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Updated Meta-analysis of Probiotics for PreventingNecrotizing Enterocolitis in Preterm Neonates
WHAT’S KNOWN ON THIS SUBJECT: Previous systematic reviewsof RCTs indicated significantly lower risk for all-cause mortalityand definite NEC and shorter time to full feeds after probioticsupplementation in preterm (�34 weeks’ gestation) VLBW (birthweight�1500 g) neonates.
WHAT THIS STUDY ADDS: The results of this conclusive updatedmeta-analysis confirm the benefits of probiotic supplements inreducing death and disease in preterm neonates. Given thetotality of the evidence, additional placebo-controlled trials areunnecessary if a suitable probiotic product is available.
abstract +
OBJECTIVE: Systematic reviews of randomized, controlled trials(RCTs) indicate lowermortality and necrotizing enterocolitis (NEC) andshorter time to full feeds after probiotic supplementation in preterm(�34 weeks’ gestation) very low birth weight (VLBW; birth weight�1500 g) neonates. The objective of this study was to update our 2007systematic review of RCTs of probiotic supplementation for prevent-ing NEC in preterm VLBW neonates.
METHODS: We searched in March 2009 the Cochrane Central register;Medline, Embase, and Cinahl databases; and proceedings of the Pedi-atric Academic Society meetings and gastroenterology conferences.Cochrane Neonatal Review Group search strategy was followed. Selec-tion criteria were RCTs of any enteral probiotic supplementation thatstartedwithin first 10 days and continued for�7 days in preterm VLBWneonates and reported on stage 2 NEC or higher (Modified BellStaging).
RESULTS: A total of 11 (N � 2176), including 4 new (n � 783), trialswere eligible for inclusion in the meta-analysis by using a fixed-effectsmodel. The risk for NEC and death was significantly lower. Risk forsepsis did not differ significantly. No significant adverse effects werereported. Trial sequential analysis) showed 30% reduction in the inci-dence of NEC (� � .05 and .01; power: 80%).
CONCLUSIONS: The results confirm the significant benefits of probi-otic supplements in reducing death and disease in preterm neonates.The dramatic effect sizes, tight confidence intervals, extremely low Pvalues, and overall evidence indicate that additional placebo-controlled trials are unnecessary if a suitable probiotic product isavailable. Pediatrics 2010;125:921–930
AUTHORS: Girish Deshpande, FRACP,a,b Shripada Rao,MD,a,b Sanjay Patole, DrPH,a,c and Max Bulsara, PhDdaDepartment of Neonatal Paediatrics, King EdwardMemorial HospitalforWomen, Perth, Western Australia, Australia; bDepartment ofNeonatal Paediatrics, PrincessMargaret Hospital for Children, Perth,Western Australia, Australia; cSchool of Women’s and Infant’s Health,University of Western Australia, Perth, Western Australia, Australia; anddDepartment of Biostatistics, University of Notre Dame, Perth, WesternAustralia, Australia
ABBREVIATIONSNEC—necrotizing enterocolitisLOS—late-onset sepsisVLBW—very low birth weightRCT—randomized, controlled trialCNRG—Cochrane Neonatal Review GroupTFF—time to full feedsTSA—trial sequential analysisRR—relative riskCI—confidence intervalNNT—numbers needed to treatTPN—total parenteral nutritionNDI—neurodevelopmental impairmentCONS—coagulase-negative StaphylococcusELBW—extremely low birth weight
Thisworkwas presented in part at Perinatal Society of Australia andNew Zealandmeeting; April 19–22, 2009; Darwin, Australia (J PediatrChild Health. 2009;45:S1–A029).
Dr Deshpande participated in the literature search, selection of trials,quality assessment of trials, contacting the authors of the trials foradditional information, performing analysis, andwriting themanuscript; Dr Rao did independent literature search, identified trialsfor inclusion and exclusion, assessed themethodologic quality ofincluded trials, and extracted and also verified the data entered by DrDeshpande in the RevMan software; Prof Bulsarawas responsible forconducting the trial sequential analysis and interpreting its results andalso contributed to the revisedmanuscript; and Dr Patolewasresponsible for the concept, design, interpretation of analysis, andwriting of the final version of themanuscript that was seen andapproved by all authors. All authors had full access to all of the data(including statistical reports and tables) in the study and takeresponsibility for the integrity of the data and the accuracy of the dataanalysis.
