SciELO - Scientific Electronic Library Online

 
vol.50 issue3EARLY FUNCTIONAL RESULTS OF BIOFEEDBACK AND ITS IMPACT ON QUALITY OF LIFE OF PATIENTS WITH ANAL INCONTINENCEARE THE SPIDER ANGIOMAS SKIN MARKERS OF HEPATOPULMONARY SYNDROME? author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand

Article

Indicators

Related links

  • Have no similar articlesSimilars in SciELO

Share


Arquivos de Gastroenterologia

Print version ISSN 0004-2803

Arq. Gastroenterol. vol.50 no.3 São Paulo July/Sept. 2013

http://dx.doi.org/10.1590/S0004-28032013000200030 

Original Articles

ROLE OF HELICOBACTER PYLORI INFECTION AND LIFESTYLE HABITS IN THE DEVELOPMENT OF GASTRODUODENAL DISEASES IN A POPULATION FROM THE BRAZILIAN AMAZON

Papel da infecção por Helicobacter pylori e hábitos de vida no desenvolvimento das doenças gastroduodenais em uma população da amazônia brasileira

Ruth Maria Dias Ferreira VINAGRE1, Adenielson VILAR-e-SILVA2, Amanda Alves FECURY2, Luisa Caricio MARTINS2

1Departamento de Clínica Médica, Hospital Ophir Loyola, Belém, PA, Brasil.

2Laboratório de Clínica e Patologia de Doenças Tropicais da Universidade do Pará, Belém, PA, Brasil.

ABSTRACT

Context

Although more than half of the world's population is colonized with Helicobacter pylori, it remains unknown why this organism is able to produce severe disease in some hosts and be innocuous in others. The clinical outcome of infection is determined by several factors, including differences in the host response to bacterial stimulation, specific virulence factors of the organism and environmental influences, or a combination of these factors.

Objectives

This study compared the prevalence of H. pylori infection and risk factors (infection with CagA+ strains, excessive alcohol consumption, smoking, and inadequate eating habits) between patients with different gastrointestinal disorders and associated these risk factors with the histopathological findings.

Methods

In a prospective study, samples were collected from 442 patients and a standardized questionnaire regarding lifestyle habits (excessive alcohol consumption, smoking, and eating habits) was applied. The presence of H. pylori and of the cagA gene was investigated by polymerase chain reaction (PCR). Gastric biopsies were obtained for histological assessment.

Results

The frequency of alcohol consumption, smoking, inadequate diet and infection with CagA+ H. pylori was higher among patients with peptic ulcer and adenocarcinoma when compared to those with gastritis. Gastric inflammation was more pronounced in patients infected with CagA+ strains.

Conclusion

We conclude that infection with CagA+ H. pylori strains, excessive alcohol consumption, smoking and inadequate eating habits increase the risk of developing peptic ulcer and gastric carcinoma.

Key words: Helicobacter pylori ; Gastrointestinal diseases; Adenocarcinoma; Peptic ulcer; Amazonian ecosystem

RESUMO

Contexto

Apesar de o H. pylori colonizar o estômago de aproximadamente metade da população mundial, ainda se desconhece por quê esse organismo é capaz de causar doença severa em certos hospedeiros e ser inofensivo em outros. As manifestações clínicas da infecção são determinadas por vários fatores, tais como: diferentes respostas do hospedeiro ao estímulo bacteriano, fatores de virulência específicos do organismo e influências ambientais ou a combinação desses fatores.

Objetivos

Esse estudo tem como objetivo comparar a prevalência e a associação dos fatores de risco, tais como: infecção por cepas H. pylori CagA+, consumo excessivo de bebidas alcoólicas, uso de tabaco e hábitos inadequados de alimentação entre pacientes com diferentes doenças gastrointestinais e a associação deles com achados histopatológicos.

Métodos

Em estudo prospectivo, foram cole-tadas amostras de 442 pacientes submetidos a um questionário padronizado com perguntas sobre hábitos de vida (uso excessivo de álcool, tabaco e, hábitos alimentares). A presença do H. pylori e do gene cagA foi detectada utilizando a Reação em Cadeia da Polimerase (PCR). As biópsias gástricas foram avaliadas histologicamente.

