Efficacy and safety of combined oral sucrose and nonnutritive sucking in pain management for infants: A systematic review and meta-analysis.

BACKGROUND: Pain management is currently important in neonatal intensive care unit (NICU). The superiority in pain relief of the combined oral sucrose (OS) and nonnutritive sucking (NNS) to other single intervention has not been well established. The administration of sucrose has been considered to potentially induce adverse events, which has been controversial. This study aims to investigate the combined effects and safety in comparison with other single intervention methods, including NNS, OS alone, breast milk and oral glucose.
METHODS: We searched databases including Medline (via Pubmed), Embase (via Ovid), web of science, and Cochrane Library for randomized controlled trials from Jan 1, 2000 to Mar 31, 2021. The data were analyzed in the meta-analysis using Review manager Version 5.3. Pain score was the primary outcome in this meta-analysis. The adverse events were assessed qualitatively.
RESULTS: A total of 16 studies were eligible in the meta-analysis. The results demonstrated a significant reduction in pain score in the NNS+OS group compared with NNS alone (SMD = -1.69, 95%CI, -1.69,-0.65) or sucrose alone (SMD = -1.39, 95% CI, -2.21,-0.57) during the painful procedures. When compared NNS+OS with breast milk, no significant difference was detected (SMD = -0.19, 95% CI: -0.5, 0.11).
CONCLUSION: The combined effects of NNS and OS might be superior to other single intervention method. However, the effects might be mild for moderate-to-severe pain.

1 Introduction

Newborns, especially preterm infants, are frequently subjected to painful procedures [1], repeated painful and stressful stimuli may develop clinical, physiologic and psychologic sequelae in the short or long term [2-4]. In some developing countries, most infants were not provided any analgesic intervention during painful procedures [5, 6]. Even in a developed country like Canada, analgesic interventions were not offered in almost half of painful procedures, according to an epidemiology study [7]. In addition, it remains unclear whether the administration of anesthetics is safe [8]; thus, selective use of pharmacological treatment is recommended, such as opioids, non-opioids, and other anesthetics [9]. On the other hand, nonpharmacological methods have been developed in recent years to help reduce multiple bedside interruptions and have been considered safe and effective in pain management [10, 11]. An updated research revealed the use rate of nonpharmacologic interventions tended to be higher than pharmacologic interventions [12].

Combined therapy was assumed to be more efficient in pain relief than single interventions and has been investigated frequently in recent studies [13-16]. For example, combined effects of music and touch [17], sucking, breast milk and tucking [18], music and sucrose [19], oral sucrose (OS) and nonnutritive sucking (NNS) [20, 21], etc. were evaluated in clinical trials. Among which the combined effects of NNS and sucrose were tested by numerous prospective randomized trials. It has been common knowledge that OS and NNS could separately alleviated pain in neonates. Previous reviews or meta-analyses have preliminarily evaluated the combined effects of NNS and sucrose [10, 13]. However, they may have neglected the variations in effectiveness and safety under different painful procedures. The superiority of the combination to various single interventions was also barely covered [8]. Therefore, this systematic review and meta-analysis aim to fill the gap by evaluating the efficacy of combining OS and NNS in different scenarios of comparison and exploring the safety of this intervention method preliminarily.

2 Methods

2.1 Literature search and screen

We conducted a systematic search from the following databases for English language articles: Medline (via PubMed), Embase (via Ovid), Cochrane Library, Web of Science. The search terms were based on three domains “newborns”, “sucrose and non-nutritive sucking” and “procedural pain”. The outcome-related terms were not restricted. Randomized controlled trials were selected using database-specific limiters. The time span of publication years was limited from Jan 1, 2000 to Mar 31, 2021. The search strategies for each database were provided (S1 Table). In addition to the electronic searches, Google Scholar and references in literature were also manually searched for potential suitable studies. The unpublished studies were not considered. The language restriction was English.

The literature research and screening procedures afterwards were performed independently by two researchers. The final eligible studies were cross-checked. If there were any disagreements, a third author made judgments.

2.2 Inclusion and exclusion criteria

2.2.1 Inclusion criteria

(1) The study was a randomized controlled trial. (2) The paticipants were preterm or full term neonates without severe illness. (3) The randomized groups in the trial should at least contain an intervention group applied with OS combined with NNS. (4) Conference abstracts eligible for the inclusion criteria above were also included.

2.2.2 Exclusion criteria

(1) Results of pain score were presented as categorical outcome. (2) The sample size, mean and standard deviation were partly provided and could not be estimated from other statistics such as mean difference and p-value or median, range and interquartile range. (3) Non-English literature was excluded. (4) The outcomes of no interest, e.g., the occurrence of startle, jerk or tremor were excluded.

2.3 Literature quality evaluation

We conducted literature quality evaluation according to the Cochrane Risk of Bias Tool (RoB), version 5.2 [22]. The risk of bias was assessed on a 3 level scale:”low”, “high” and “unclear” risk based on the following seven domains: random sequence generation, allocation concealment, blinding of participants and personnel, blinding of outcome assessment, incomplete outcome data, selective reporting, intention-to-treat analysis, and a completeness of follow-up.

2.4 Data extraction

Data extraction was performed by two reviewers independently. The summary of studies was established from the following information: first author, year of publication, country, study design, gestational age, pain procedure, sample size and specific intervention descriptions in each group and outcome.

