Skip to main content
SpringerLink
Log in

Cystic Fibrosis: A Simple and Customized Strategy for Genetic Screening Able to Detect Over 90% of Identified Mutated Alleles in Brazilian Newborns

  • Original Research Article
  • Published:
Molecular Diagnosis & Therapy Aims and scope Submit manuscript

Abstract

Introduction

The incorporation of molecular genetic testing into cystic fibrosis (CF) screening programs increases the specificity of the diagnostic strategy and has the potential to decrease the rate of false- positive results. In this sense, our objective was to develop a genotyping assay that could detect 25 pathogenic variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene with high sensitivity and that could be incorporated into the routine of newborn screening, complementing the current existing protocol used in our public health institution.

Methods

A mini-sequencing assay was standardized using single-base extension in a previously genotyped control sample. This strategy was validated in a Brazilian cohort of CF patients by Sanger sequencing.

Results

The inclusion of the 25 variants in the current newborn screening program increased the identification rates of two alleles from 33 to 52.43% in CF patients. This new approach was able to detect a total of 37 variants, which represents 93.01% of all mutated alleles described in the last CF Brazilian Register.

Conclusions

Mini-sequencing for the simultaneous detection of 25 CFTR gene variants improves the screening of Brazilian newborns and decreases the number of inconclusive cases. This method uses minimal hands-on time and is suited for rapid screening, which reduces sample processing costs.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  1. O’neal WK, Knowles MR. Cystic fibrosis disease modifiers: complex genetics defines the phenotypic diversity in a monogenic disease. Annu Rev Genom Hum Genet. 2018;19:201–22.

    Article  Google Scholar 

  2. Drumm ML, Pope HA, Cliff WH, et al. Correction of the cystic fibrosis defect in vitro by retrovirus-mediated gene transfer. Cell. 1990;62:1227–33.

    Article  CAS  Google Scholar 

  3. Collins FS, Drumm ML, Cole JL, et al. Construction of a general human chromosome jumping library, with application to cystic fibrosis. Science. 1987;235:1046–9.

    Article  CAS  Google Scholar 

  4. Castellani C, Assael BM. Cystic fibrosis: a clinical view. Cell Mol Life Sci. 2017;74(1):129–40 (Epub ahead of print).

    Article  CAS  Google Scholar 

  5. De Boeck K, Vermeulen F, Dupont L. The diagnosis of cystic fibrosis. Presse Med. 2017;46:e97–108.

    Article  Google Scholar 

  6. Sosnay PR, White TB, Farrell PM, et al. Diagnosis of cystic fibrosis in nonscreened populations. J Pediatr. 2017;181S:S52–S57.e2.

    Article  Google Scholar 

  7. Sathe M, Houwen R. Meconium ileus in cystic fibrosis. J Cyst Fibros. 2017;16(Suppl 2):S32–9.

    Article  Google Scholar 

  8. Castellani C, Massie J, Sontag M, et al. Newborn screening for cystic fibrosis. Lancet Respir Med. 2016;4:653–61.

    Article  Google Scholar 

  9. Rodrigues R, Magalhaes PKR, Fernandes MIM, et al. Neonatal screening for cystic fibrosis in São Paulo State, Brazil: a pilot study. 2009. www.bjournal.com.br. Accessed 2 June 2019.

  10. Athanazio RA, da Silva Filho LVRF, Vergara AA, et al. Brazilian guidelines for the diagnosis and treatment of cystic fibrosis. J Bras Pneumol. 2017;43:219–45.

    Article  Google Scholar 

  11. Servidoni MF, Gomez CCS, Marson FAL, et al. Sweat test and cystic fibrosis: overview of test performance at public and private centers in the state of São Paulo, Brazil. J Bras Pneumol. 2017;43:121–8.

    Article  Google Scholar 

  12. Taylor CJ, Hardcastle J, Southern KW. Physiological measurements confirming the diagnosis of cystic fibrosis: the sweat test and measurements of transepithelial potential difference. Paediatr Respir Rev. 2009;10:220–6.

    Article  CAS  Google Scholar 

  13. Farrell PM, Rosenstein BJ, White TB, et al. Guidelines for diagnosis of cystic fibrosis in newborns through older adults: Cystic Fibrosis Foundation consensus report. J Pediatr. 2008;153:S4–14.

    Article  Google Scholar 

  14. Rispoli T, Martins de Castro S, Grandi T, et al. A low-cost and simple genetic screening for cystic fibrosis provided by the Brazilian public health system. J Pediatr. 2018;199:272–7.

    Article  Google Scholar 

  15. Faucz FR, Souza DAS, Olandoski M, et al. CFTR allelic heterogeneity in Brazil: historical and geographical perspectives and implications for screening and counseling for cystic fibrosis in this country. J Hum Genet. 2010;55:71–6.

    Article  Google Scholar 

  16. Mousquer GT, Maciel LP, Pompeu Saraiva AC, et al. Validation of a quick and low-cost DNA extraction protocol applicable to long-stored blood samples. Anal Biochem. 2018;561–562:47–51.

    Article  Google Scholar 

  17. Grupo Brasileiro de Estudos de Fibrose Cística, The Brazilian Cystic Fibrosis Patient Registry, 2016. http://www.gbefc.org.br. Accessed 2 June 2019.

