Perinatal outcomes resulting from the management of gestations with restricted intrauterine growth using different protocols

Laís Nascimento de Melo Silva1, Érika Maria Alves da Silva2, Manoella Mirella da Silva Vieira Araújo3, Gracielly Karine Tavares Souza4, Auricarla Gonçalves de Souza5, Bárbara Regina Britto de Oliveira Vieira6, Marcelo Marques de Souza Lima 7, Gilvânia Patrícia do Nascimento Paixão8

1-6Federal University of Pernambuco. Pernambuco (PE), Brazil.

7Dom Malan Hospital. Petrolina (PE), Brazil.

8State University of Bahia. Salvador (BA), Brazil.

Introduction

Intrauterine Growth Restriction (IUGR) is one of the complications of pregnancy that classifies it as high risk and affects about 5 to 10% of gestations. It is considered an important cause of mortality and of negative perinatal outcomes that requires adequate monitoring in order to avoid adverse outcomes for the mother-fetus binomial.1-2 In 2018, Brazil estimated a prevalence of 9.2% of IUGR, and about 18% of babies born in the country were born 'small' (< 2500 g). In relation to low birth weight, the prevalence was 0,7% in Latin America, 1,6% in Africa, and 2,3% in Asia. These children are 62 times more likely to die than those with no such characteristics.3-4   

IUGR is characterized by gestation with a fetus framed in the lower than the 10th  percentile, excluding Small for Gestational Age (SGA) fetuses that present a lower risk of unfavorable perinatal outcomes. Two classifications are included: moderate IUGR, with the fetus between the 3rd and the 10th percentiles; and severe IUGR, for those below the 3rd percentile. There is also early IUGR, when the fetus is < 32 weeks of gestational age, and late IUGR, with > 32 weeks.5-6

However, there is no consensus in the literature on the diagnostic criteria for this condition, creating uncertainty regarding management and the ideal timing for shortening birth. Ideally, the goal is to ensure that pregnancy continues until the fetus achieves sufficient lung maturity, avoiding irreversible fetal damage; however, the optimal timing for such measures remains a major challenge.2,6

In addition, calculating estimated fetal weight is an extremely important parameter in assessing growth restriction. The most commonly used formula in clinical practice for this purpose is Hadlock's formula.7 However, it should be noted that pregnant women with maternal diseases that could interfere with fetal growth, as well as twin pregnancies, were excluded from the development of this formula.8-10

The most viable intervention for most cases of IUGR is early delivery, even when there is uncertainty about the ideal timing, since delay may increase the chances of hypoxia and fetal neurological damage, but also considering that early delivery carries the risks of prematurity.2,6 In most cases, prematurity ends up being determined by the need to rescue the fetus from unfavorable intrauterine conditions, considering that technological and care advances in neonatal intensive care have resulted in better prognoses.5,11

In 2017, at the hospital where the on-screen study was conducted, the IUGR management protocol was performed with monitoring of Doppler velocimetry changes, cardiotocography, and fetal biophysical profile to assess fetoplacental circulation and predict the degree of compromise. The schedule for termination of pregnancy was up to 34 weeks. In the presence of zero diastole in the umbilical artery, corticosteroid therapy was expected to be completed before termination; reverse diastole would indicate immediate termination.12

In 2018, changes were made to the protocol in order to improve care. In 2019, the new protocol was established, being currently in force. In it, obstetric ultrasound scans are performed every two weeks, Doppler ultrasound scans and fetal biophysical profiles are performed weekly, and, according to the changes, it is determined whether it is possible to wait until 37 weeks for delivery. In the presence of zero diastole in the umbilical artery, termination is scheduled for the 34th week, while with reverse diastole, termination is indicated from 30 weeks onwards.13

Given that care for the binomial with IUGR is complex and involves several decisions that influence fetal prognosis, it is necessary to evaluate the outcomes of the protocols established in order to understand which one presents the best results. Thus, the objective of this study is to describe the perinatal outcomes using different protocols for managing gestations with intrauterine growth restriction in a maternal and child referral hospital.

