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    Flow chart for hepatitis B vaccination performed in rural settlers in the southwest region of central Brazil, 2011.

  • 1.

    Michel ML, Tiollais P, 2010. Hepatitis B vaccines: protective efficacy and therapeutic potential. Pathol Biol (Paris) 58: 288295.

  • 2.

    Franco E, Bagnato B, Marino MG, Meleleo C, Serino L, Zaratti L, 2012. Hepatitis B: epidemiology and prevention in developing countries. World J Hepatol 4: 7480.

    • Search Google Scholar
    • Export Citation
  • 3.

    Zanetti AR, Van Damme P, Shouval D, 2008. The global impact of vaccination against hepatitis B: a historical overview. Vaccine 26: 62666273.

  • 4.

    Shouval D, Ilan Y, Adler R, Deepen R, Panet A, Even-Chen Z, Gorecki M, Gerlich WH, 1994. Improved immunogenicity in mice of a mammalian cell-derived recombinant hepatitis B vaccine containing pre-S and pre-S2 antigens as compared with conventional yeast-derived vaccines. Vaccine 12: 14531459.

    • Search Google Scholar
    • Export Citation
  • 5.

    Martins RM, Bensabath G, Arraes LC, Oliveira MLA, Miguel JC, Barbosa GG, Camacho LAB, 2004. Multicenter study on the immunogenicity and safety of two recombinant vaccines against hepatitis B. Mem Inst Oswaldo Cruz 99: 865871.

    • Search Google Scholar
    • Export Citation
  • 6.

    Moraes JC, Luna EJA, Grimaldi RA, 2010. Imunogenicidade da vacina brasileira contra hepatite B em adultos. Rev Saude Publica 44: 353359.

  • 7.

    Ministério da Saúde Brasil, 2015. Protocolo Clínico e Diretrizes Terapêuticas para Atenção Integral às Pessoas com Infecções Sexualmente Transmissíveis. Departamento de DST, Aids e Hepatites Virais, ed. Brasília (Brasil): Ministério da Saúde, Secretaria de Vigilância em Saúde, Departamento de DST, Aids e Hepatites Virais, 120.

    • Search Google Scholar
    • Export Citation
  • 8.

    Souto FJD, 2016. Distribution of hepatitis B infection in Brazil: the epidemiological situation at the beginning of the 21st century. Rev Soc Bras Med Trop 49: 1123.

    • Search Google Scholar
    • Export Citation
  • 9.

    Ximenes RAA, Figueiredo GM, Cardoso MRA, Stein AT, Moreira RC, Coral G, Crespo D, Santos AA, Montarroyos UR, Braga MC, Pereira LMMB, 2015. Population-based multicentric survey of hepatitis B infection and risk factors in the north, south, and southeast regions of Brazil, 10–20 years after the beginning of vaccination. Am J Trop Med Hyg 93: 13411348.

    • Search Google Scholar
    • Export Citation
  • 10.

    Vermeiren AP, Hoebe CJ, Dukers-Muijrers NH, 2013. High non-responsiveness of males and the elderly to standard hepatitis B vaccination among a large cohort of healthy employees. J Clin Virol 58: 262264.

    • Search Google Scholar
    • Export Citation
  • 11.

    Chathuranga LS, Noordeen F, Abeykoon AM, 2013. Immune response to hepatitis B vaccine in a group of health care workers in Sri Lanka. Int J Infect Dis 17: 10781079.

    • Search Google Scholar
    • Export Citation
  • 12.

    Motta-Castro ARC, Gomes SA, Yoshida CFT, Miguel JC, Teles SA, Martins RMB, 2009. Compliance with and response to hepatitis B vaccination in remaining quilombo communities in central Brazil. Cad Saude Publica 25: 738742.

    • Search Google Scholar
    • Export Citation
  • 13.

    Klein SL, Marriott I, Fish EN, 2015. Sex-based differences in immune function and responses to vaccination. Trans R Soc Trop Med Hyg 109: 915.

    • Search Google Scholar
    • Export Citation
  • 14.

    Gilbert CL, Klopfer SO, Martin JC, Schödel FP, Bhuyan PK, 2011. Safety and immunogenicity of a modified process hepatitis B vaccine in healthy adults ≥50 years. Hum Vaccin 7: 13361342.

    • Search Google Scholar
    • Export Citation
  • 15.