We implemented the meta-analysis on pain score measured in different painful procedures, including Premature Infant Pain Profile(PIPP) [23], the premature infant pain profile-revised (PIPP-R) [24], Neonatal Infant Pain Scale (NIPS) [25], the Neonatal Facial Coding System(NFCS) [26] Neonatal Pain Agitation and Sedation Scale (N-PASS) [27]. The results of pain score were required to be presented as continuous variables. The Means and standard deviations (SDs) were extracted directly from the original data if available. Otherwise, they were estimated using Hozo et al.’s Method or Bland’s Method under the scenario where median, IQR or ranges were available [28-30]. Calculations using p-value, confidence interval and mean difference were performed following the instructions in Cochrane handbook 5.1 [22]. Statistics were estimated from figures if applicable when no digits were provided. Missing data were not imputed. For cross-over designs trials, paired analysis was performed to standardize the means and SDs to account for the within-subject correlation [31]. If no information about within-subject correlation was provided, we decided to assume the correlation to be 0 which was a conservative way since there were no similar data to be referred in the included studies.

For evaluating the effects at different timepoints, we extracted the outcome data in 2 phases, (1) during the procedure phase (during or within the first minute immediately after the painful procedure), and (2) the recovery phase (1 to 5 min after the procedure). Pairwise comparisons were performed between the intervention group of interest, i.e., sucrose and NNS group and any other relevant groups including NNS (NNS+water)/sucrose alone, breastfeeding, breast milk, glucose or routine care group. In crossover trials, the overall effects were extracted instead of looking at a single sequence.

2.5 Statistical analysis

Results of pain score were presented as standardized mean difference (SMD) if holding different scales and 95% CI considering variety among pain measurements. If studies shared the same measurement then mean difference (MD) could be presented. A p-value <0.05 was considered as significant. Statistical heterogeneity was assessed using I2 tests. Subgroup analysis or random-effects model applied when the statistical heterogeneity was high (I2 >50%) [22]. Otherwise, a fixed-effects model was conducted. Sensitivity analyses were conducted to assess the stability and validity of results. Factors were considered when removing certain studies, including article types, outcome measurements (PIPP or non-PIPP), target population (preterm/full term), etc. Publication bias was evaluated by Egger’s Test and funnel plots. All the statistical analysis was performed using Review Manager 5.3 [32]. Egger’s Test was performed using SAS version 9.4. Besides, the safety of the intervention was qualitatively evaluated based on the occurrence and types of adverse events.

3 Results

3.1 Outline of eligible studies

A total of 232 records were identified using our searching strategy from databases and two from the references in a previous review [10]. After removing duplicates, 79 records were obtained and 63 of them were screened in the first stage by looking through abstracts and titles. The first round of screening led to 31 records for the following full-text review. Finally, 16 studies [33–40, 42–49] were included in our systematic review and meta-analysis. The screening process was described as the PRISMA flow diagram (Fig 1).

Fig 1

Flow diagram of study inclusion and exclusion.

There were three conference abstracts among included studies [33-35]. Of all the RCTs included, two were based on a cross-over design [34, 36]. Eligible Infants enrolled were categorized by preterm (n = 8), full-term (n = 5) and both (n = 3). Infants underwent different painful procedures including heel stick (n = 8), retinopathy of prematurity (ROP) screening (n = 5), venipuncture (n = 2) and wound dressing (n = 1). PIPP score was mostly widely used for pain assessment (n = 10) while PIPP-R(n = 1), NIPS (n = 3), NFCS (n = 1) and N-pass score(n = 1) were also applied in other studies. Characteristics of eligible studies were outlined in Table 1. Fig 2A, 2B demonstrated the risk of bias assessment for each study.

Table 1

The characteristics of included studies.