Method

This is a cross-sectional, descriptive study with a quantitative approach, conducted in a high-risk obstetric hospital located in Petrolina, Pernambuco (Brazil). This unit provides care to the population of the Pernambuco/Bahia network (PEBA Network), corresponding to 53 municipalities. The service is the only referral center for high-complexity obstetric and neonatal care in its Integrated Region of Economic Development (RIDE - acronym in Portuguese).

A documentary analysis of medical records of patients with confirmed diagnosis of IUGR admitted to the High Risk Clinic was conducted between April and September 2021.

The inclusion criteria were medical records of pregnant women with gestational age between 21 and 41 weeks and six days, admitted in 2017 and 2019, with a minimum hospital stay of five days. Women with multiple pregnancies, congenital malformations, uterine anatomical anomalies, and daily use of corticosteroids were excluded.

Data were collected using a semi-structured instrument containing the following clinical and obstetric variables: number of pregnancies, number of medical appointments, associated clinical conditions, previous comorbidities, gestational age at admission, gestational age at corticosteroid use, gestational age at termination, and indication for termination of pregnancy.

The neonatal variables included were weight at birth, Apgar score, neonatal mortality, neonatal diagnosis, length of hospital stay, devices used during hospitalization, and admission to the Intensive Care Unit/Neonatal Intensive Care Unit (ICU/NICU).

The data were entered into an Excel spreadsheet and analyzed using the Statistical Package for Social Sciences (SPSS 20) program through descriptive statistics, with calculations of absolute and relative frequencies, and measures of central tendency and dispersion.

The study, conducted in accordance with Resolution No. 466/12 of the National Health Council, complied with the Declaration of Helsinki (1964, revised in 1975, 1983, 1989, 1996, and 2000) on research involving human subjects, and was submitted to and approved by the Human Research Ethics Committee of the Prof. Fernando Figueira Institute of Integral Medicine, CEP-IMIP, under CAAE 27394619.1.0000.5201.

Results

114 patients diagnosed with IUGR pregnancy who met the proposed eligibility criteria were identified, with 36 in 2017 and 78 in 2019. Regarding gestational characteristics, it was observed that in both years the majority were multigravida (61,1% in 2017, 66,7% in 2019), were between 21 and 32 weeks of gestation at admission (66,7% in 2017, 59% in 2019), and had a diagnosis of IUGR at ≤ 32 weeks of gestation (58,3% in 2017, 53,8% in 2019). In 2019, patients had more prenatal appointments (60,3% had ≥ 6) (Table 1).

Regarding fetal percentile in 2017, most were in the < 3rd percentile (41,7%), followed by fetuses in the < 3rd percentile with Doppler velocimetry abnormalities (27,8%). In 2019, more than half were in the < 3rd percentile with Doppler velocimetry abnormalities (55,1%). The most frequent period of pregnancy interruption was between 28 and 33 weeks and 6 days in 2017 (41,7%) and ≥ 37 weeks in 2019 (38,5%). Data on a history of prematurity and IUGR were not available in the medical records (Table 1).

Table 1 - Frequency of gestational characteristics in women with IUGR in 2017 and 2019. Petrolina (PE), Brazil, 2021.

Variables

2017

2019

 

N=36

%

N=78

%

Number of pregnancies

 

 

 

 

Primigravida

14

38,9%

26

33,3%

Multigravida

22

61,1%

52

66,7%

Gestational age at admission

 

 

 

 

21 weeks to 32 weeks

24

66,7%

46

59,0%

32 weeks and 1 day to 37 weeks

12

33,3%

32

41,0%

Number of appointments

 

 

 

 

≤ 5 appointments

18

50,0%

31

39,7%

≥ 6 appointments

18

50,0%

47

60,3%

Gestational age suspected of IUGR

 

 

 

 