Resultados

O consumo de bebida alcoólica, uso de tabaco, dieta inadequada e infecção por cepas H. pylori CagA+ foram maiores entre pacientes com úlcera péptica e adenocarcinoma do que em pacientes com gastrite. Os pacientes infectados por cepas H. pylori CagA+ apresentaram inflamação gástrica de maior intensidade.

Conclusão

A presença de infecção por cepas H. pylori CagA+, o consumo excessivo de álcool, de tabaco e hábitos de alimentação inadequados aumentam o risco de desenvolvimento de úlcera péptica e carcinoma gástrico.

Palavras-Chave: Helicobacter pylori ; Gastroenteropatias; Adenocarcinoma; Úlcera péptica; Ecossistema amazônico

INTRODUCTION

Helicobacter pylori colonizes the human stomach mucosa, causing chronic gastritis, and is an important risk factor for the development of peptic ulcers and gastric carcinoma(21). However, only some infected individuals develop severe diseases(16). Various factors, particularly bacterial virulence factors, have been studied in an attempt to explain this clinical diversity in H. pylori infection(11). One virulence factor is cytotoxin-associated gene A (CagA), an immunodominant protein with a high molecular weight (140-160 kDa) which is encoded by the cagA gene present on the cag pathogenicity island. The CagA protein is injected into gastric epithelial cells by a type IV secretion system and induces rearrangements of the actin cytoskeleton that alter the morphology and function of epithelial cells(20). Studies have demonstrated a high frequency of anti-CagA IgG antibodies in patients with atrophic gastritis, peptic ulcers, and gastric carcinoma(4 27).

The combined presence of adverse environmental factors, such as unhealthy lifestyle habits of the patients, and H. pylori infection has been associated with progression to more severe gastroduodenal disease. Excessive alcohol consumption, smoking and inadequate eating habits, such as excess salt, fats and canned products, have been related to the development of peptic ulcers and stomach cancer(12 18 24).

The objective of the present study was to compare the prevalence of H. pylori infection and risk factors (bacterial virulence factors and lifestyle habits) between patients with different gastrointestinal diseases and to associate these risk factors with the histopathological findings.

METHODS

Patients

Gastric biopsy samples were collected from 442 patients with different gastrointestinal disorders seen at the Endoscopy Department of the Ofir Loiola Hospital between 2010 and 2011.

Four biopsy fragments were obtained from the stomach of each patient during endoscopy: two from the gastric lesion for histological analysis and two from the antrum for analysis by molecular methods. In cases suggestive of malignancy, biopsies were obtained from the cancer lesion and the adjacent area (perilesion) for histological analysis. None of the patients had received antimicrobial drugs, H2-receptor antagonists, acid pump inhibitors, nonsteroidal anti-inflammatory drugs, or any medication for at least 60 days prior to endoscopy.

All patients included in the study were of the same socioeconomic level, had similar cultural habits, were born in the state of Pará, and had the same ethnic background (approximately 50% Portuguese, 40% Amerindian, and 10% African)(3). The study was approved by the Ethics Committee of the Nucleus of Tropical Medicine, Federal University of Pará.

Evaluation of lifestyle habits

A standardized questionnaire regarding lifestyle habits (excessive alcohol consumption, smoking, and eating habits) was used. Average drinking levels were evaluated to calculate the amount of alcohol consumed: sugarcane rum = 40%; beer = 5%; wine = 12%; other distilled beverages = 55%. The threshold of alcohol consumption was standardized by calculating average daily intake in grams, with an acceptable dose of 70 g alcohol per week for women and 140 g for men(7).

Smoking intensity was defined as the total number of cigarettes consumed per day, with each hand-rolled cigarette being equivalent to five manufactured cigarettes(17). Subjects smoking more than five cigarettes per day were classified as smokers and those smoking fewer than five cigarettes per day or did not smoke at all were classified as non-smokers. The patients were also divided into two groups according to age (>50 years and <50 years).

Dietary habits were classified as inadequate when the subjects reported to consume large amounts of salty, fatty and canned foods and low amounts of fruits and vegetable, and as adequate when the subjects reported to frequently consume fruits and vegetables and to rarely consume fatty, salty and canned foods.

