Abstract
Two different approaches, real-time PCR and in silico analysis, were used to study the abundance of bacterial genes encoding the enzymes producing biomarker metabolites in metagenomes of gut microbiota of children with autism spectrum disorders (ASD) and healthy children. As a result of the analysis of the published data, the following biomarker metabolites of ASD were chosen for the research: p-cresol, indole, propionic acid, and D-lactic acid. For the real-time PCR and the transcriptomic analysis, primer sequences were used specifically developed for conservative regions of the genes involved in production of these bacterial metabolites. Nucleotide sequences of the conservative regions were also united into a catalog and were searched in metagenomic assemblies as part of in silico analysis. Comparison of the results revealed a decrease in abundance of the genes in the metagenomes of children with ASD during both PCR and bioinformatics analyses. In addition, real-time PCR made it possible to detect a significant increase in abundance of the genes encoding p-hydroxyphenylacetate decarboxylase and D-lactate dehydrogenase in Bacteroides fragilis and Alistipes finegoldii species and methylmalonyl-CoA decarboxylase in B. fragilis, Alistipes shahii, and Eubacterium rectali species. A low level of the transcripts in total RNA was found in microbiota of a child with severe form of ASD.
Similar content being viewed by others
REFERENCES
Averina, O.V. and Danilenko, V.N., The human gut microbiota: role in the formation and functioning of nervous system, Microbiology, 2017, vol. 86, no. 1, pp. 1—19. https://doi.org/10.1134/S0026261717010040
Dinan, T.G. and Cryan, J.F., Gut—brain axis in 2016: drain—gut—microbiota axis—mood, metabolism and behavior, Nat. Rev. Gastroenterol. Hepatol., 2017, vol. 14, no. 2, pp. 69—70. https://doi.org/10.1038/nrgastro.2016.200
Borre, Y.E., Moloney, R.D., Clarke, G., et al., The impact of microbiota on brain and behavior: mechanisms and therapeutic potential, Microbial Endocrinology: The Microbiota—Gut—Brain Axis in Health and Disease, New York: Springer-Verlag, 2014, pp. 373—403.
Poletaev, A.B. and Shenderov, B.A., Autism and autoimmunity: genetics or epigenetics, Clin. Pathophysiol., 2016, vol. 4, pp. 1—14.
Poletaev, A.B. and Shenderov, B.A., Autism: genetics or epigenetics?, ARC J. Immun. Vaccines, 2016, vol. 1, no. 2, pp. 1—7.
Kovtun, A.S., Averina, O.V., Alekseeva, M.G., and Danilenko, V.N., Antibiotic resistance genes in the gut microbiota of children with autistic spectrum disorder as possible predictors of the disease, Microbial Drug Resist., 2020 (in press).
Hughes, H.K., Rose, D., and Ashwood, P., The gut microbiota and dysbiosis in autism spectrum disorders, Curr. Neurol. Neurosci. Rep., 2018, vol. 18, no. 81, pp. 1—15. https://doi.org/10.1007/s11910-018-0887-6
Kang, D.W., Ilhan, Z.E., Isern, N.G., et al., Differences in fecal microbial metabolites and microbiota of children with autism spectrum disorders, Anaerobe, 2018, vol. 49, pp. 121—131. https://doi.org/10.1016/j.anaerobe.2017.12.007
Wang, L., Christophersen, C.T., Sorich, M.J., et al., Elevated fecal short chain fatty acid and ammonia concentrations in children with autism spectrum disorder, Diges. Dis. Sci., 2012, vol. 54, no. 1, pp. 1—7. https://doi.org/10.1007/s10620-012-2167-7
Macfabe, D.F., Short-chain fatty acid fermentation products of the gut microbiome: implications in autism spectrum disorders, Microb. Ecol. Health. Dis., 2012, vol. 23, pp. 1—25. https://doi.org/10.3402/mehd.v23i0.19260
Reigstad, C.S., Salmonson, C.E., Rainey, J.F., et al., Gut microbes promote colonic serotonin production through an effect of short-chain fatty acids on enterochromaffin cells, FASEB J., 2015, vol. 29, no. 4, pp. 1395—1403. https://doi.org/10.1096/fj.14-259598
MacFabe, D.F., Cain, N.E., Boon, F., et al., Effects of the enteric bacterial metabolic product propionic acid on object-directed behavior, social behavior, cognition, and neuroinflammation in adolescent rats: relevance to autism spectrum disorder, Behav. Brain. Res., 2011, vol. 217, pp. 47—54. https://doi.org/10.1016/j.bbr.2010.10.005
De Angelis, M., Piccolo, M., Vannini, L., et al., Fecal microbiota and metabolome of children with autism and pervasive developmental disorder not otherwise specified, PLoS One, 2013, vol. 8, no. 10, p. e76993. https://doi.org/10.1371/journal.pone. 0076993
Persico, A.M. and Napolioni, V., Urinary p-cresol in autism spectrum disorder, Neurotoxicol. Teratol., 2013, vol. 36, pp. 82—90. https://doi.org/10.1016/j.ntt.2012.09.002
Mack, D.R., D(–)-lactic acid-producing probiotics, D(–)-lactic acidosis and infants, Can. J. Gastroenterol., 2004, vol. 18, no. 11, pp. 671—675. https://doi.org/10.1155/2004/342583
Kovtun, A.S., Alekseeva, M.G., Averina, O.V., and Danilenko, V.N., Identification of aminoglycoside phosphotransferases of clinical bacterial strains in the microbiota of Russia residents, Vestn. Ross. Gos. Med. Univ., 2017, vol. 2, pp. 14—19. https://doi.org/10.24075/brsmu.2017-02-02
Srikantha, P. and Mohajeri, M.H., The possible role of the microbiota—gut—brain—axis in autism spectrum disorder, Int. J. Mol. Sci., 2019, vol. 20, no. 9, p. E2115. https://doi.org/10.3390/ijms20092115
Funding
This study was supported by the Russian Science Foundation (project 17-15-01488).
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest. The authors declare that they have no conflict of interest.
Statement of compliance with standards of research involving humans as subjects. All procedures carried out in a study with the participation of people comply with the ethical standards of the institutional and/or national research ethics committee and the 1964 Helsinki Declaration and its subsequent changes or comparable standards of ethics.
Informed voluntary consent was obtained from each of the participants in the study.
Additional information
Translated by A. Kashevarova
Rights and permissions
About this article
Cite this article
Averina, O.V., Kovtun, A.S. & Danilenko, V.N. Bacterial Genes of Metabolite Biomarkers of Autism Spectrum Disorders in Gut Microbiota of Young Children: Detection by Real-Time PCR and In Silico Analysis. Russ J Genet 56, 1260–1268 (2020). https://doi.org/10.1134/S1022795420100026
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1134/S1022795420100026