≤ 32 weeks

21

58,3%

42

53,8%

> 32 weeks and 1 day

15

41,7%

36

46,2%

Fetal percentile

 

 

 

 

< 3rd Percentile

15

41,7%

23

29,5%

< 3rd Percentile + Doppler velocimetry alteration

10

27,8%

43

55,1%

< 10th Percentile

6

16,7%

0

0%

< 10th Percentile + Doppler velocimetry alteration

5

13,9%

12

15,4%

Gestational age at termination of pregnancy

 

 

 

 

< 28 weeks

2

  5,6%

2

  2,6%

28 to 33 weeks and 6 days

15

41,7%

19

24,4%

34 to 36 weeks and 6 days

9

25,0%

27

34,6%

≥ 37 weeks

10

27,8%

30

38,5%

Table 2 shows maternal diagnoses during pregnancy with IUGR and indications for termination of pregnancy. Among the most frequent in the follow-up with both protocols, there was a significant presence of preeclampsia, 63,3% in 2017 and 70,5% in 2019, followed by urinary tract infection (46,2% and 33,3%, respectively) and chronic hypertension (36,1% and 43,6%, respectively). The main reasons for termination of pregnancy were alterations in Doppler velocimetry or fetal centralization (47,2% in 2017 and 25,6% in 2019) and acute fetal distress (41,7% in 2017 and 19,2% in 2019).

Table 2 - Frequency of diagnoses in pregnant women with fetuses with IUGR and indications for termination of pregnancy in 2017 and 2019. Petrolina (PE), Brazil, 2021.

Variables

2017

2019

N=36

%

N=78

%

Gestational diagnoses

 

 

 

 

Preeclampsia

23

63,9%

55

70,5%

Chronic arterial hypertension/Hypertension

13

36,1%

34

43,6%

Diabetes mellitus/ Gestational diabetes mellitus

5

13,9%

12

15,4%

Thrombocytopenia/ Bleeding during pregnancy

9

25,0%

11

14,1%

Premature rupture of the ovular membranes

4

11,1%

5

6,4%

Vaginitis

10

27,8%

16

20,5%

Urinary tract infection

17

47,2%

26

33,3%

Smoking or alcoholism

3

8,3%

5

6,4%

Obstetric malpractice

5

13,9%

6

7,7%

Autoimmune disease

1

2,8%

6

7,7%

History of abortion or fetal death or stillbirth

9

25,0%

25

32,1%

Premature labor

4

11,1%

8

10,3%

Indication for termination of pregnancy

 

 

 

 

Pelvic

1

2,8%

4

5,1%

Blood pressure instability

7

19,4%

11

14,1%

Acute fetal distress

15

41,7%

15

19,2%

Doppler alteration or centralization

17

47,2%

20

25,6%

Premature labor

3

8,3%

10

12,8%

Preeclampsia

12

33,3%

2

2,6%

Alterations in amniotic fluid

6

16,7%

0

0%

Thrombocytopenia or gestational bleeding

3

8,3%

3

3,8%

Diabetes Mellitus

1

2,8%

0

0%

Full-term

3

8,3%

29

37,2%

Note: The data in the table are cumulative, allowing the same patient to have more than one diagnosis.

With regard to neonatal variables, the 2017 protocol observed a prevalence of newborns (NBs) weighing less than 2.500 g (41,7%); an Apgar score < 7 in the first minute (55,6%) and > 7 in the fifth minute (94,4%); and admission to the ICU/ITU (69,4%), with the vast majority not progressing to death (83,3%). In 2019, the main difference was that more than half were born with Apgar scores > 7 in the first minute and fifth minute, with a higher expression in the fifth minute (60,3% and 88,5%, respectively) (Table 3).

The most frequent neonatal complications were transient tachypnea of the newborn in both years (69,4% in 2017 and 55,1% in 2019), followed by jaundice (55,6% in 2017 and 43,6% in 2019) and healthcare-associated infection (30,6% and 19,2%, respectively). Newborns who required continuous positive airway pressure (CPAP) or mechanical ventilation with venous access predominated in 2017 (38,9%); in 2019, most newborns did not require any device (37,2%) (Table 3).