Extraction of DNA and PCR

Gastric fragments were collected from the antrum for histopathological analysis and extraction of bacterial DNA(14). Total DNA was extracted from frozen gastric biopsy specimens by adding 10 µl proteinase K and 300 µL lysis buffer (200 mM Tris-HCl, 25 mM EDTA, 300 mM NaCl, 1.2% sodium dodecyl sulfate) and incubating the mixture for 12 h at 55°C. The lysate was extracted with an equal volume of phenol-chloroform, precipitated with isopropanol, and washed with 70% ethanol. The pellet was dried and resuspended in 200 µL sterile distilled water. Extracted DNA was stored at - 20°C until the time for analysis.

A set of primers (p1 and p2)(10) amplifying a fragment of 298 bp of the urease gene present in all H. pylori strains was used to detect bacterial DNA. Only positive samples were used for amplification of the cagA gene. The protocol and primers described by Tumuru et al.(25) were used for amplification of the cagA gene. Negative and positive controls were included in all reactions. The PCR products were submitted to electrophoresis on 2% agarose gel stained with ethidium bromide and visualized under a UV transilluminator.

Histopathological analysis

The gastric biopsies were stained with hematoxylin-eosin for histological analysis. The histopathological parameters were scored from 0 to 3 according to the Sydney classification system(6) for the analysis of polymorphonuclear and mononuclear cell infiltration.

Statistical analysis

Odds ratios and G-tests were used to evaluate the proportion of risk factors in the control and patient groups. The level of significance was set at 95%. Statistical analysis was performed using the Biostat 5.0 program(2).

RESULTS

Of the 442 patients with different gastrointestinal diseases studied, 45% (198/442) had gastritis, 17% (78/442) had gastric ulcer, 22% (96/442) had duodenal ulcer, and 16% (70/442) gastric adenocarcinoma (intestinal type). Among patients with gastritis, all had superficial gastritis based on the endoscopy report and a second histopathological analysis revealed chronic gastritis without atrophy in all patients.

Excessive alcohol consumption, smoking and inadequate eating habits were more frequent among patients with peptic ulcers and gastric adenocarcinoma compared to those with gastritis (Table 1). Male patients and those older than 50 years predominated in the group with gastric adenocarcinoma (Table 1).

TABLE 1. Distribution of variables analyzed in patients with different gastrointestinal diseases 

VariableGastritis (n = 198) (%)Duodenal ulcer (n = 96) (%)Gastric ulcer (n = 78) (%)Gastric cancer (n = 70) (%)
Sex
 Male89 (45)60 (62.5)48 (62)50 (71)
 Female109 (55)36 (37.5)30 (38)20 (29)
P * nsns<0.05
Age
 >50 years60 (30)35 (36)30 (38)42 (60)
 <50 years138 (70)61 (64)48 (62)28 (40)
P * nsns<0.05
Alcohol consumption
 Yes109 (55)67 (70)65 (83)45 (64)
 No89 (45)29 (30)13 (17)25 (36)
P * <0.05<0.05<0.05
Smoking
 Yes81 (41)72 (75)55 (71)45 (64)
 No117 (59)24 (25)23 (29)25 (36)
P * <0.05<0.05<0.05
Diet
 Adequate83 (42)7 (7)2 (3)5 (7)
 Inadequate115 (58)89 (93)76 (97)65 (93)
P * -<0.05<0.05<0.05
H. pylori
 Positive168 (85)90 (94)76 (97)67 (96)
 Negative30 (15)6 (6)2 (3)3 (4)
P * -<0.05<0.05<0.05
CagA (H. pylori +)
 Positive117 (59)86 (89)69 (88)60 (86)
 Negative51 (26)4 (6)7 (9)7 (10)
P * -<0.05<0.05<0.05

G-test.

*

P value. ns: not significant

Investigation of the presence of H. pylori infection showed a higher prevalence of the bacterium and of CagA-positive strains in patients with gastric ulcers, duodenal ulcers and gastric adenocarcinoma when compared to patients with gastritis. However, no difference in prevalence was observed between patients with peptic ulcers and gastric adenocarcinoma (Table 1).

Table 2 shows the association between histopathological findings and lifestyle risk factors in the patients studied. Excessive and prolonged smoking and inadequate dietary habits were associated with a high degree of inflammation and intense neutrophilia. No association was observed with excessive alcohol consumption.