First author, year, country	Study Type/design	Objects	Procedure	Sample size	Gestational age and weight (inclusion criteria)	Intervention/Control groups	Outcomes
Asmerom,2013, United States [46]	Prospective double-blind randomized controlled study	preterm	heel lance	sucrose+NNS group: n = 44	premature infants ≤ 36.5	sucrose+NNS group:	PIPP scored from the time of heel lance to 30 seconds post the lance.
						received a single dose of 24% sucrose via syringe to the anterior tongue along with a pacifier (NNS) two minutes before the heel lance.
				water+NNS group: n = 45	weeks gestation who weighed ≥800 grams
						Water+NNS group: the placebo group received an equal volume of sterile water to the anterior portion of the tongue along with a pacifier.
Benoit,2021, Canada [47]	Single-blind randomized controlled trial	healthy full-term	heel lance	sucrose+NNS: n = 19	healthy, full-term (born ≥ 37 0/7 weeks’gestation)	Intervention group: 24% oral sucrose combined with offered NNS and containment in a blanket while in an infant cot. Control group: direct breastfeeding	1.pain-related brain activity
							2.PIPP-R at 30-, 60-, 90-, and 120-s following heel lance
				breastfeeding: n = 18
							3.adverse events
Boyle, 2016, Canada [42]	Prospective, Randomised, placebo controlled study	preterm	ROP screening	group 1: n = 10	< 32 weeks’ PMA	Group 1: 1 ml sterile water given by mouth using a syringe	PIPP scores during eye examination
				group 2: n = 10
						Group 2: 1 ml sucrose 33% given by mouth using a syringe
				group 3: n = 9		Group 3: 1 ml sterile water given by mouth using a syringe and pacifier put into the mouth
				group 4: n = 11		Group 4: 1 ml sucrose 33% given by mouth using a syringe and pacifier put into the mouth
Collados,2018, Spain [36]	Multicentre randomised, non-inferiority, cross-over trial	preterm	venipuncture	EBM-sucrose sequence: n = 33	gestational age of less than 37 weeks at birth and weigh less than 2,500 grams.	In group one, the neonate received EBM during the first venipuncture that was included in the study and 24% sucrose during the second venipuncture. The process was reversed for group two.	PIPP, the duration of crying, oxygen saturation and heart rate
				sucrose-EBM sequence: n = 33
						This accompanied throughout by non-nutritive sucking and swaddling.
Cullas,2012, Turkey [33]	Prospective randomised study	preterm	ROP screening	sucrose+NNS: n = 21	patients under 32 weeks of gestational age	Group 1: oral sucrose solution given two minutes before examination. Pacifier was used.	PIPP, time of crying
				water+NNS: n = 19
						Group 2: sterile water given two minutes before examination. Pacifier was used.
De Bernardo,2019, Italy [48]	Randomized double-blinded case–control pilot study	full-term	venipuncture	sucrose+NNS: n = 33	neonates 37–42 weeks gestational age at birth and >1 week old at the time of the intervention with body weight 2,500–4,500 g and able to feed orally.	Intervention group: received both 1 mL 24% sucrose orally via syringe 1 minute before venipuncture and 1 mL during the procedure. A pacifier was offered to all neonates immediately following sucrose administration each infant.	NIPS
							Outcome measurements (HR, SpO2) were obtained before (T0), during (T1), and 1 minute after (T2) venipuncture
				glucose+NNS: n = 33
						Control: received 1 mL 10% glucose orally via syringe with a pacifier 1 minute before venipuncture and during the procedure
Dilli, 2014, Turkey [49]	Prospective randomised and placebo-controlled study	preterm	ROP screening	Group 1: n = 32	-	Group 1: 0.5 mL/kg of sucrose 24% was given by mouth using a syringe, and pacifier was placed in the mouth.	PIPP score during examination crying time
				Group 2: n = 32
						Group 2: 0.5 mL/kg of sterile water was given by mouth using a syringe, and pacifier was placed in the mouth.
Gao,2018, China [44]	Randomized controlled trial	Preterm infants	heel stick	1.NNS group: n = 22	before 37 weeks of gestation	1.NNS group: pacifier given in 2 minutes before, and throughout the recovery phase of the heel stick.	1.PIPP scale in the blood collection phase (0-60s) and recovery phase (after 1 min)
						2. sucrose group: Sucrose 20% (0.2 mL/kg) was administrated to the preterm infant’s mouth by 1 ml syringe in 2 minutes before the heel stick procedure	2.heart rate and oxygen saturation
							3.the percentage of crying time respectively in the blood collection phase and recovery phase
						3.sucrose+NNS: Sucrose 20% (0.2 mL/kg) was administrated to the preterm infant’s mouth by 1 ml syringe in 2 minutes before the heel stick procedure and a pacifier was given until the recovery phase of the heel stick.
						4. routine care group: received only routine comfort through gentle touch when he cried after the heel stick procedure.
				2.sucrose group: n = 21
				3.NNS+sucrose group: n = 22
				4.routine care group: n = 21
Gibbins,2002, Canada [45]	Randomized controlled trial	Preterm/term	heel lance	1.sucrose+NNS: n = 64	born between 27 and 43 weeks gestation	1. sucrose+NNS group: received 0.5 ml of 24% sucrose via a syringe followed immediately by the insertion of a pacifier into the mouth. The pacifier was held in place as required 2 minutes before, during, and 5 minutes following the heel lance.	the PIPP are numerically scored on 30/60 seconds following an acute painful stimulus.
				2.sucrose alone: n = 62
				3.water+NNS: n = 64
						2. sucrose group: received 0.5 ml of 24% sucrose via a syringe.
						3.water+NNS group: received 0.5 ml of sterile water via a syringe. No pacifier was offered.
Leng,2016, China [43]	Prospective, multi-centred, randomized controlled clinical trial	full term	shallow or deep heel stick procedures	NS group: n = 167	gestational age between 37 and 42 weeks at delivery;	Group S: 2 ml of 24% sucrose was administrated by syringe 2 min before the heelstick procedure.	NFCS score
						Group NS: 2 ml of 24% sucrose was administrated by syringe 2 min before the heel stick procedure, and then a standard silicone newborn pacifier was placed into the infant’s mouth until the end of the process.
				S group: n = 176
					Birthweight between 2500 g and 4000 g;
Miller, 2009, United States [34]	Repeated-measures crossover design.	preterm/full term	heel stick	NNS+sucrose-no treatment sequence: n = 7	between the ages of 32	1.In the treatment condition, infants were offered NNS with sucrose.	NIPS score, heart rate and oxygen saturation
				no treatment-NNS+sucrose sequence: n = 7	weeks to younger than or equal to 42 weeks	2.In the control condition, infants were not offered any treatment
Mandee,2020, Thailand [37]	Prospective randomized control trial	preterm/full term	Wound dressing	sucrose+NNS group: n = 16	-	Sucrose+NNS group: participants were first administered a dose of 24% sucrose and then were given the pacifier.	The NIPS scores were assessed at 30, 120, and 240 seconds
						NNS group: participants were administered a pacifier while their wound dressing was performed.	from the commencement of the wound dressing, crying time
				NNS group: n = 16
Mitchell,2016, United States [39]	Double-blind 2×2 factorial randomized controlled trial	full term	heel stick	sucrose+NNS group: n = 37	healthy term infants between 37 and 42 weeks	1.sucrose+NNS: received 1±0.1 ml of the 24% sucrose solution orally with pacifier at 2 ±0.5 minutes prior to the procedure2.water+NNS: received 1 ±0.1 ml of sterile water with pacifier	PIPP, heart rate variability (HRV), and salivary cortisol
				water+NNS group: n = 39
O’Sullivan,2010, Ireland [38]	Randomised placebo controlled study	preterm	ROP screening	sucrose+NNS group: n = 20	< 1501 g	1. sucrose+NNS: infants were swaddled and received 0.2 ml of sucrose 24% given by mouth using a syringe and a soother	N-PASS score
				water+NNS group: n = 20	< 32 Weeks gestation Infants
						2. water+NNS: infantsswaddled, and received 0.2 ml of sterile water given by mouth using a syringe and a soother.
Thakkar,2016, India [40]	Randomized controlled trial	full term	heel-stick	group I (sucrose): n = 45	(>37 weeks PMA), with birthweight > 2200 g	1.group I received 30% sucrose solution by sterile syringe;	PIPP score, heart rate, oxygen saturation, duration of crying
				group II (NNS): n = 45
						2.group II received NNS in which sterile gauze was held gently in neonate’s mouth and the palate tickled to stimulate sucking;
				group III (sucrose+NNS): n = 45
						3.group III received both the interventions (sucrose and NNS);
						4.group IV received no intervention.
				group IV: n = 45
Ucar S,2014, Turkey [35]	Randomised, controlled study	preterm	prematurity (ROP) screening	sucrose group: n = 27	-	group 1 received 24% sucrose oral,	PIPP
						group 2 received 24% sucrose with pacifier
				sucrose+NNS: n = 27
				water+NNS: n = 27
						group 3 received sterile water with pacifier.