    Kulkarni PS, Raut SK, Patki PS, Phadke MA, Jadhav SS, Kapre SV, Dhorje SP, Godse SR, 2006. Immunogenicity of a new, low-cost recombinant hepatitis B vaccine derived from Hansenula polymorpha in adults. Vaccine 24: 34573460.

    • Search Google Scholar
    • Export Citation
  • 16.

    Meeren OVD, Crasta P, Cheuvart B, Ridder MD, 2015. Characterization of an age-response relationship to GSK's recombinant hepatitis B vaccine in healthy adults: an integrated analysis. Hum Vaccin Immunother 11: 17261729.

    • Search Google Scholar
    • Export Citation
  • 17.

    Rosenberg C, Bovin NV, Bram LV, Flyvbjerg E, Erlandsen M, Vorup-Jensen T, Petersen E, 2013. Age is an important determinant in humoral and T cell responses to immunization with hepatitis B surface antigen. Hum Vaccin Immunother 9: 14661476.

    • Search Google Scholar
    • Export Citation
  • 18.

    Vijayakumar V, Hari R, Parthiban R, Mehta J, Thyagarajan SP, 2004. Evaluation of immunogenicity and safety of Genevac B: a new recombinant hepatitis b vaccine in comparison with Engerix B and Shanvac B in healthy adults. Indian J Med Microbiol 22: 3438.

    • Search Google Scholar
    • Export Citation
  • 19.

    Ministério da Saúde Brasil, 2015. Protocolo Clínico e Diretrizes Terapêuticas para Atenção Integral às Pessoas com Infecções Sexualmente Transmissíveis. Departamento de DST, Aids e Hepatites Virais, ed. Brasília (Brasil): Ministério da Saúde, Secretaria de Vigilância em Saúde, Departamento de DST, Aids e Hepatites Virais, 120.

    • Search Google Scholar
    • Export Citation

 

 

 

 

Low Immunogenicity of Recombinant Hepatitis B Vaccine Derived from Hansenula polymorpha in Adults Aged Over 40 Years

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  • 1 Nursing School, Federal University of Goias, Goiania, Brazil.
  • 2 Institute of Tropical Pathology and Public Health, Federal University of Goias, Goiania, Brazil.
  • 3 Nursing School, State University of Mato Grosso do Sul, Dourados, Brazil.

The Brazilian recombinant hepatitis B vaccine (VrHB-IB) is based on the expression of the recombinant antigen in Hansenula polymorpha yeast cells. Currently, data on the immunogenicity of this vaccine in older adults are nonexistent. This study aimed to evaluate the immunogenicity of VrHB-IB in adults over 40 years of age. From May to October 2011, 235 rural settlers between 2 and 93 years of age from the State of Goias in Brazil were eligible for vaccination. Of these, 180 accepted the first dose of the vaccine and 106 (58.9%) completed the vaccination schedule. Multivariate analysis revealed that individuals ≥ 40 years of age responded significantly less well to vaccination than younger adults. Also, a greater proportion of male nonresponders was observed (versus women; P = 0.02). These results point to the need for better evaluation of the immunogenicity of VrHB-IB in older adults.

Vaccination against hepatitis B is the safest and most effective method of reducing the incidence and prevalence of infection with hepatitis B virus (HBV).1 Indeed, HBV vaccination confers protection in over 90% of subjects who receive it.2 The first-generation HBV vaccine, which was produced from human HBV surface antigen (HBsAg) plasma from chronically infected patients, became commercially available in the early 1980s and is still used in some countries.3 The second-generation vaccine, also marketed in the 1980s, is produced using recombinant DNA technology. This vaccine is safe and effective, and allows for unlimited production and widespread distribution throughout the world. Currently, many countries produce the HBV vaccine using this technology, and various expression systems have been used to obtain recombinant HBsAg.1,4 Engerix-B, the gold-standard vaccine, uses Saccharomyces cerevisiae as the expression system for recombinant HBsAg.1

In Brazil, the Brazilian recombinant hepatitis B vaccine (VrHB-IB) is obtained by culturing genetically engineered Hansenula polymorpha yeast cells with the HBsAg gene. Initially, this vaccine was produced as a recombinant antigen (concentration 20 μg/mL). However, research has shown that in adults 30–40 years of age, the immunogenicity of the Brazilian vaccine is not equivalent to the gold-standard vaccine (Engerix B),5 and the concentration of HBsAg was therefore increased to 25 μg/mL. A clinical trial comparing the immunogenicity of VrHB-B with Engerix-B in patients in this age group found that the vaccine produced in Brazil at the current concentration is equivalent to the gold standard.6

The aim of our study was to evaluate the immunogenicity of VrHB-IB vaccination in rural settlers in central Brazil. Rural settlers who were susceptible to HBV comprised the vaccination group, and most of them were > 40 years of age. At present, published studies on the immunogenicity of the Brazilian HBV vaccine in adults in this age group are rare.