TABLE 2. Association of the degree of inflammation and neutrophil activity with risk factors analyzed in patients with gastric diseases 

Risk factorDegree of inflammationOR (95% CI)Neutrophil activityOR (95% CI)
LowHigh P valueLowHigh P value
Smoking
 Yes68 (27%)185 (73%)2.69 (1.80-4.00)72 (17%)181 (41%)2.82 (1.90-4.19)
 No95 (50%)94 (50%)0.0089 (20%)100 (22%)0.00
Alcohol consumption
 Yes
 No106 (37%)180 (63%)1.02 (0.68-1.53)99 (35%)187 (65%)0.76 (0.50-1.13)
57 (36%)99 (64%)0.9964 (41%)92 (59%)0.22
Diet
 Adequate59 (60%)38 (40%)3.59 (2.25-5.74)56 (57%)41 (43%)3.03 (1.91-4.82)
 Inadequate104 (30%)241 (70%)0.00107 (31%)238 (69%)0.00

OR: odds ratio, 95% CI: 95% confidence interval

The degree of inflammation and neutrophil activity was higher in patients infected with H. pylori than in uninfected subjects (Table 3). Similarly, patients infected with CagA positive strains presented a higher degree of inflammation and neutrophil activity than patients infected with CagA negative H. pylori strains (Table 3).

TABLE 3. Association of the degree of inflammation and neutrophil activity with the presence of Helicobacter pylori DNA and CagA in the group of patients with gastric diseases 

Risk factorDegree of inflammationOR (95% CI)Neutrophil activityOR (95% CI)
LowHigh P valueLowHigh P value
H. pylori
 Positive136 (34%)265 (66%)3.75 (1.90-7.40)138 (34%)263 (66%)2.42 (1.27-4.66)
 Negative27 (65%)14 (35%)0.0023 (56%)18 (44%)0.00
CagA
 Positive82 (25%)250 (75%)13.13 (6.80-25.23)85 (25%)247 (75%)10.46 (5.61-19.50)
 Negative56 (81%)13 (19%)0.0054 (78%)15 (22%)0.00

OR: odds ratio, 95% CI: 95% confidence interval

DISCUSSION

The clinical manifestations of H. pylori infection can vary from one region to another as a result of differences in the geographic distribution of H. pylori genotypes(26). CagA, a cytotoxin produced by H. pylori, is a marker of bacterial virulence and is associated with gastric disorders such as peptic ulcer and gastric carcinoma in western countries(20 22). A high incidence of gastric carcinoma and peptic ulcer, as well as a high prevalence of H. pylori infection, is observed in Brazil, particularly in the state of Pará(1 13 27).

The frequency of the cagA gene observed in the patients studied here was higher than that reported in other studies conducted in different Brazilian states(4 23). Furthermore, the results showed an association between the presence of CagA and the development of peptic ulcer and gastric carcinoma. This finding agrees with other studies carried out in Brazil(14 19 27) and in other countries(8 9 16). Noqueira et al.(15), studying Portuguese patients, found a significant association between the presence of the cagA gene and a higher degree of inflammation and neutrophil activity in gastric mucosa. In the present study, histopathological analysis also demonstrated an association between the presence of CagA and a high degree of inflammation and neutrophil activity in gastric mucosa.

The present study analyzed the presence of H. pylori infection and other risk factors in patients with gastroduodenal diseases. Smoking, alcohol consumption and inadequate dietary habits were the most frequent risk factors. Heavy and prolonged smoking and inadequate dietary habits were associated with a higher degree of inflammation and intense neutrophilia. However, no association was observed with excessive alcohol consumption. Epidemiological studies suggest that alcohol plays a fundamental role in the development of gastrointestinal diseases such as peptic ulcer and gastric cancer since it causes gastric mucosal injuries and stimulates parietal cells to produce gastric acid, in addition to its synergistic action with smoking(5 29). Inadequate dietary habits such as excess intake of salt, preservatives and smoked produced, as well as diets poor in vegetable and fruits, are also strongly associated with the development of gastric cancer(24 28).

Food diaries have been used in studies investigating the association between diet and cancer in different populations in order to obtain nutritional information. However, this tool is limited since it is based on the information provided by the interviewed person and on the skills of the interviewer(24). Careful analysis is therefore needed before attributing a carcinogenic or anticarcinogenic role to one or more dietary factors and studies on risk factors and cancer are potentially prone to errors. Nevertheless, despite the errors inherent to surveys of habitual food intake, these tools are important since they provide knowledge of dietary patterns and their temporal variation in different populations worldwide. In this respect, evidence indicates the negative influence of the modern western diet (high in fat and processed foods and low in fibers) on the development of various types of cancer in developed and developing countries(28).