ROP: retinopathy of prematurity; NNS: nonnutritive sucking; PIPP: Premature Infant Pain Profile; NFCS: Neonatal Facial Coding System; NIPS: Neonatal Infant Pain Scale

Fig 2

Risk of bias assessment for each included trial.

A, Risk of bias graph; B: Risk of bias summary.

3.2 Pain score

3.2.1 NNS+OS group versus NNS group

A total of 11 studies with 677 participants explored the effect of OS combined with NNS compared with NNS alone during the painful procedures, most of which reported PIPP score, while one study reported NIPS [37] and one reported N-pass score [38]. In both heel-stick and ROP subgroups, a significant effect was observed under the random-effects model, with the standard mean difference being -1.59 (95%CI: -2.49, -0.68) and -1.05(95%CI: -1.56, -0.55), respectively. However, there was no significant difference in the wound dressing subgroup (Fig 3). A sensitivity analysis was conducted based on two aspects: removing conference abstracts and removing articles using non-PIPP measurements. The results remained significant after two conference abstracts were removed in the ROP group [33, 35]. Besides, unifying the measurements by removing two studies reporting NIPS or N-pass score did not change the results or reduce heterogeneity neither. The analysis showed no effect in four studies involving term infants regardless of the type of procedures (p = 0.16) with SMD: -0.81 (95%CI: -1.94, 0.33) [37, 39–41].

Fig 3

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with applying NNS alone during different painful procedures.

NNS: nonnutritive sucking.

Fewer studies provided information in the recovery phase. The available 4 studies showed the combined sucrose and NNS did not provide a better effect in relieving pain in both heel stick and wound dressing groups (Fig 4). However, this finding was not robust in the heel stick subgroup due to the reverse result (MD = -3.23, 95%CI: -4.56, -1.89, P<0.001) after removing Mitchell’s study in the sensitivity analysis [39].

Fig 4

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with applying NNS alone in the recovery phase.

NNS: nonnutritive sucking.

3.2.2. NNS+OS versus OS alone

Six studies [35, 40, 42–45] involving 677 infants or newborns assessed the pain in the group applying sucrose alone. One study measured pain as NFCS score [43] while the other five studies reported PIPP. The meta-analysis showed a significant effect of the combined interventions than OS (Fig 5). No significant difference, however, was detected when studying the term subgroup (SMD = -1.30, 95%CI: -2.81, 0.22, P = 0.09) [40, 41, 43]. In the recovery phase, the effect was also significant (MD = -3.48, 95%CI: -5.41, -1.54) (Fig 6).

Fig 5

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with applying sucrose alone during heel stick and ROP.

NNS: nonnutritive sucking; ROP: retinopathy of prematurity.

Fig 6

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with applying sucrose alone in recovery phase.

NNS: nonnutritive sucking.

3.2.3 NNS+OS versus routine care group

Five studies involving 268 participants were included in the analysis to compare the effect between the NNS+sucrose group and the routine care group [34, 40, 42, 44, 46]. The results showed a better impact of NNS+sucrose in both heel stick (SMD = -2.66, 95%CI: -4.53, -0.78) and ROP group (SMD = -1.1, 95%CI: -2.03,-0.17) (Fig 7). Removal of any study did not change the results significantly.

Fig 7

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with routine care group during the procedures.

NNS: nonnutritive sucking.

3.2.4 NNS+OS versus breastfeeding or breast milk

The comparison between OS+NNS and breast milk showed no difference in effects for pain relief despite the heterogeneity in study design, intervention methods, pain procedures, outcome assessment and target population between the two studies (Fig 8). In Benoit’s study [47], full term infants underwent heel stick and were direct breastfed, whose pain was assessed by PIPP-R. In contrast, 66 preterm infants underwent venipuncture in two sequences. During the controlling period, infants were given breast milk via a pacifier, whose pain was assessed by PIPP in Collados’s study [36].

Fig 8

Forest plot of pain score comparing the combined intervention (NNS and sucrose) with breastmilk during the procedures.

NNS: nonnutritive sucking.