From May to October 2011, 467 rural settlers from the southwest region of the State of Goias, central Brazil, were recruited. All agreed to participate in the study and signed a consent form. The participants were tested initially for HBV markers (HBsAg, anti-HBc, and anti-HBs). Those who were negative for all serologic markers and had not reported a previous vaccination history for hepatitis B were identified as susceptible to the virus and eligible for vaccination. This study was approved by the ethics committee of the Federal University of Goias (protocol CEPMHA/HC/UFG No. 127/2010).

The following Brazilian VrHB-IB vaccine lot numbers (Instituto Butantan, Sao Paulo, Brazil) were used: 1006117 and 1006116. Each mL of vaccine contains 25 μg/mL of recombinant HBsAg expressed in H. polymorpha. The vaccine was administered intramuscularly in three doses, with an interval of 1 month between the first and second doses, and 5 months between the second and third doses. Forty-five to 60 days after the third vaccination dose, eligible individuals were contacted for blood collection and quantitative detection of anti-HBs markers, which were measured using a microparticle enzyme immunoassay (AxSYM® Ausab®; Abbott, Wiesbaden, Germany). Seropositivity was defined as an anti-HBs antibody concentration ≥ 1 mIU/mL, and seroprotection as ≥ 10 mIU/mL.

Data were analyzed using the statistical program, STATA 11 (StataCorp., College Station, TX). Geometric mean anti-HBs titers were calculated using a confidence interval (CI) of 95%. The χ2 test and Fisher's exact test were used to test the significance of the differences between the proportions. P values of < 0.05 were considered statistically significant. A total of 77 subjects were sufficient to achieve 80% power to detect a difference in seroprotection rate of 35% between nonresponder and responder groups. The significance level of the test was 0.05.

Of the total study participants (467), 51 (10.9%; 95% CI = 8.4–14.1) presented some serological markers of exposure to infection by HBV. In 90 individuals (19.3%; 95% CI = 15.9–23.1), positivity was observed for isolated anti-HBs markers, indicating previous vaccination against hepatitis B.

Ninety-one individuals reported previous hepatitis B vaccination histories. Therefore, nearly half of the participants (235/467) between 2 and 93 years of age were susceptible to infection and were eligible for vaccination. Of those, 180 accepted the first dose of the vaccine and 106 (58.9%; 106/180) completed the vaccination schedule. The vaccine response was evaluated for 77 individuals (Figure 1).

Figure 1.
Figure 1.

Flow chart for hepatitis B vaccination performed in rural settlers in the southwest region of central Brazil, 2011.

Citation: The American Society of Tropical Medicine and Hygiene 96, 1; 10.4269/ajtmh.16-0475

The characteristics of the individuals who completed the vaccination series and were evaluated for their vaccine responses were similar to those susceptible to hepatitis B, except for age and marital status (Table 1). Over half of the individuals were female (51.9%) and almost half had up to 4 years of primary-level schooling (49.3%). Also, most of these were married (87%), native to the midwest region of Brazil (71.4%), and > 40 years of age (71.4%). Only 68.8% (53/77) had protective anti-HBs titers, and of these, 30 developed titers > 100 mIU/mL. The geometric mean of the anti-HBs antibodies was 27.92 mIU/mL. Multivariate analysis revealed that individuals ≥ 40 years of age responded significantly less well to vaccination than young adults. Also, men comprised the greater proportion of nonresponders (versus women; P = 0.02) (Table 2).

Table 1

Characteristics of participants eligible for HBV vaccination (N = 235), central Brazil, 2011