  18. Rodrigues R, Magalhaes PKR, Fernandes CS, et al. Neonatal screening for cystic fibrosis in São Paulo State, Brazil: a pilot study. Braz J Medical Res. 2009;42(10):973–8.

    Article  CAS  Google Scholar 

  19. Sih T, Godinho R, Franco LP, et al. Cystic fibrosis: Brazilian ENT experience. Int J Otolaryngol. 2012;2012:204696.

    Article  Google Scholar 

  20. Nunes LM, Ribeiro R, Niewiadonski VDT, et al. A new insight into CFTR allele frequency in Brazil through next generation sequencing. Pediatr Pulmonol. 2017;52:1300–5.

    Article  Google Scholar 

  21. Salinas DB, Sosnay PR, Azen C, et al. Benign and deleterious cystic fibrosis transmembrane conductance regulator mutations identified by sequencing in positive cystic fibrosis newborn screen children from California. PLoS One. 2016;11:e0155624.

    Article  Google Scholar 

  22. Latini FRM, Gazito D, Arnoni CP, et al. A new strategy to identify rare blood donors: single polymerase chain reaction multiplex SNaPshot reaction for detection of 16 blood group alleles. Blood Transfus. 2014;12(Suppl 1):S256–63.

    PubMed  PubMed Central  Google Scholar 

Download references

Author information

Author notes

  1. José Eduardo Vargas and Maria L. R. Rossetti contributed equally to this work.

Authors and Affiliations

  1. Programa de Pós-Graduação em Biologia Celular e Molecular, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil

    Thaiane Rispoli, Mayara J. Prado & Maria L. R. Rossetti

  2. Centro de Desenvolvimento Científico e Tecnológico, Secretaria da Saúde do Estado do Rio Grande do Sul (SES-RS), Porto Alegre, Rio Grande do Sul, Brazil

    Thaiane Rispoli, Grazielle M. Rodrigues, Mayara J. Prado, Cláudia M. D. da Silva & Tarciana Grandi

  3. Faculdade de Farmácia, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil

    Grazielle M. Rodrigues & Simone M. de Castro

  4. Serviço de Referência em Triagem Neonatal, Hospital Materno Infantil Presidente Vargas, Porto Alegre, Rio Grande do Sul, Brazil

    Simone M. de Castro & Helena T. Mocelin

  5. Hospital da Criança Santo Antônio, Irmandade da Santa Casa de Misericórdia de Porto Alegre, Porto Alegre, Rio Grande do Sul, Brazil

    Gilberto B. Fischer

  6. Hospital São Lucas da PUCRS, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil

    Leonardo A. Pinto

  7. Hospital de Clínicas de Porto Alegre, Porto Alegre, Rio Grande do Sul, Brazil

    Paulo J. C. Maróstica

  8. Programa de Pós-Graduação em Ciências Pneumológicas, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil

    Laís C. R. Scortegagna

  9. Instituto de Ciências Biológicas, Universidade de Passo Fundo, Passo Fundo, Rio Grande do Sul, Brazil

    José Eduardo Vargas

  10. Programa de Pós-Graduação em Biologia Molecular Aplicada a Saúde, Canoas, Universidade Luterana do Brazil, Canoas, Rio Grande do Sul, Brazil

    Maria L. R. Rossetti

Authors
  1. Thaiane Rispoli
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Grazielle M. Rodrigues
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Simone M. de Castro
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Mayara J. Prado
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Cláudia M. D. da Silva
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Tarciana Grandi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Gilberto B. Fischer
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Leonardo A. Pinto
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Paulo J. C. Maróstica
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Laís C. R. Scortegagna
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Helena T. Mocelin
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. José Eduardo Vargas
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Maria L. R. Rossetti
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Thaiane Rispoli or José Eduardo Vargas.

Ethics declarations

Funding

The study was financed in part by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) (Finance Code 001) and Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS) (1260-2551/13-0).

Ethics approval

This study was approved by the ethics committee of the Secretaria da Saúde do Estado do Rio Grande do Sul, Universidade Federal do Rio Grande do Sul, Hospital Materno Infantil Presidente Vargas, Hospital da Criança Santo Antônio, Hospital São Lucas da PUCRS, and Hospital de Clínicas de Porto Alegre.

Patient consent

Obtained.

Conflict of interest

The authors, Thaiane Rispoli, Grazielle M. Rodrigues, Simone M. de Castro, Mayara J. Prado, Cláudia M.D. da Silva, Tarciana Grandi, Gilberto B Fischer, Leonardo A. Pinto, Paulo J.C. Maróstica, Laís C.R. Scortegagna, Helena T. Mocelin, José E. Vargas, and Maria L.R. Rossetti, have no conflict of interest to declare.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Rispoli, T., Rodrigues, G.M., de Castro, S.M. et al. Cystic Fibrosis: A Simple and Customized Strategy for Genetic Screening Able to Detect Over 90% of Identified Mutated Alleles in Brazilian Newborns. Mol Diagn Ther 24, 315–325 (2020). https://doi.org/10.1007/s40291-020-00456-9

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s40291-020-00456-9

Search

Navigation