3.2.5 NNS+OS versus glucose

Only one study enrolling 66 term newborns set glucose+NNS as the control group [48]. The result demonstrated a significant difference (p < 0.05) in both two phases. NIPS scores were significantly lower in the NNS+sucrose group (range 1–2 and median 0) compared with the NNS+glucose group (range 5–7 and median 6) during venipuncture.

3.2.6 Publication bias

Publication bias was tested by Egger’s Test and funnel plots (Fig 9A, 9B). The results of Egger’s test indicated no publication bias in both scenarios. The bias (intercept) was estimated -3.3 (95%CI -10.90, 4.30) with p value 0.3515 in the comparison between NNS+OS and NNS alone (11 studies). The estimates of bias were -5.60 (95%CI -13.40, 2.20) with p value 0.117 in the comparison between NNS+OS and OS alone (6 studies). The funnel plots presented slight asymmetry especially in heel-stick subgroup. More studies are needed to justify this publication bias.

Fig 9

Funnel plots of publication bias on NNS+OS versus NNS alone (A) and NNS+OS versus OS alone (B) during painful procedures. NNS: nonnutritive sucking; OS: oral sucrose.

3.3 Adverse events

Among the included studies, ten trials [36–38, 40, 43–45, 47–49] reported the occurrence of adverse events. Five trials observed that the occurrence rate of adverse events was zero in all treatment groups [36, 37, 43, 47, 48]. Oxygen desaturation was the most common adverse event, reported in 4 trials [38, 40, 45, 49]. Gao reported 5 episodes of vomit and abdominal distension whereas Dilli and O’Sullivan reported 13 and 4 episodes of bradycardia, respectively. A significant difference in tachycardia between the NNS+sucrose and the NNS alone group was observed by Dilli, whereas no significant adverse effects were observed in other trials (Table 2). Three trials reported heart rate and oxygen saturation as mean (SD) as a secondary outcome (Table 3) [34, 44, 46]. They all observed a significant effect of NNS+sucrose on oxygen saturation and heart rate except a non-significant result in Asmerom’s for heart rate.

Table 2

Episodes of adverse events in included studies.

Study	NNS+sucrose: N(Total)	NNS alone: N(Total)	Sucrose alone: N (Total)	Routine care: N(Total)	Clinical significance
Dilli, 2014 [49]	bradycardia:6(32)	bradycardia:7 (32)	-	-	bradycardia: p = 0.75
	tachycardia:12(32)	tachycardia:19 (32)			tachycardia: p = 0.08
	desaturations:6(32)	desaturations:7 (32)			desaturations: p = 0.75
Gao, 2018 [44]	vomit:1(22)	vomit:1(22)	-	Vomit:1	vomit: p = 0.800;
				abdominal distension:1	abdominal distension: p = 0.562
	abdominal distension:1(22)
Gibbins, 2002 [45]	No adverse events	oxygen desaturation:2(64)	oxygen desaturation:3(64)	-	p>0.05
		choke:1(64)
O’Sullivan, 2010 [38]	bradycardia:1(20)	bradycardia:3(20)	-	-	bradycardia: p = 0.328
	desaturations:1(20)	desaturations:3(20)			desaturations: p = 0.328
Thakkar, 2016 [40]	oxygen desaturation:1(45)	oxygen desaturation:1(45)	oxygen desaturation:1(45)	oxygen desaturation:2(45)	p>0.05

Table 3

Summarized statistics of oxygen desaturation and Heart rate in included studies during painful procedures.

Study	Sucrose+NNS		NNS		sucrose		Routine care		p-value
	Mean (SD)	N	Mean (SD)	N	Mean (SD)	N	Mean (SD)	N
oxygen saturation
Asmerom, 2013 [46]	96.4(0.6)	44	95.8(0.6)	45	-	-	-	-	<0.0001
Gao, 2018 [44]	95.2(1.6)	22	92.9(2.4)	22	93.5(1.7)	21	92.9(2.1)	21	<0.05
Miller, 2009 [34]	97.69(2.41)	14	-	-	-	-	94.30(3.74)	14	<0.001
Heart rate
Asmerom, 2013 [46]	170.5(14.7)	44	164.9(14.6)	45	-	-	-	-	0.07
Gao, 2018 [44]	138.6(7.9)	22	154.2(9.0)	22	151.6(9.6)	21	156.8(7.2)	21	<0.0001
Miller, 2009 [34]	135.64(7.71)	14	-	-	-	-	150.64(7.53)	14	<0.001

NNS: nonnutritive sucking; SD: Standard deviation.

4 Discussion

Pain management has been well established in recent years, especially in procedures involving skin punctures [12, 50]. However, there is not a practical guideline on how to reduce pain and stress in a prolonged procedure such as ROP and wound treatment [51]. Therefore, this systematic review and meta-analysis aims to determine the effect of combined NNS and sucrose intervention in different types of painful procedures.

Overall, the combined interventions of NNS and sucrose showed the superiority in relieving mild pain during the painful procedures compared to applying NNS alone or sucrose alone or standard care. This has been confirmed by previous reviews [10, 13]. Non-nutritive sucking was believed to be associated with antinociceptive mechanisms and sucrose appears to enhance the effect of NNS, leading to an increase of endogenous endorphins [9, 52]. However, the mechanism of the combined effects is not completely clear [53].

On the other hand, the superiority is not significant when compared with direct breastfeeding or giving breast milk via a pacifier [36, 47]. The similarity between sucking through a pacifier and breastfeeding might explain this result [52]. Besides, swaddling and a blanket used in the two trials helped establish a simulated environment as breastfeeding, calming and comforting the babies. The similar effectiveness in healthy term neonates or stable late preterm neonates between sucrose/glucose administration and breast milk was also proposed in Shah’s review [54].

