the Cente
| Short Abstract Hypoglycin A (HGA) and methylenecyclopropylglycine (MCPG) are naturally-occurring amino acids found in soapberry (Sapindaceae) fruits. HGA is found in ackee fruit and is known to cause Jamaican Vomiting Sickness, or ackee poisoning. MCPG is a structural analogue of HGA and was recently implicated as the causative agent of seasonal hypoglycemic encephalopathy outbreaks in India. To verify the role of soapberry fruits in these outbreaks, we developed an agricultural method to quantify HGA and MCPG in fruit arils as well as a clinical method to quantify the urinary metabolites of both toxins. |
Long Abstract
Unexplained outbreaks of hypoglycemic encephalopathy have occurred seasonally near litchi-growing regions for the past two decades.(1-5) These outbreaks coincide with the litchi-harvesting season and largely occur in young children. Recently, methylenecyclopropylglycine (MCPG), a naturally-occurring amino acid in litchi fruit, was implicated as the causative agent of the illness. MCPG and its structural analogue hypoglycin A (HGA) are found in the soapberry (Sapindaceae) family. For example, MCPG has been reported in the seeds and arils of litchi fruit, and HGA is found in ackee fruit. HGA is known to be the causative agent of Jamaican Vomiting Sickness as well as Seasonal Pasture Myopathy in horses. In rat studies, both MCPG and HGA were found to be “powerfully hypoglycemic.”(6) As a technical assist for a public health investigation of hypoglycemic encephalopathy outbreaks, we developed clinical and agricultural methods to evaluate exposure to MCPG and HGA.
The clinical method was developed to quantify urinary metabolites of MCPG and HGA. The sample preparation required 10 µL of urine and used an isotope-dilution sample preparation followed by HPLC-MS/MS analysis. We applied the method to a convenience set of 96 individual urine samples obtained commercially with no expected exposure to MCPG or HGA. The method was also applied in a public health investigation to assess exposure to HGA and MCPG and provided the first reported identification of urinary metabolites of HGA and MCPG in human urine.(7) Prior to the development of this clinical method, evidence linking MCPG to hypoglycemic encephalopathy was largely circumstantial due to the lack of analytical methods to evaluate human exposure. Furthermore, the detection of HGA metabolites in suspected hypoglycemic encephalopathy case specimens linked hypoglycin A to hypoglycemic encephalopathy outbreaks in Asia for the first time.
In addition to the clinical method which provides exposure assessment, there was a need for a method to evaluate MCPG and HGA concentrations in litchi and other soapberry fruits. We developed an agricultural method to quantify both MCPG and HGA in the edible arils of soapberry fruit. The sample preparation required 2-3 mg of fruit sample. The analytes were extracted by homogenizing the fruit in 80% ethanol followed by derivatization with dansyl chloride and solid-phase extraction. Quantitative analysis was then carried out by HPLC-MS/MS. We determined the presence of both HGA and MCPG in litchi arils obtained during the public health investigation. We were also the first to report HGA in litchi, longan, and mamoncillo arils.(8)
The popularity of soapberry fruits worldwide and the risk they may pose to certain groups presents a public health challenge. Ingestion of HGA has been reported as the cause of hundreds of deaths worldwide,(9, 10) and MCPG is being implicated in recent illness outbreaks in Asia.(2-4) This work is not only the first to detect metabolites of both MCPG and HGA in human urine but is also the first to provide simultaneous quantification of MCPG and HGA in soapberry arils. Assessing exposure to these toxins as well as evaluating their concentrations in fruit is essential to the prevention of future illness outbreaks.
References:
1. Tanaka K, Ikeda Y. Hypoglycin and Jamaican Vomiting Sickness. Prog Clin Biol Res. 1990, 321, 167-84.
2. John TJ, Das M. Acute Encephalitis Syndrome in children in Muzaffarpur: Hypothesis of toxic origin. Curr Sci India 2014, 106, 1184-5.
3. Shrivastava A, Srikantiah P, Kumar A, Bhushan G, Goel K, Kumar S, et al. Outbreaks of unexplained neurologic illness – Muzaffarpur, India, 2013-2014. MMWR. 2015, 64, 49-53.
4. Pulla P. A child-killing toxin emerges from shadows: Scientists link mystery deaths in India to consumption of lychees. . Science. 2015, 348, 15-6.
5. Yadav R, editor Outbreak of acute neurologic illness outbreak, Muzaffarpur, Bihar, India, 2014. 64th Annual Epidemic Intelligence Service Conference; 2015; Centers for Disease Control and Prevention: Atlanta, GA.
6. Melde K, Jackson S, Bartlett K, Sherratt SA, Ghisla S. Metabolic consequences of methylenecyclopropylglycine poisoning in rats. Biochem J 1991, 274, 395-400.
7. Isenberg SL, Carter MD, Graham LA, Mathews TP, Johnson D, Thomas JD, et al. Quantification of metabolites for assessing human exposure to soapberry toxins hypoglycin A and methylenecyclopropylglycine. Chem Res Toxicol 2015, 28, 1753-9.