www.pediatrics.org/cgi/doi/10.1542/peds.2009-1301
doi:10.1542/peds.2009-1301
Accepted for publication Jan 15, 2010
Address correspondence to Sanjay Patole, DrPH, Department ofNeonatal Paediatrics, King Edward Memorial Hospital forWomen, 374 Bagot Rd, Subiaco, Perth, Western Australia 6008,Australia. E-mail: [email protected]
Mortality and morbidities such as ne-crotizing enterocolitis (NEC), late-onset sepsis (LOS), and feeding diffi-culties as a result of immature bowelfunction are a major issue in preterm,especially extremely preterm (�28weeks’ gestation) neonates world-wide. Probiotics may prevent NEC bypromoting colonization of the gut withbeneficial organisms, preventing colo-nization by pathogens, improving thematurity and function of gut mucosalbarrier, and modulating the immunesystem (eg, TLR4 receptor, nuclearfactor-�B, inflammatory cytokines) tothe advantage of the host.1,2 Many clin-ical trials have evaluated the safetyand benefits of probiotic supplementa-tion in preterm very low birth weight(VLBW) neonates. Deshpande et al3
first reported a systematic review ofrandomized, controlled trials (RCTs) ofprobiotic supplementation in pretermVLBW neonates. The results of theirsystematic review and meta-analysiswere based on 7 trials that involved1393 preterm VLBW neonates with ges-tation �33 weeks. The risk for all-cause mortality and NEC was reducedby 53% and 64%, respectively, in neo-nates who received probiotic supple-mentation compared with controlgroup neonates. The time to achievefull milk feeds was also significantlyless (by an average of �3 days) inthose who received probiotic supple-mentation.3 These significant resultswere subsequently confirmed by 2more systematic reviews that indi-cated the tremendous potential of pro-biotic supplementation in savingpreterm neonates from death and dis-ease.4,5 Expert bodies such as the Co-chrane Neonatal Review Group (CNRG)have concluded that except for thosewhoweigh�1000 g (because of lack ofspecific data in this high-risk popula-tion), a change in practice is sup-ported by the data.5 Individual expertshave also commented that on the basisof current data, those who wish to of-
fer probiotic supplementation as aroutine therapy for preterm neonatescannot be faulted.6 Subsequent tothese systematic reviews, 4 more RCTs(including 1 multicenter trial) that in-volved 783 preterm neonates havebeen reported.7–10 Given the global bur-den related to death, NEC, LOS, andfeeding difficulties in preterm VLBWneonates and the reported very signif-icant benefits of this low-cost, simple,and easily available intervention, weaimed to update our systematic re-view3 of probiotic supplementation ofpreterm VLBW neonates and studythe implications of its results tothe current clinical practice andresearch.11–13
METHODS
Guidelines from CNRG,14 Centre for Re-views and Dissemination,15 and theQUOROM statement were followed forthis systematic review and meta-analysis.16 The following prespecifiedcriteria, similar to our previous sys-tematic review, justified inclusion ofany trial in the analysis: (1) RCT involv-ing preterm VLBW neonates (�34weeks’ gestation and birth weight�1500 g) and reporting on stage 2 NECor higher (Modified Bell staging crite-ria)17,18 and (2) enteral administrationof any probiotic commencedwithin thefirst 10 days of life and continued for atleast 7 days. The details of the searchstrategy and approach to analysis aregiven in the Appendix, which is pub-lished as supplemental information atwww.pediatrics.org/content/full/125/5/921.
The primary outcome was efficacy ofprobiotic supplement in preventingstage 2 NEC or higher, safety in termsof blood culture–positive sepsis in-cluding that by the organism(s) in theprobiotic supplement, and any otheradverse events reported by the au-thors. The secondary outcomes in-cluded the time to reach full feeds (TFF;
120–150 mL/kg per day enteral feedsor as per the prestated definitions byauthors) and duration of hospital stay.