Table 3 - Characterization of newborns with Intrauterine Growth Restriction in 2017 and 2019. Petrolina (PE), Brazil, 2021.

Variables

2017

2019

 

N=36

%

N=78

%

Weight at birth

 

 

 

 

< 1000 g

6

16,7%

5

6,4%

< 1500 g

12

33,3%

21

26,9%

< 2500 g

15

41,7%

48

61,5%

≥ 2500 g

3

8,3%

4

5,1%

Apgar first minute

 

 

 

 

< 7

22

55,6%

31

39,8%

≥ 7

16

44,4%

47

60,3%

Apgar fifth minute

 

 

 

 

< 7

2

5,6%

9

11,6%

≥ 7

34

94,4%

69

88,5%

Admission to neonatal ITU/ICU

 

 

 

 

Yes

25

69,4%

45

57,7%

No

11

30,6%

32

42,3%

Death

 

 

 

 

No

30

83,3%

69

88,5%

Intrauterine

1

2,8%

2

2,6%

Neonatal

5

13,9%

7

9,0%

Neonatal complications*

 

 

 

 

TTN

25

69,4%

43

55,1%

Hypoglycemia

8

22,2%

15

19,2%

Jaundice

20

55,6%

34

43,6%

HAIs

11

30,6%

15

19,2%

Intraventricular hemorrhage

4

11,1%

2

2,6%

Apnea of prematurity

3

8,3%

3

3,8%

Anemia of prematurity

5

13,9%

6

7,7%

Necrotizing enterocolitis

0

0,0%

2

2,6%

Cardiac arrest / death

6

16,7%

8

10,3%

Neonatal devices

 

 

 

 

None

7

19,4%

29

37,2%

CPAP or MV

1

2,8%

1

1,3%

Venous access

0

0,0%

3

3,8%

Phototherapy

2

5,6%

3

3,8%

CPAP or MV + Venous access

14

38,9%

15

19,2%

Venous access + Phototherapy

1

2,8%

2

2,6%

CPAP or MV + Phototherapy

1

2,8%

2

2,6%

All devices

10

27,8%

23

29,5%

*The same newborn may present more than one complication. Captions: CPAP = continuous positive airway pressure; VM = mechanical ventilation; HAIs = healthcare-associated infections; TTN = transient tachypnea of the newborn.

In 2017, most newborns who underwent corticosteroid therapy for lung maturation were between 24 and 34 weeks GA, weighed less than 2.500 g (70,9%), had an Apgar score < 7 in the first minute (45,1%) and ≥ 7 in the fifth minute (70,9%), and were in the < 3rd percentile (54,9%). In 2019, the data differed in terms of Apgar scores in the first minute, when most scored ≥ 7. Most NBs admitted to the neonatal ICU/ITU weighed less than 2.500 g in 2017 (68,5%) and 2019 (56,5%), being in the < 3rd percentile in 2017 (51,4%) and 2019 (50%). The minority progressed to neonatal death in both years, with a higher prevalence in 2017 among those weighing less than 2.500 g (14,2%), Apgar ≥ 7 at the fifth minute of life (11,4%); and with percentile < 3 (11,4%), while in 2019 the majority of deaths occurred among the NBs weighing less than 2.500 g (9,21%) and percentile < 3 (7,89%) (Table 4).

Table 4 - Description of perinatal data of newborns with IUGR regarding gestational age at corticosteroid use, neonatal death, admission to neonatal ICU/ITU, weight at birth, Apgar score, and fetal percentile in 2017 (n=36) and 2019 (n=78). Petrolina (PE), Brazil, 2021.