ROP examinations and wound dressing have two points in common. Both procedures last for several minutes and tend to give giving stronger painful stimuli. The insertion of an eyelid speculum and scleral depression usually increases the intensity of the pain during ROP screening [55]. Although a significant effect was observed in ROP examinations in a preterm population, we found the means of PIPP score even in the combined NNS and sucrose group still around or higher than 12, qualitatively, indicating a moderate-to-severe level. Considering the preterm infants tend to have lower scores than term infants, such high scores indicated the finite effect of this combined intervention [56]. Besides, a non-significant effect was detected during wound dressing in Mandee’s study where moderate-to-severe pain were often observed [37]. This might not be indicative because few studies researched on pain managements in wound dressing. However, previous studies found the effect of sucrose were time-dependent and could not last for a long time [10, 41]. This might be able to explain the minor effect. More innovations are needed on non-pharmacological interventions for those intensive procedures. For example, a recent study recommended the physiological flexion position called ROP position and it was demonstrated more effective than the combination of sucrose and NNS [57]. The combination of multiple non-pharmacological methods, including non-nutritive sucking, oral breast milk, and facilitated tucking, music, etc. might also help enhance the effects of pain relief [17, 18, 58]. Investigations on more effective methods are warranted, especially for lowering moderate or even severe pain.

Few studies in our analysis reported the pain measurements in the recovery phase, which might lead to an unreliable result in the meta-analysis. Our analysis found the effect has vanished in heel stick procedure when comparing the combined interventions with NNS alone group during this period. This finding could be reversed by removing a study with nonsignificant result in sensitivity analysis [39]. In this trial, both intervention and control groups received a pacifier and facilitated tucking resulting a low PIPP and sucrose did not contribute much to reducing pain. In fact, facilitated tucking has been reported to be effective in pain management [59].

Subgroup analysis on sucrose administration was not performed in the meta-analysis because of the varieties in different trials in terms of volumes (from 0.1 mL to 2 mL), concentrations (range from 12% to 33%) and frequencies. However, Steven found no difference in effectiveness among doses by proposing the minimal effective dose of only 0.1 mL in relieving procedural pain [60]. Thus, the wide range of volumes of sucrose might not influence the comparison.

OS has been considered to cause adverse events such as desaturation and bradycardia, especially in preterm infants [10]. Our analysis indeed indicates the trend of oxygen saturation increasing while heart rate decreasing under intervention. However, we did not observe a higher occurrence of bradycardia, tachycardia or desaturations in groups with OS. Besides, adverse events indeed occurred more in preterm subjects. Trials with zero adverse events were mostly based on full-term infants. This might indicate the vulnerability of preterm infants attributes more to adverse events than OS. Nonetheless, the evidence might be influenced by reporting bias because some studies did not report any information on the safety during the intervention procedure.

Although the combined intervention of sucrose and NNS was considered effective in reducing pain among the studies included, sucrose was often administered 2 min prior to the painful procedure in these studies. This time interval was unjustified by Meesters in 2017 [61]. Besides, the recovery phase was neglected in half of the studies. The recovery from pain is important as it also affects the baby in a long-term way. We might improve the intervention methods to deal with moderate and severe pain and pay more attention to the recovery phase.

De Bernardo’s trial found sucrose was more effective then oral glucose when both combined with a pacifier. This conflicts with a previous review which take glucose as an acceptable alternative to sucrose [62]. More studies could be conducted in this direction in order to expand the available and effective intervention methods.

There are three limitations in our analysis. Firstly, the heterogeneity is non-negligible in heel-stick subgroups but acceptable in other painful procedures. Heel-stick was involved in most studies. The variations in their protocols, such as population, the operating process, the pain measurements, or the sample size might interactively contribute to moderate to high heterogeneity. For example, some studies enrolled both preterm and term infants in the trial, while others enrolled purely preterm or full-term infants, which added the difficulty in explaining the effects on the certain population. Different scales and standards in pain measurements also influenced pooling data in quantitative analysis. We have conducted post hoc subgroup analyses to test the source of heterogeneity in terms of these factors. High heterogeneity might induce an unreliable conclusion even under a random-effects model. Results should be explained with caution. Secondly, pain score was the only one outcome to measure pain in this analysis and the different assessments were pooled. Those pain assessment methods are distinguishable from their target population, scales and evaluation items [63]. It was hard to unify because PIPP was the most common and other assessments accounted for no more than 25% of included studies. A sensitivity analysis was conducted to reduce the potential bias though. Other measurements including crying time were not investigated because of the limited number of studies. There were very few studies reporting crying time and comparing the combined intervention with sucrose alone or routine care. A single outcome of pain score might not be robust enough since different scales were used among studies. Finally, publication bias seems to exist in our analysis.

5 Conclusion

In conclusion, our systematic review and meta-analysis indicated the superiority of the combined intervention of sucrose and NNS than any single intervention except for breastfeeding. However, the effect appears to be mild in alleviating moderate-to-severe pain. More exploration and improvement of intervention were needed.

PRISMA 2009 checklist.

(DOCX)

Click here for additional data file.

Full searching strategy (from Jan 1, 2000 to Mar 31, 2021).

(DOCX)

Click here for additional data file.

27 Dec 2021

A rebuttal letter that responds to each point raised by the academic editor and reviewer(s). You should upload this letter as a separate file labeled 'Response to Reviewers'.