VariablesParticipants who refused vaccination (%)Participants who started the vaccine series (%)Participants vaccinated and evaluated for immunogenicity (%)Total (%)P value
N = 55N = 103N = 77
Gender
 Female22 (40)52 (50.5)40 (51.9)114 (48.5)0.346
 Male33 (60)51 (49.5)37 (48.1)121 (51.5)
Age (years; range 2–93)
 ≤ 3941 (74.5)33 (32)22 (28.6)96 (40.9)0.000
 40–497 (12.7)23 (22.3)21 (27.3)51 (21.7)
 ≥ 507 (12.7)47 (45.6)34 (44.1)88 (37.4)
Civil status
 Married/steady partner15 (27.3)81 (78.6)67 (87)163 (69.4)0.000
 Single40 (72.7)16 (15.5)8 (10.4)64 (27.2)
 Separated0 (0)2 (1.9)1 (1.3)3 (1.3)
 Widowed0 (0)4 (3.9)1 (1.3)5 (2.1)
Education (years)
 ≤ 419 (40.4)48 (46.6)38 (49.3)105 (46.3)0.718
 5–821 (44.7)40 (38.8)25 (32.5)86 (37.9)
 ≥ 97 (14.9)15 (14.6)14 (18.2)36 (15.9)
 NA: 8
Region
 Midwestern47 (85.5)84 (81.6)55 (71.4)186 (79.1)0.380
 Northeast4 (7.3)7 (6.8)14 (18.2)25 (10.6)
 North0 (0)0 (0)1 (1.3)1 (0.4)
 Southeast1 (6.7)9 (8.7)5 (6.5)15 (6.4)
 South3 (5.5)3 (2.9)2 (2.6)8 (3.4)
Tobacco user
 No28 (87.5)74 (72.5)54 (70.1)156 (73.9)0.154
 Yes4 (12.5)28 (27.5)23 (29.9)55 (26.1)
 NA: 24

HBV = hepatitis B virus; NA = not applicable.

Table 2

Characteristics associated with seroprotection after vaccination schedule for hepatitis B in 77 individuals, central Brazil, 2011

 Univariate analysisMultivariate analysis*
ResponderNonresponderP value
N = 53 (%)N = 24 (%)OR (95% CI)P value
Age (years)  0.003  
 ≤ 3921 (39.6)1 (4.2) 1.0 
 40–4913 (24.5)8 (33.3) 11.4 (1.1–115.6)0.04
 ≥ 5019 (35.9)15 (62.5) 14.2 (1.6–124.9)0.02
Tobacco user11 (20.8)12 (50.0)0.0091.55 (0.8–8.5)0.12
Male20 (37.7)17 (70.8)0.0073.8 (1.2–12.3)0.02

CI = confidence interval; OR = odds ratio.

Logistic regression.

In 1998, the Ministry of Health of Brazil began offering the HBV vaccine free of charge for babies up to 1 year of age, a service that was gradually extended to older people. Nowadays, HBV vaccine is offered to the entire population, regardless of age or vulnerability.7 Despite this policy, differences exist in the vaccination uptake across different age groups and low immunization rates are apparent in some groups.8,9 Also, in older age groups, the immunogenicity of this vaccine is not known, and there are many factors that may affect how immunogenic the vaccine is in these groups.1

As has been observed for the gold-standard vaccine by other authors,1012 sex represents a statistically significant difference in terms of vaccine responders and nonresponders. Different biological characteristics and behaviors contribute to men developing a weaker immune response to vaccination than women.13

The concentration of VrHB-IB is greater than recombinant HBsAg to the gold standard, Engerix-B.6,14,15 According to Moraes and others,6 with an HBsAg concentration of 25 μg/mL, the immunogenicity of the vaccine produced in Brazil for adults 31–40 years of age is equivalent to the gold standard, for which 98.6% of the vaccinees developed protective titers. In fact, in subjects < 40 years of age, the vaccine response was equivalent to that reported by other authors using vaccines expressed in S. cerevisiae (Engerix-B)16 and H. polymorpha (GeneVac B).15 In older individuals, surprisingly, there was a much lower percentage of seroprotection than desired in an age range of the Brazilian population that does not present comparative data. Among those aged 40–49 and 50–59 years, only 61.9% and 55.9% were responders, respectively. Investigations using the gold-standard vaccine illustrate higher response rates in older adults,10,14 as was also found in a study using the GeneVac-B vaccine.15

Indeed, in a multicenter study of 144 individuals ≥ 50 years of age who received three doses of Engerix-B, 84% developed protective titers.14 In the Netherlands, 11,439 individuals were evaluated after completing the vaccination schedule and, for men and women, a response rate of 90% was found for ages 41 and 53, respectively. In the same study, this rate declined to 80% for male and female vaccinees 54 and 68 years of age, respectively.10 Concerning the GeneVac-B vaccine, a greater response rate was also found in older individuals compared with the data from this investigation. Those aged 40–49 years and 50–57 years, which represents 93% and 85% of the participants, respectively, developed protective titers.15