 

Corticosteroid GA**

Neonatal Death

AdmissIon to ITU/ICU

 

2017

2019

2017

2019

2017

2019

Weight at birth

24-34 w

34,1-37

24-34 w

34,1-37

Yes

No

Yes

No

Yes

No

Yes

No

< 2.500 g

22

(70,9%)

1

22,5%

48

(64%)

23

(30,6%)

5

14,2%)

27

(77,1%)

7

(9,21%)

65

(85,5%)

24

(68,5%)

8

(22,8%)

43

(56,5%)

29

(38,1%)

≥ 2.500 g

1

(3,2%)

1

(3,2%)

0

4

(5,33%)

0

3

(8,57%)

0

4

(5,26%)

1

(2,86%)

2

(5,71%)

1

(1,32%)

3

(3,96%)

Apgar 1st minute

 

 

 

 

 

 

 

 

 

 

 

 

< 7

14

(45,1%)

3

(9,68%)

19

(25,3%)

10

13,3%)

3

(8,57%)

16

(45,7%)

5

(6,58%)

24

(31,5%)

17

(48,5%)

2

(5,71%)

24

(31,5%)

5

(6,58%)

≥ 7

9

(29,0%)

5

(16,1%)

29

(38,6%)

17

(22,6%)

2

(5,71%)

14

(40%)

2

(2,63%)

45

(59,2%)

8

(22,8%)

8

(22,8%)

20

(26,3%)

27

(35,5%)

Apgar 2nd minute

 

 

 

 

 

 

 

 

 

 

 

 

< 7

1

(3,23%)

0

4

(5,33%)

3

(4%)

1

(2,86%)

0

2

(2,63%)

5

(6,5%)

1

(2,86%)

0

5

(6,58%)

2

(2,63%)

≥ 7

22

(70,9%)

8

(25,8%)

44

(58,6%)

24

(32%)

4

(11,4%)

30

(85,7%)

5

(5,68%)

64

(84,2%)

24

(68,5%)

10

(28,5%)

39

(51,32%)

30

(39,4%)

Percentile

 

 

 

 

 

 

 

 

 

 

 

 

< 3*

17

(54,8%)

5

(16,1%)

41

(54,6%)

22

(29,3%)

4

(11,4%)

20

(57,1%)

6

(7,89%)

58

(76,3%)

18

(51,4%)

6

(17,1%)

38

(50%)

26

(34,2%)

<10*

6

(19,3%)

3

(9,68%)

7

(9,33%)

5

(6,67%)

1

(2,86%)

10

(28,5%)

1

(1,32%)

11

(14,4%)

7

(20%)

4

(11,4%)

6

(7,89%)

6

(7,89%)

* With or without Doppler velocimetry alteration.

** Gestational age at corticosteroid use, stated in weeks.

Discussion

The Ministry of Health (MH) recommends at least six prenatal medical appointments for standard-risk pregnancies; in cases of reassessment as gestational risk, this number is increased and the interval between appointments adjusted.12 Among the objectives of prenatal care is the identification of maternal and fetal health problems and, in the case of the subject under study, the early identification of fetuses with IUGR, so that interventions can be made to reduce fetal and neonatal morbidity and mortality.14-15 In the on-screen study, it was observed that in 2019 the percentage of women with more than six appointments was higher when compared to 2017. Some studies present data that show a higher frequency of low weight at birth, as well as other complications with women who had fewer than six prenatal appointments.16-17

There is generally a relationship between early IUGR diagnosis and greater fetal and neonatal complications associated with placental insufficiency and chronic fetal hypoxia, best verified with the aid of Doppler velocimetry.5,18-19 In the study in question, there was a higher prevalence of fetuses with a diagnosis of early IUGR who had lower Apgar scores at the first minute, although with a lower percentage in 2019. This finding may be related to the higher frequency of screening tests in the new protocol introduced at the hospital in 2019. These data are consistent with a study conducted in Recife-PE, which showed higher complications of early IUGR, as well as higher morbidity and mortality rates.20