A marked-up copy of your manuscript that highlights changes made to the original version. You should upload this as a separate file labeled 'Revised Manuscript with Track Changes'.

An unmarked version of your revised paper without tracked changes. You should upload this as a separate file labeled 'Manuscript'.

If you would like to make changes to your financial disclosure, please include your updated statement in your cover letter. Guidelines for resubmitting your figure files are available below the reviewer comments at the end of this letter.

If applicable, we recommend that you deposit your laboratory protocols in protocols.io to enhance the reproducibility of your results. Protocols.io assigns your protocol its own identifier (DOI) so that it can be cited independently in the future. For instructions see: https://journals.plos.org/plosone/s/submission-guidelines#loc-laboratory-protocols. Additionally, PLOS ONE offers an option for publishing peer-reviewed Lab Protocol articles, which describe protocols hosted on protocols.io. Read more information on sharing protocols at https://plos.org/protocols?utm_medium=editorial-email&utm_source=authorletters&utm_campaign=protocols.

We look forward to receiving your revised manuscript.

Kind regards,

Girish Chandra Bhatt, MD, FASN

Academic Editor

PLOS ONE

Journal Requirements:

When submitting your revision, we need you to address these additional requirements.

1. Please ensure that your manuscript meets PLOS ONE's style requirements, including those for file naming. The PLOS ONE style templates can be found at

https://journals.plos.org/plosone/s/file?id=wjVg/PLOSOne_formatting_sample_main_body.pdf and https://journals.plos.org/plosone/s/file?id=ba62/PLOSOne_formatting_sample_title_authors_affiliations.pdf

2. Thank you for stating in your Funding Statement:

“Our paper is supported by Project of Sichuan Provincial Health Commission in 2014 (NO: 140108 ).”

Please provide an amended statement that declares *all* the funding or sources of support (whether external or internal to your organization) received during this study, as detailed online in our guide for authors at http://journals.plos.org/plosone/s/submit-now.  Please also include the statement “There was no additional external funding received for this study.” in your updated Funding Statement.

Please include your amended Funding Statement within your cover letter. We will change the online submission form on your behalf.

One or more authors are affiliated with the funder, but authors state that the funder had no role

Thank you for stating the following financial disclosure:

“The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.”

We note that one or more of the authors is affiliated with the funding organization, indicating the funder may have had some role in the design, data collection, analysis or preparation of your manuscript for publication; in other words, the funder played an indirect role through the participation of the co-authors. If the funding organization did not play a role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript and only provided financial support in the form of authors' salaries and/or research materials, please do the following:

a. Review your statements relating to the author contributions, and ensure you have specifically and accurately indicated the role(s) that these authors had in your study. These amendments should be made in the online form.

b. Confirm in your cover letter that you agree with the following statement, and we will change the online submission form on your behalf:

“The funder provided support in the form of salaries for authors [insert relevant initials], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.

3. We note that you have indicated that data from this study are available upon request. PLOS only allows data to be available upon request if there are legal or ethical restrictions on sharing data publicly. For more information on unacceptable data access restrictions, please see http://journals.plos.org/plosone/s/data-availability#loc-unacceptable-data-access-restrictions.

In your revised cover letter, please address the following prompts:

a) If there are ethical or legal restrictions on sharing a de-identified data set, please explain them in detail (e.g., data contain potentially sensitive information, data are owned by a third-party organization, etc.) and who has imposed them (e.g., an ethics committee). Please also provide contact information for a data access committee, ethics committee, or other institutional body to which data requests may be sent.

b) If there are no restrictions, please upload the minimal anonymized data set necessary to replicate your study findings as either Supporting Information files or to a stable, public repository and provide us with the relevant URLs, DOIs, or accession numbers. For a list of acceptable repositories, please see http://journals.plos.org/plosone/s/data-availability#loc-recommended-repositories.

We will update your Data Availability statement on your behalf to reflect the information you provide.

4. PLOS requires an ORCID iD for the corresponding author in Editorial Manager on papers submitted after December 6th, 2016. Please ensure that you have an ORCID iD and that it is validated in Editorial Manager. To do this, go to ‘Update my Information’ (in the upper left-hand corner of the main menu), and click on the Fetch/Validate link next to the ORCID field. This will take you to the ORCID site and allow you to create a new iD or authenticate a pre-existing iD in Editorial Manager. Please see the following video for instructions on linking an ORCID iD to your Editorial Manager account: https://www.youtube.com/watch?v=_xcclfuvtxQ.

5. Please include captions for your Supporting Information files at the end of your manuscript, and update any in-text citations to match accordingly. Please see our Supporting Information guidelines for more information: http://journals.plos.org/plosone/s/supporting-information.

6. Please include a copy of Table 2 & 3 which you refer to in your text on page 13.

[Note: HTML markup is below. Please do not edit.]

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

**********

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: No

**********

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

**********

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: No

**********

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Interesting, well written and well designed review and meta--analysis on efficacy and safety of non-pharmacological interventions on pain relief in infants.

Here my comments:

- introduction:once you introduce the abbreviation e.g OS, I suggest to use it instead of repeating oral sucrose.

- methods: please clarify if the research was restricted for english language articles or not; the statements were not clear for this point.

- discussion: you were discussiong about non pharamcological intervention; I think that citing studies about ketorolac was unnecessary.

Reviewer #2: The authors have made great efforts in doing this systematic review and meta-analysis. There are few concerns :

1. How was the cross over trial managed and which data was included in the meta-analysis is not mentioned. The methods section should clearly indicate how the cross-over designs will be dealt with.