Trial sequential analysis (TSA) was re-cently reported as a useful tool to es-tablish when firm evidence is reachedin a cumulative meta-analysis.19 Wetherefore conducted TSA to evaluatewhether the findings of our updatedmeta-analysis are conclusive.2,20,21 Cu-mulative z curves, information size,and sequential monitoring boundarieswere estimated on the basis of risk fortype I error of 5% and type II error of20%. An intervention effect of 30% forthe prevention of NEC was regarded asclinically significant. Moderate hetero-geneity was assumed, and a heteroge-neity correction factor of 1.33 was ap-plied. Cumulative z curve of eachcumulative meta-analysis was calcu-lated with a fixed-effect and a random-effect model. The monitoring bound-aries were calculated by using themethod reported by Reboussin et al.22
RESULTS
A total of 38 potentially relevant cita-tions were obtained by our searchmethod. An additional citation wasidentified by using text words andsearching related articles.10 The selec-tion process details are given in Fig 1.Four new trials were finally included inthe updated analysis after extractionof data from the publications,7–10 andreceipt of additional data for pretermVLBW neonates who were �34weeks’ gestation.7 One trial (N � 20)was subsequently excluded becauseof unavailability of necessary datafrom the author.25 Search of otherdatabases mentioned already did notidentify any additional eligible stud-ies. Characteristics of the 11 trials (4new and the 7 from our previous re-port3) that were included in the anal-ysis (N � 2176) are summarized inTable 1.7–10,27–33 The quality of the tri-als assessed by Jadad score is sum-
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marized in Table 2.34 The results ofquality assessment were similar byusing CNRG guidelines.
Effect of Probiotics on Stage 2 orHigher (Definite) NEC
Data on definite NEC were reportedby all 11 trials involving 2176 neo-nates7–10,27–33 (Fig 2). A higher propor-tion of neonates in the control group(no probiotics) developed definite NECcompared with the probiotics group(71 [6.56%] of 1082 vs 26 [2.37%] of1094). Meta-analysis of data by using afixed-effects model estimated a lowerrisk (relative risk [RR]: 0.35 [95% con-fidence interval (CI): 0.23–0.55]; P �.00001) of NEC in the probiotic group.There was no significant heterogeneity(I2 � 0%, P � .57) among the trials.Individually, only 4 trials reported sig-nificantly higher risk for NEC in thecontrol group.8,9,29,30 The numbersneeded to treat (NNT) with probioticsto prevent 1 case of NEC was 25 (95%CI: 17–34).
Effect of Probiotics on BloodCulture–Positive Sepsis
Meta-analysis of data from10 trials7–10,28–33
(N � 2138) estimated no significantdifference in the risk for sepsis be-tween the probiotics and controlgroup neonates (RR: 0.98 [95% CI:0.81–1.18] P � .80; Fig 3); however,there was significant heterogeneity(I2� 52.1%, P� .03) among the trials.Only 1 trial reported significantly lowerrisk for sepsis in the probiotic group,7
and another reported higher risk forsepsis in the probiotic group, whichwas not significant after adjustmentfor gestation and birth weight.6
Effect of Probiotics on Mortality
Pooling of data from 9 trials,7–10,28–30,32,33
(N� 2051) showed a reduced risk fordeath from all causes in the probioticversus the control group (RR: 0.42[95% CI: 0.29–0.62]; P� .00001; Fig 4).No significant heterogeneity was notedbetween the trials (I2 � 0%, P � .86).The NNT to prevent 1 death from all
causes by treatment with probioticswas 20 (95% CI: 14–34). Pooling of data(N � 1335) from 5 trials,8,28,30,32,33
showed no significant difference in therisk for mortality as a result of NEC inthe probiotic versus the control group(RR: 0.30 [95% CI: 0.08–1.08]). Therewas no significant heterogeneity amongthese trials (I2� 0%, P� 0.53).
Effect of Probiotics on TFF
Meta-analysis of data (N� 936) from 5trials 8,9,30,32,33 showed significant re-duction in TFF in the probiotic versusthe control group (weighted mean dif-ference:�5.03 days [95% CI:�5.62 to�4.44]; P � .0001). There was signifi-cant heterogeneity (I2 � 83.3%, P �.0001) among the trials. This differencewas not significant after using therandom-effectsmodel (weightedmeandifference:�2.39 days [95% CI:�5.53to 0.75]; P� .14).
Sensitivity Analysis
Only 5 of 11 included trials had a pri-mary outcome of NEC or of death andNEC.8,9,28–30 A sensitivity analysis ofthese 5 trials (N � 1717) showed sig-nificant reduction of NEC in the probi-otic group (0.29 [95% CI: 0.17–0.49]; P� .00001) and reduction in mortali-ty(0.39 [95% CI: 0.25–0.59]; P �.00001). There was no heterogeneityamong all 5 trials (I2 � 0%, P �.71).8,9,28–30 Four of these 5 trials withprimary outcome of NEC or of deathand NEC showed significant reductionof NEC in the probiotic group8,9,29,30;however, no difference in sepsis wasnoted. All trials included in the meta-analysis had Jadad quality score �3(Table 2). A sensitivity analysis basedon Jadad score �3 vs �3 thereforewas not required. The roughly symmet-rical funnel plot (Fig 5) suggests thatpublication bias was unlikely. All re-sults except TFF remained similar byusing the random-effects model.