8. Isenberg SL, Carter MD, Hayes SR, Graham LA, Johnson D, Mathews TP, et al. Quantification of Toxins in Soapberry (Sapindaceae) Arils: Hypoglycin A and Methylenecyclopropylglycine. Journal of Agricultural and Food Chemistry. 2016, 64, 5607-13.
9. Gaillard Y, Carlier J, Berscht M, Mazoyer C, Bevalot F, Guitton J, et al. Fatal intoxication due to ackee (Blighia sapida) in Suriname and French Guyana. GC-MS detection and quantification of Hypoglycin-A. Forensic Sci Int. 2011, 206, e103-e7.
10. Joskow R, Belson M, Vesper H, Backer L, Rubin C. Ackee fruit poisoning: An outbreak investigation in Haiti 2000-2001, and review of the literature. . Clin Toxicol. 2006, 44, 267-73.
Acknowledgements:
The authors would like to thank Gary R. Takeoka and Leslie A. Harden of the U.S. Department of Health and Human Services for collaboration on the agricultural method for soapberry fruit. The authors acknowledge Scott Landvatter and Rich Tyburski for their efforts towards the initial custom synthesis that led to the commercial availability of the standards and isotopically-labeled internal standards that were imperative for this work. The authors would also like to thank Chariety Sapp of the CDC’s Incident Response Laboratory (IRL) for dispensing convenience set and public health investigation samples prior to analysis.
Disclaimer:
The findings and conclusions in this study are those of the authors and do not necessarily represent the views of the U.S. Department of Health and Human Services, the U.S. Centers for Disease Control and Prevention, or the U.S. Department of Agriculture. The use of trade names and commercial sources is for identification only and does not constitute endorsement by the U.S. Department of Health and Human Services, the U.S. Centers for Disease Control and Prevention, or the U.S. Department
References & Acknowledgements:
Acknowledgements:
The authors would like to thank Gary R. Takeoka and Leslie A. Harden of the U.S. Department of Health and Human Services for collaboration on the agricultural method for soapberry fruit. The authors acknowledge Scott Landvatter and Rich Tyburski for their efforts towards the initial custom synthesis that led to the commercial availability of the standards and isotopically-labeled internal standards that were imperative for this work. The authors would also like to thank Chariety Sapp of the CDC’s Incident Response Laboratory (IRL) for dispensing convenience set and public health investigation samples prior to analysis.
Disclaimer:
The findings and conclusions in this study are those of the authors and do not necessarily represent the views of the U.S. Department of Health and Human Services, the U.S. Centers for Disease Control and Prevention, or the U.S. Department of Agriculture. The use of trade names and commercial sources is for identification only and does not constitute endorsement by the U.S. Department of Health and Human Services, the U.S. Centers for Disease Control and Prevention, or the U.S. Department of Agriculture.
References:
1. Tanaka K, Ikeda Y. Hypoglycin and Jamaican Vomiting Sickness. Prog Clin Biol Res. 1990, 321, 167-84.
2. John TJ, Das M. Acute Encephalitis Syndrome in children in Muzaffarpur: Hypothesis of toxic origin. Curr Sci India 2014, 106, 1184-5.
3. Shrivastava A, Srikantiah P, Kumar A, Bhushan G, Goel K, Kumar S, et al. Outbreaks of unexplained neurologic illness – Muzaffarpur, India, 2013-2014. MMWR. 2015, 64, 49-53.
4. Pulla P. A child-killing toxin emerges from shadows: Scientists link mystery deaths in India to consumption of lychees. . Science. 2015, 348, 15-6.
5. Yadav R, editor Outbreak of acute neurologic illness outbreak, Muzaffarpur, Bihar, India, 2014. 64th Annual Epidemic Intelligence Service Conference; 2015; Centers for Disease Control and Prevention: Atlanta, GA.
6. Melde K, Jackson S, Bartlett K, Sherratt SA, Ghisla S. Metabolic consequences of methylenecyclopropylglycine poisoning in rats. Biochem J 1991, 274, 395-400.
7. Isenberg SL, Carter MD, Graham LA, Mathews TP, Johnson D, Thomas JD, et al. Quantification of metabolites for assessing human exposure to soapberry toxins hypoglycin A and methylenecyclopropylglycine. Chem Res Toxicol 2015, 28, 1753-9.
8. Isenberg SL, Carter MD, Hayes SR, Graham LA, Johnson D, Mathews TP, et al. Quantification of Toxins in Soapberry (Sapindaceae) Arils: Hypoglycin A and Methylenecyclopropylglycine. Journal of Agricultural and Food Chemistry. 2016, 64, 5607-13.
9. Gaillard Y, Carlier J, Berscht M, Mazoyer C, Bevalot F, Guitton J, et al. Fatal intoxication due to ackee (Blighia sapida) in Suriname and French Guyana. GC-MS detection and quantification of Hypoglycin-A. Forensic Sci Int. 2011, 206, e103-e7.
10. Joskow R, Belson M, Vesper H, Backer L, Rubin C. Ackee fruit poisoning: An outbreak investigation in Haiti 2000-2001, and review of the literature. . Clin Toxicol. 2006, 44, 267-73.
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