Immune system aging, which is referred to as immunosenescence, affects responses to the HBV vaccine17; hence, the immunogenicity rate for this vaccine in older individuals is generally lower. However, our findings suggest that the immunogenicity of the Brazilian recombinant vaccine is even lower in individuals > 40 years of age compared with the gold-standard vaccine and others.6,18 The greatest discrepancy in protection rates was found between the two vaccines that are expressed in H. polymorpha, and which also use the same adsorbent (aluminum hydroxide).6,18 Therefore, it seems likely that factors such as storage, transport, and packaging may influence the effectiveness of the Brazilian vaccine. However, all these factors were thoroughly checked during the developmental stages of this investigation, according to Brazilian recommendations.19

This study has some limitations. Rural settlers are not representative of the national population as a whole. Also, budget constraints meant we were unable to form a control group and evaluate the response rates using the gold-standard vaccine; however, our findings for subjects < 40 years of age were similar to those of the gold-standard vaccine.16 Factors that may have interfered with vaccine responses, such as obesity and immunosuppression, were not investigated. Furthermore, the vaccine response was evaluated in only 77 individuals who represent 43% of those who received the first vaccine dose. Nevertheless, these results represent the first data on Brazilian vaccine immune responses among people ≥ 40 years of age, and point to the need for better evaluation of the immunogenicity of the Brazilian HBV vaccine in older adults. With the gradual proliferation of such a vaccine, the burden of infection is moving to older age groups, whereas the results of this study suggest that poor vaccine responses occur among individuals in these groups.

ACKNOWLEDGMENTS

We thank the Center for Research on Epidemiology and Infectious Diseases of the Federal University of Goias Nursing School for their excellent assistance during data collection.

  • 1.

    Michel ML, Tiollais P, 2010. Hepatitis B vaccines: protective efficacy and therapeutic potential. Pathol Biol (Paris) 58: 288295.

  • 2.

    Franco E, Bagnato B, Marino MG, Meleleo C, Serino L, Zaratti L, 2012. Hepatitis B: epidemiology and prevention in developing countries. World J Hepatol 4: 7480.

    • Search Google Scholar
    • Export Citation
  • 3.

    Zanetti AR, Van Damme P, Shouval D, 2008. The global impact of vaccination against hepatitis B: a historical overview. Vaccine 26: 62666273.

  • 4.

    Shouval D, Ilan Y, Adler R, Deepen R, Panet A, Even-Chen Z, Gorecki M, Gerlich WH, 1994. Improved immunogenicity in mice of a mammalian cell-derived recombinant hepatitis B vaccine containing pre-S and pre-S2 antigens as compared with conventional yeast-derived vaccines. Vaccine 12: 14531459.

    • Search Google Scholar
    • Export Citation
  • 5.

    Martins RM, Bensabath G, Arraes LC, Oliveira MLA, Miguel JC, Barbosa GG, Camacho LAB, 2004. Multicenter study on the immunogenicity and safety of two recombinant vaccines against hepatitis B. Mem Inst Oswaldo Cruz 99: 865871.

    • Search Google Scholar
    • Export Citation
  • 6.

    Moraes JC, Luna EJA, Grimaldi RA, 2010. Imunogenicidade da vacina brasileira contra hepatite B em adultos. Rev Saude Publica 44: 353359.

  • 7.

    Ministério da Saúde Brasil, 2015. Protocolo Clínico e Diretrizes Terapêuticas para Atenção Integral às Pessoas com Infecções Sexualmente Transmissíveis. Departamento de DST, Aids e Hepatites Virais, ed. Brasília (Brasil): Ministério da Saúde, Secretaria de Vigilância em Saúde, Departamento de DST, Aids e Hepatites Virais, 120.

    • Search Google Scholar
    • Export Citation
  • 8.

    Souto FJD, 2016. Distribution of hepatitis B infection in Brazil: the epidemiological situation at the beginning of the 21st century. Rev Soc Bras Med Trop 49: 1123.

    • Search Google Scholar
    • Export Citation
  • 9.

    Ximenes RAA, Figueiredo GM, Cardoso MRA, Stein AT, Moreira RC, Coral G, Crespo D, Santos AA, Montarroyos UR, Braga MC, Pereira LMMB, 2015. Population-based multicentric survey of hepatitis B infection and risk factors in the north, south, and southeast regions of Brazil, 10–20 years after the beginning of vaccination. Am J Trop Med Hyg 93: 13411348.