Fetuses with percentiles < 3 also prevailed, but in 2019, the highest number was of those with Doppler velocimetry alterations. There is still no consensus on the diagnosis of IUGR; however, these are classified as severe IUGR because they are below the 3rd percentile.10 When the percentile is associated with abnormal Doppler in maternal or fetal vessels due to progressive destruction of the villous vasculature, follow-up tests need to be intensified to reduce perinatal mortality.15

Regarding the change in protocol, there was an increase in the gestational age at termination of pregnancy in 2019, with most reaching full term in comparison with 2017. One of the objectives of care required for IUGR pregnancies is to avoid prematurity and associated complications, allowing the pregnancy to continue for lung maturation while seeking to avoid irreversible fetal damage.2,6

In the two studied years, similar results related to preeclampsia were found in more than half of the sample, followed by other hypertensive disorders. The pathophysiology of preeclampsia, in particular, establishes a relationship with abnormal decidualization and vascularization of uterine cells, which is essential for the implantation and growth of the fetal allograft. These placental alterations reduce uteroplacental perfusion, leading to consequences such as IUGR and low weight at birth.22-22 A study showed that NBs of mothers with blood pressure alterations had higher perinatal mortality, prematurity, were SGA, and presented IUGR.22

Adaptive blood supply responses are essential for maintaining cerebral oxygenation in the presence of chronic hypoxia, and are mainly identified by alterations in Doppler velocimetry. In the presence of fetal compromise with a risk of neurological damage, termination of pregnancy is indicated, weighing the risks and benefits.1 It was observed that, among the diagnoses used as justification for pregnancy termination in both years evaluated in this study, altered Doppler velocimetry or centralization and acute fetal distress were the most common (with higher frequency in 2017).

Fetal weight is an essential parameter in ultrasound scan assessment, as it is one of the determining factors in the decision to continue or terminate high-risk pregnancies.8-9 Low birth weight NBs are at greater risk of death compared to those with adequate weight, which is more significant when they are born with extremely low birth weight.16 Neonatal death in this study was more prevalent in low birth weight newborns and in the < 3rd percentile, slightly higher in 2017. It is known that, among other factors related to intrauterine injuries suffered by the fetus, neonatal deaths are also related to the conditions of care in the peripartum period.23 Knowing the needs of fetuses with IUGR, priority should be given to the quality of care throughout the prepartum, delivery, and neonatal periods.

An important factor at birth, including for NBs with IUGR, is the Apgar score. Scores below 7 at five minutes of life are associated with neonatal asphyxia, and outcomes worsen when associated with low birth weight.24 Nevertheless, the present study found that most newborns weighing less than 2.500 g at birth had Apgar scores higher than 7 at five minutes of life in both years. Identifying low birth weight newborns and poor vitality at birth, including related risk factors, facilitates planning interventions that can improve care and reduce the chances of neonatal mortality.25-26

Due to the need for continuous surveillance and monitoring, these newborns must be admitted to a neonatal ITU/ICU, usually because they are babies with hypoglycemia and respiratory distress.10 Fetuses with low percentiles have longer stays in the neonatal ITU/ICU when compared to fetuses with the appropriate percentile. No significant difference was observed among NBs admitted to neonatal ITU/ICU in 2017 or 2019. However, in 2017 there was a higher number of NBs admitted to neonatal ITU/ICU with low birth weight and a percentile below 3.

The risk of morbidity and mortality is increased in infants with IUGR due to compromised growth and low energy reserves, leaving them more vulnerable.27 Perinatal mortality increases as growth restriction becomes more severe, increasing considerably when there is low weight and especially extreme low weight at birth.10 Babies with restriction have higher rates of need for mechanical ventilation and respiratory distress when compared to healthy babies.19 Transient Tachypnea of the Newborn was also prevalent in the present study, especially in 2017.