1 Excluded due to unavailability of relevant data after contacting authors 25
1 Reporting additional data from previous paper 26
Number of studies included in the Meta analysis: 4 7-10
Potentially relevant citations identified after screening of the electronic search (n=38)
31 Articles excluded: study population not preterm VLBW infants or the intervention was not a probiotic
1Review article23
1Not a RCT 24
7 Articles retrieved for more information 1 Article retrieved by using text words and searching related articles 10
FIGURE 1Flow diagram of the study selection process after screening of electronic search.
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TSA results showed evidence to sup-port a 30% reduction in the incidenceof NEC (� � .05; power: 80%; Fig 6). Thecumulative z curve crossed the moni-toring boundary. The conclusion wasunchanged when � � .01 was used.Adjusting for 1 interim analysis (firstmeta-analysis)3 did not change theconclusion, because the boundary wasstill crossed. The results were similarfor random- and fixed-effect models.
DISCUSSION
The results of our update confirmthose of the previous systematic re-views while improving their precisionand further reducing the role ofchance alone. The dramatic benefits interms of reduced risk for all-causemortality and definite NEC are sus-tained; however, despite the additionof 4 new trials (N� 783) to the existing
data, there is still no evidence that pro-biotic supplementation reduces therisk for LOS.
The incidence of NEC (5%–6% of VLBWneonates) has not changed signifi-cantly despite advances in neonatal in-tensive care.12 Definite NEC (stage 2 orhigher) continues to be a potentiallydisastrous illness in preterm neo-nates, with significant mortality (atleast 20%–25%) and morbidity, includ-ing need for surgery and survival withshort bowel syndrome and its conse-quences such as recurrent sepsis anddependence on total parenteral nutri-tion (TPN).12,35 Surgical NEC is associ-ated with prolonged (�6 months)hospitalization and long-term neuro-developmental impairment (NDI).36–38
Results of a systematic review of ob-servational studies indicated that therisk for long-term NDI was significantlyhigher in the presence of at least stage
2 NEC versus no NEC (odds ratio: 1.82[95% CI: 1.46–2.27]) in preterm VLBWneonates. Those who required surgerywere at higher risk for NDI than werethose who were treated medically(odds ratio: 1.99 [95% CI: 1.26–3.14]).38
On the basis of the length of stay, theestimated annual additional hospitalcharges for NEC have been reported tobe as high as $216 666 per survivor inthe United States.39 Given the signifi-cant burden of NEC, the benefits of pro-biotics in this area are extremelysignificant.
Feeding difficulties that lead to pro-longed deprivation of enteral feedsand dependence on TPN are a majorissue in extremely preterm neonates.The lack of scientific guidelines for de-fining and managing signs of “feed in-tolerance” and the fear of NEC are as-sociated with frequent stoppage ofenteral feeds in this high-risk popula-
TABLE 1 Characteristics of Trials Included in the Analysis
Source Birth Weight/GA Probiotic Agent/s Dosage and Duration Type of Milk Primary Outcome
Kitajima et al,33 1997 �1500 g BB 0.5� 109 organisms once dailyfrom first feed for 28 d
MM or FM Gut colonization by BB
Dani et al,28 2002 �33 wk or�1500 g LB-GG (Dicloflor) 6� 109 CFU once daily fromfirst feed until discharge
MM, DM, or FM UTI, sepsis, NEC
Costalos et al,31 2003 28–32 wk SB 109/kg twice daily from firstfeed for 30 d
FM Gut function and stoolcolonization
Bin Nun et al,30 2005 �1500 g BI, ST, BBB BI 0.35� 109 CFU, ST 0.35�109 CFU, and BBB 0.35� 109
CFU once daily from firstfeed to 36 wk corrected age
MM or FM NEC
Lin et al,29 2005 �1500 g LB-A, BI LB-A 1004356 and BI 1015697organisms twice daily fromday 7 until discharge
MM or DM NEC or death
Manzoni et al,32 2006 �1500 g LB-C (Dicloflor) 6� 109 CFU once daily fromday 3 of life to 6 wk ordischarge from NICU
MM or DM Gut colonization by Candiaspecies
Mohan et al,27 2006 �37 wka BB-L 1.6� 109 CFU once daily fromday 1 to day 3 4.8� 109 CFUonce daily from day 4 today 21
FM Gut colonization by BB-Land enteric pathogens
Stratiki et al,7 2007 27 to 37 wka BB-L Preterm formula 1� 107 CFU/gstarted within 48 h to 30 d
FM Intestinal permeability
Lin et al,8 2008 �34 wk and�1500 g BBB, LB-A 2� 109 CFU/d for 6 wk MM or FM NEC or deathSamanta et al,9 2009 �34 wk and�1500 g BBB, BB-L, BI, LB-A 2.5� 109 CFU/d until discharge MM or FM NEC, TFF, sepsis, death, and
hospital stayRougé et al,10 2009 �32 wk and�1500 g BB-LG, LB GG 1� 108 CFU/d until discharge MM, DM, or FM Enteral feed intake at
day 14
GA indicates gestational age; BB, Bifidobacterium breve; LB GG, Lactobacillus GG; SB, Saccharomyces boulardii; BI, Bifidobacteria infantis; ST, Streptococcus thermophilus; BBB, Bifidobac-terium bifidus; LB-A, Lactobacillus acidophilus; LB-C, Lactobacillus casei; BB-L, Bifidobacterium lactis; BB-LG, Bifidobacterium longum; CFU, colony-forming units; MM, mother’s milk; DM,donor milk; FM, formula milk; UTI, urinary tract infection.a Data for�34 weeks and�1500 g obtained by contacting the authors.