The present results and those of other studies led us to conclude that infection with CagA-positive H. pylori strains, excessive alcohol consumption, smoking and inadequate eating habits increase the risk of developing peptic ulcer and gastric carcinoma.

REFERENCES

1. Araújo-Filho I, Brandão-Neto J, Pinheiro LA, Azevedo IM, Freire FH, Medeiros AC. Prevalence of Helicobacter pylori infection in advanced gastric carcinoma. Arq. Gastroenterol. 2006;43:288-92. [ Links ]

2. Ayres M, Ayres MJ, Ayres DL, Santos AS. Bioestat 5.0. Aplicações estatísticas nas áreas das ciências biológicas e médicas. Sociedade Civil Mamirauá MCT - CNPq, Belém, 2007:364p. [ Links ]

3. Batista dos Santos SE, Rodrigues JD, Ribeiro-dos-Santos AK, Zago MA. Differential contribution of indigenous men and women to the formation of an urban population in the Amazon region as revealed by mtDNA and Y-DNA. Am J Phys Anthropol. 1999;109:175-80. [ Links ]

4. Brito CA, Silva LM, Jucá N, Leal NC, de Souza W, Queiroz D, Cordeiro F, Silva NL. Prevalence of cagA and vacA genes in isolates from patients with Helicobacter pylori-associated gastroduodenal diseases in Recife, Pernambuco, Brazil. Mem Inst Oswaldo Cruz. 2003;98:817-21. [ Links ]

5. Bujanda L. The effects of alcohol consumption upon the gastrointestinal tract. Am J Gastroenterol. 2000;95:3374-82. [ Links ]

6. Dixon MF, Genta RM, Yardley JH, Correa P. Histological classification of gastritis and Helicobacter pylori infection: an agreement at last? The International Workshop on the Histopathology of Gastritis. Helicobacter. 1997:S17-24. [ Links ]

7. Dufour MC. What is moderate drinking? Defining “drinks” and drinking levels. Alcohol Res Health. 1999;23:5-14. [ Links ]

8. Figueiredo C, Van Doorn LJ, Nogueira C, Soares JM, Pinho C, Figueira P, Quint WG, Carneiro F. Helicobacter pylori genotypes are associated with clinical outcome in Portuguese patients and show a high prevalence of infections with multiple strains. Scand J Gastroenterol. 2001;36:128-35. [ Links ]

9. Gwack J, Shin A, Kim CS, Ko KP, Kim Y, Jun JK, Bae J, Park SK, Hong YC, Kang D, Chang SH, Shin HR, Yoo KY. CagA-producing Helicobacter pylori and increased risk of gastric cancer: a nested case–control study in Korea. Br J Cancer. 2006;95:639–64. [ Links ]

10. Hammar M, Tyszkiewicz T, Wadström T, O'Toole PW. Rapid detection of Helicobacter pylori in gastric biopsy material by polymerase chain reaction. J. Clin. Microbiol. 1992;30:54–8. [ Links ]

11. Higashi H, Tsutsumi R, Fujita A, Yamazaki S, Asaka M, Azuma T, Hatakeyama M. Biological activity of the Helicobacter pylori virulence factors CagA is determined by variation in the tyrosine phosphorylation sites. Proc Natl Acad Sci USA. 2002;99:14428-33. [ Links ]

12. Maity P, Biswas K, Roy S, Banerjee RK, Bandyopadhyay U. Smoking and the pathogenesis of gastroduodenal ulcer recent mechanistic update. Mol Cell Biochem. 2003;253:329-38. [ Links ]

13. Martins LC, Corvelo TCO, Oti HT, Barile KAS. [Seroprevalence of antibodies against Helicobacter pylori CagA antigen in patients with gastric ulcer in the North region of Brazil]. Rev Soc Bras Med Trop. 2002;35:307-10. [ Links ]

14. Martins LC, Corvelo TC, Demachki S, Araujo MT, Assumpcao MB, Vilar SC, Freitas FB, Barbosa HP, Fecury AA, do Amaral RK, Dos Santos SE. Clinical and pathological importance of vacA allele heterogeneity and cagA status in peptic ulcer disease in patients from North Brazil. Mem Inst Oswaldo Cruz. 2005;100:875-81. [ Links ]