The 2019 protocol showed better results in terms of the number of NBs who did not require any type of neonatal device, whereas in 2017 the use of CPAP or MV with venous access prevailed. A study shows that chronic hypoxia in IUGR hinders normal lung development and increases the chance of respiratory impairment; even babies with restriction born at full term have worse respiratory outcomes than babies without restriction.27 Prenatal corticosteroid therapy reduces the incidence of respiratory distress syndrome; however, in this study, there was no difference in respiratory distress in either year.28-29

In summary, it was observed that the implementation of the new IUGR management protocol in 2019 at the institution under study led to improvements in important (albeit modest) aspects of the NB. The main findings show that with the 2019 protocol, Apgar scores and gestational age at termination of pregnancy were higher, and there was less need for ventilatory support for the NB.

Thus, it can be assumed that there is a positive superiority in the protocol established in 2019, according to which it is possible to wait up to 37 weeks of gestation for delivery depending on ultrasound scan characteristics, on Doppler, on the presence of zero diastole in the umbilical artery, and on reverse diastole. The combined assessment with Doppler parameters, as well as their correlation, provides more reliable information on the risk of adverse perinatal outcomes.

It is suggested that characteristics of the ultrasound scan test such as fetal percentile, abdominal circumference, and Doppler velocimetry of maternal and fetal vessels correlated with neonatal outcomes be explored in another article. The limitation of this study refers to the method of analysis, which does not allow for the indication of cause and effect. Thus, analytical/associative studies are essential to complement this gap.

Finally, this study provides insight into a field of practice focused on neonatal and obstetric care, from admission to high-risk units, health-related infection control and intensive care, as well as the importance of implementing and discussing evidence-based protocols with multidisciplinary healthcare teams.

Conclusion

Discrete but positive differences were observed during the years of the research investigation. The most unfavorable neonatal outcomes were identified in 2017 with regard to the NB weighing < 2,500 g, percentile < 3, Apgar < 7, admission to the neonatal ICU/ITU, and neonatal death. Therefore, it can be assumed that the implementation of the new 2019 protocol has led to outcomes that are more favorable to the NB.

The results have the potential to contribute to clinical practice and scientific advancement with regard to institutional protocol changes and their impact on newborns considering IUGR. The data demonstrate that monitoring new evidence and updated studies, as well as incorporating them into these protocols allow for better outcomes in IUGR conditions, in this case increasing the gestational age for termination.

Further studies are required, with in-depth analysis to understand the real impact and relationship of each variable in the different protocols.

 Authors’ contributions

Study design: Laís Nascimento de Melo Silva. Data collection: Laís Nascimento de Melo Silva. Data analysis and interpretation: Laís Nascimento de Melo Silva, Érika Maria Alves da Silva, Manoella Mirella da Silva Vieira Araujo, Gracielly Karine Tavares Souza, Gilvânia Patrícia do Nascimento Paixão, Marcelo Marques de Souza Lima, Bárbara Regina Britto de Oliveira Vieira, Auricarla Gonçalves de Souza. Manuscript writing: Laís Nascimento de Melo Silva, Érika Maria Alves da Silva, Manoella Mirella da Silva Vieira Araujo, Gracielly Karine Tavares Souza, Gilvânia Patrícia do Nascimento Paixão, Marcelo Marques de Souza Lima, Bárbara Regina Britto de Oliveira Vieira, Auricarla Gonçalves de Souza. Manuscript critical revision:  Érika Maria Alves da Silva, Manoella Mirella da Silva Vieira Araujo, Gracielly Karine Tavares Souza, Gilvânia Patrícia do Nascimento Paixão, Marcelo Marques de Souza Lima, Bárbara Regina Britto de Oliveira Vieira, Auricarla Gonçalves de Souza. Approval of the final version of the text: Érika Maria Alves da Silva, Manoella Mirella da Silva Vieira Araujo, Gracielly Karine Tavares Souza, Gilvânia Patrícia do Nascimento Paixão, Marcelo Marques de Souza Lima, Bárbara Regina Britto de Oliveira Vieira, Auricarla Gonçalves de Souza.

 Conflict of interest

The authors have declared that there is no conflict of interest.

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