2. The forests plots show critical levels of heterogeneity despite the subgroup analysis. The manuscript doesn't describe any efforts to explore the reasons for this heterogeneity. It is also worth considering if the meta-analysis with such high level of heterogeneity is required. If yes please add few lines about exploring heterogeneity. It is acceptable to have unexplained heterogeneity but a justification and reasoning with references is required.

3. Authors have interpreted the funnel plot as showing publication bias. It would be better to add Eggers test and then describe the funnel plots. Only consider funnel plots if number of studies in an analysis are more then 10

4. Authors have mentioned "sensitivity analysis conducted on 2 aspects..." please describe these 2 aspects and also add some details about sensitivity analysis in the method section

5. Overall a relook at the grammar and rephrasing of certain sentences is advised.

**********

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

21 Feb 2022

Response letter

Dear Editors and Reviewers,

Thank you for your valuable comments. We have revised them one by one according to these comments. We hope that the revised manuscript is now acceptable for publication in your journal.

Thank you again for your time and consideration.

Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented.

Reviewer #1: Yes

Reviewer #2: Partly

2. Has the statistical analysis been performed appropriately and rigorously?

Reviewer #1: Yes

Reviewer #2: No

3. Have the authors made all data underlying the findings in their manuscript fully available?

The PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data—e.g. participant privacy or use of data from a third party—those must be specified.

Reviewer #1: Yes

Reviewer #2: Yes

4. Is the manuscript presented in an intelligible fashion and written in standard English?

PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.

Reviewer #1: Yes

Reviewer #2: No

5. Review Comments to the Author

Please use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. (Please upload your review as an attachment if it exceeds 20,000 characters)

Reviewer #1: Interesting, well written and well designed review and meta--analysis on efficacy and safety of non-pharmacological interventions on pain relief in infants.

Here my comments:

- introduction:once you introduce the abbreviation e.g OS, I suggest to use it instead of repeating oral sucrose.

Reply: Thanks. This problem has been updated.

- methods: please clarify if the research was restricted for english language articles or not; the statements were not clear for this point.

Reply: “Non-English literature were excluded.” was clarified in the exclusion criteria.(page 5 line 97)

- discussion: you were discussing about non pharamcological intervention; I think that citing studies about ketorolac was unnecessary.

Reply: Thanks for your suggestion. The references and related opinions have been deleted.

Reviewer #2: The authors have made great efforts in doing this systematic review and meta-analysis. There are few concerns :

1. How was the cross over trial managed and which data was included in the meta-analysis is not mentioned. The methods section should clearly indicate how the cross-over designs will be dealt with.

Reply: The results of cross-over design trials were standardized. The details were added to the method section. (page 7 line 130)

2. The forests plots show critical levels of heterogeneity despite the subgroup analysis. The manuscript doesn't describe any efforts to explore the reasons for this heterogeneity. It is also worth considering if the meta-analysis with such high level of heterogeneity is required. If yes please add few lines about exploring heterogeneity. It is acceptable to have unexplained heterogeneity but a justification and reasoning with references is required.

Reply: Possible explanations of high heterogeneity have been added to the limitation part. (page 17 line 342)

3. Authors have interpreted the funnel plot as showing publication bias. It would be better to add Eggers test and then describe the funnel plots. Only consider funnel plots if number of studies in an analysis are more then 10

Reply: Thanks for your advice. Egger’s Test has been added. (page 12 line 235)

4. Authors have mentioned "sensitivity analysis conducted on 2 aspects..." please describe these 2 aspects and also add some details about sensitivity analysis in the method section

Reply: Two aspects were described(page 10 line 192). Some sentences about sensitivity analysis were added to the last paragraph in method section. (page 7 line 148)

5. Overall a relook at the grammar and rephrasing of certain sentences is advised.

Reply: Thank you. The whole manuscript has been updated.

6. PLOS authors have the option to publish the peer review history of their article (what does this mean?). If published, this will include your full peer review and any attached files.

If you choose “no”, your identity will remain anonymous but your review may still be made public.

Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.

Reviewer #1: No

Reviewer #2: No

[NOTE: If reviewer comments were submitted as an attachment file, they will be attached to this email and accessible via the submission site. Please log into your account, locate the manuscript record, and check for the action link "View Attachments". If this link does not appear, there are no attachment files.]

While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/. PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org. Please note that Supporting Information files do not need this step.

Finally, we would like to express my sincere appreciations to you for your hard work, dear editors and reviewers. We look forward to having more opportunities to learn from you in the future!

Best regards!

Submitted filename: Response to Reviewers.doc

Click here for additional data file.

21 Apr 2022

Efficacy and safety of combined oral sucrose and nonnutritive sucking in pain management for infants: A systematic review and meta-analysis

PONE-D-21-21132R1

Dear Dr. Zou,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

Within one week, you’ll receive an e-mail detailing the required amendments. When these have been addressed, you’ll receive a formal acceptance letter and your manuscript will be scheduled for publication.

An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.

If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they’ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact onepress@plos.org.

Kind regards,

Girish Chandra Bhatt, MD, FASN

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments:

28 Apr 2022

PONE-D-21-21132R1

Efficacy and safety of combined oral sucrose and nonnutritive sucking in pain management for infants: A systematic review and meta-analysis

Dear Dr. Zou:

I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

If we can help with anything else, please email us at plosone@plos.org.

Thank you for submitting your work to PLOS ONE and supporting open access.

Kind regards,

PLOS ONE Editorial Office Staff

on behalf of

Dr. Girish Chandra Bhatt

Academic Editor

PLOS ONE