15. Nogueira C, Figueredo C, Carneiro F, Gomes AT, Barreira R, Figueira P, Salgado C, Belo L, Peixoto A, Bravo JC, Bravo LE, Realpe JL, Plaisier AP, Quint WG, Ruiz B, Correa P, Van Dooorn LJ. Helicobacter pylori genotypes may determine gastric histopathology. Am. J Pathol. 2001;158:647-54. [ Links ]

16. Nomura AM, Lee J, Stemmermann GN, Nomura RY, Perez-Perez G.I, Blaser M.J. Helicobacter pylori CagA seropositivity and gastric carcinoma risk in a Japanese American population. J Infect Dis. 2002;186:1138-44. [ Links ]

17. Peleteiro B, Bastos J, Barros H, Lunet B. Systematic review of the prevalence of gastric intestinal metaplasia and its area-level association with smoking. Gac Sanit. 2008;22:236-47. [ Links ]

18. Resende ALS, Mattos IE, Koifman S. Dieta e câncer gástrico: aspectos históricos associados ao padrão de consumo alimentar no estado do Pará. Rev Nutr. 2006;19:511-9. [ Links ]

19. Ribeiro ML, Godoy AP, Benvengo YH, Mendonca S, Pedrazzoli J Jr. Clinical relevance of the cagA, vacA and iceA genotypes of Helicobacter pylori in Brazilian clinical isolates. FEMS Immunol Med Microbiol. 2003;36:181-5. [ Links ]

20. Shimoyama T, Crabtree JE. Bacterial factors and immune pathogenesis in Helicobacter pylori infection. Gut. 1998;43:S2-5. [ Links ]

21. Suerbaum S, Michetti P. Helicobacter pylori infection. N Engl J Med. 2002;347:1175-86. [ Links ]

22. Tham KT, Peek RM Jr, Atherton JC, Cover TL, Perez-Perez GI, Shyr Y, Blaser MJ. Helicobacter pylori genotypes, host factors, and gastric mucosal histopathology in peptic ulcer disease. Hum Pathol. 2001;32:264-73. [ Links ]

23. Thomazini CM, Pinheiro NA, Pardini MI, Naresse LE, Rodrigues MAM. Helicobacter pylori and gastric cancer: distribution of cagA and vacA genotypes in patients with gastric carcinoma. J Bras Patol Med Lab. 2006;42:25-30. [ Links ]

24. Tsugane S. Salt, salted food intake, and risk of gastric cancer: epidemiologic evidence. Cancer Sci. 2005;96:1-6. [ Links ]

25. Tummuru MK, Cover TL, Blaser MJ. Cloning and expression of a high-molecular-mass major antigen of Helicobacter pylori: evidence of linkage to cytotoxin production. Infect Immun. 1993;61:1799–809. [ Links ]

26. Van Doorn LJ, Figueiredo C, Mégraud F, Pena S, Midolo P, Queiroz DM, Carneiro F, Vanderborght B, Pegado MD, Sanna R, De Boer W, Schneeberger PM, Correa P, Ng EK, Atherton J, Blaser MJ, Quint WG. Geographic distribution of vacA allelic types of Helicobacter pylori. Gastroenterology.1999;116:823-30. [ Links ]

27. Vinagre RM, Corvelo TC, Arnaud VC, Leite AC, Martins LC. Determinação das cepas do Helicobacter pylori e do polimorfismo do gene da interleucina-8 em pacientes com câncer gástrico. Arq. Gastroenterol. 2011;48:46-51. [ Links ]

28. Vinagre RM, Campos BP, Sousa RM. Case study of stomach adenocarcinoma conducted at a cancer referral hospital in northern Brazil. Arq. Gastroenterol. 2012;49:125-29. [ Links ]

29. Wu WK, Cho CH. The pharmacological actions of nicotine on the gastrointestinal tract. J Pharmacol Sci. 2004;94:348-8. [ Links ]

Received: December19, , 2012; Accepted: April24, , 2013

Correspondence: Luisa Caricio Martins. Trav. Mauriti, 3269, ap. 704 B, Bairro: Marco - 66095-360 – Belém, PA, Brasil. E-mail: luisacaricio@uol.com.br

Declared conflict of interest of all authors: none

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.