    • Search Google Scholar
    • Export Citation
  • 10.

    Vermeiren AP, Hoebe CJ, Dukers-Muijrers NH, 2013. High non-responsiveness of males and the elderly to standard hepatitis B vaccination among a large cohort of healthy employees. J Clin Virol 58: 262264.

    • Search Google Scholar
    • Export Citation
  • 11.

    Chathuranga LS, Noordeen F, Abeykoon AM, 2013. Immune response to hepatitis B vaccine in a group of health care workers in Sri Lanka. Int J Infect Dis 17: 10781079.

    • Search Google Scholar
    • Export Citation
  • 12.

    Motta-Castro ARC, Gomes SA, Yoshida CFT, Miguel JC, Teles SA, Martins RMB, 2009. Compliance with and response to hepatitis B vaccination in remaining quilombo communities in central Brazil. Cad Saude Publica 25: 738742.

    • Search Google Scholar
    • Export Citation
  • 13.

    Klein SL, Marriott I, Fish EN, 2015. Sex-based differences in immune function and responses to vaccination. Trans R Soc Trop Med Hyg 109: 915.

    • Search Google Scholar
    • Export Citation
  • 14.

    Gilbert CL, Klopfer SO, Martin JC, Schödel FP, Bhuyan PK, 2011. Safety and immunogenicity of a modified process hepatitis B vaccine in healthy adults ≥50 years. Hum Vaccin 7: 13361342.

    • Search Google Scholar
    • Export Citation
  • 15.

    Kulkarni PS, Raut SK, Patki PS, Phadke MA, Jadhav SS, Kapre SV, Dhorje SP, Godse SR, 2006. Immunogenicity of a new, low-cost recombinant hepatitis B vaccine derived from Hansenula polymorpha in adults. Vaccine 24: 34573460.

    • Search Google Scholar
    • Export Citation
  • 16.

    Meeren OVD, Crasta P, Cheuvart B, Ridder MD, 2015. Characterization of an age-response relationship to GSK's recombinant hepatitis B vaccine in healthy adults: an integrated analysis. Hum Vaccin Immunother 11: 17261729.

    • Search Google Scholar
    • Export Citation
  • 17.

    Rosenberg C, Bovin NV, Bram LV, Flyvbjerg E, Erlandsen M, Vorup-Jensen T, Petersen E, 2013. Age is an important determinant in humoral and T cell responses to immunization with hepatitis B surface antigen. Hum Vaccin Immunother 9: 14661476.

    • Search Google Scholar
    • Export Citation
  • 18.

    Vijayakumar V, Hari R, Parthiban R, Mehta J, Thyagarajan SP, 2004. Evaluation of immunogenicity and safety of Genevac B: a new recombinant hepatitis b vaccine in comparison with Engerix B and Shanvac B in healthy adults. Indian J Med Microbiol 22: 3438.

    • Search Google Scholar
    • Export Citation
  • 19.

    Ministério da Saúde Brasil, 2015. Protocolo Clínico e Diretrizes Terapêuticas para Atenção Integral às Pessoas com Infecções Sexualmente Transmissíveis. Departamento de DST, Aids e Hepatites Virais, ed. Brasília (Brasil): Ministério da Saúde, Secretaria de Vigilância em Saúde, Departamento de DST, Aids e Hepatites Virais, 120.

    • Search Google Scholar
    • Export Citation

Author Notes

* Address correspondence to Karlla Antonieta Amorim Caetano, Faculdade de Enfermagem, Universidade Federal de Goias, Rua 227, s/n–Setor Leste Universitário, Goiania 74605-220, Brazil. E-mail: karllacaetano@gmail.com

Authors' addresses: Karlla Antonieta Amorim Caetano, Raquel Silva Pinheiro, and Sheila Araujo Teles, Faculdade de Enfermagem, Universidade Federal de Goias, Goiania, Brazil, E-mails: karllacaetano@gmail.com, raquelsilva932@gmail.com, and sheila.fen@gmail.com. Nativa Helena Alves Del-Rios and Megmar Aparecida dos Santos Carneiro, Instituto de Patologia Tropical e Saúde Pública, Universidade Federal de Goias, Goiania, Brazil, E-mails: nativa_rios@yahoo.com.br and megmar242@gmail.com. Fabiana Perez Rodrigues Bergamaschi, Ciências da Saúde, Universidade Estadual do Mato Grosso do Sul, Dourados, Brazil, E-mail: fa.rodrigues@hotmail.com.

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