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tion.13 Despite the aggressive ap-proach to enteral and parental nutri-tion, postnatal growth restrictioncontinues to remain a significant issuein this population.40,41 Given the impor-tance of optimal enteral nutrition inearly postnatal life, the benefits of pro-biotics (eg, improved gastric emptyingand gut barrier function, reduction inTFF) have significant implications inimproving the overall prognosis of thishigh-risk population.7,25 The change inresults, from significant (fixed-effectsmodel) to no significant (random-effects model) reduction in TFF, mayrelate to significant heterogeneity inthe feeding protocols in various trials.
Given the lack of specific data on neo-nates who are fed exclusively breastmilk/formula/mixed milk feeds, evalu-ation of the benefits of probiotics in thepresence of different types of milkfeeds is difficult. It is important to notethat despite its significant benefits,preferential use of breast milk alonehas not eliminated the risk for NEC sig-nificantly in preterm VLBW neonates.Although specific data are not avail-able from all trials included in our sys-tematic review, the reduction in all-cause mortality may relate to thesignificant reduction in the incidenceof definite NEC and possibly TFF andsevere sepsis, leading to an improve-ment in the general well-being.8,42
The reasons for the lack of reduction inthe risk for LOS need to be discussed.Colonization of the gut by aberrantflora and its translocation play an im-portant role in LOS in preterm neo-nates.11,43 Probiotic microorganismsare expected to colonize the gut, com-pete with pathogens, improve the gutbarrier function and permeability, andmodulate immune function.7,25 The gas-trointestinal tract is reported to be themain reservoir of coagulase-negativeStaphylococcus (CONS), the most fre-quent organism responsible for LOS inextremely preterm neonates.44 The in-
ability of probiotics alone to overcomethe burden of LOS may thus relate tothe presence of not only a single (gut)but also multiple (eg, endotrachealtubes, central venous catheters, TPNsolutions, lipid infusions) sources ofvarious pathogens (CONS, Gram-negative, fungi) in the presence of fre-quent exposure to broad-spectrum an-tibiotics, prolonged deprivation ofenteral nutrition, and an immature im-mune system in this high-risk popula-tion.8 Mortality from CONS sepsis islow, whereas that related to virulentpathogens (eg, Gram-negative organ-isms) is high.44–46 It is not knownwhether the immunomodulating ef-fects of probiotics are different inCONS versus non-CONS organisms.47
Benefits of probioticsmay thus dependon the type of microorganisms respon-sible for LOS and, as with any interven-tion, on the baseline incidence of LOS invarious settings. Although probioticsepsis has been reported in immuno-compromised hosts and neonates,48–50
it is reassuring to know that no signif-icant adverse events, especially probi-otic sepsis, have been reported in anyof the trials included in our analysisdespite the diversity of the populationsand the settings of the trials. Neverthe-less, we emphasize the need for care-ful surveillance not only for probioticsepsis but also for the development ofantibiotic resistance and altered im-mune responses in the long-term.51,52
Although we do not have the specificdata to support this, the risk for trans-location of probiotic bacteria across acompromised gut barrier followed bysepsis may be higher in critically sickand/or extremely low birth weight(ELBW) neonates.
Overall, the results of our updated sys-tematic review and meta-analysis (11good-quality RCTs and N � 2176) con-firm the dramatic benefits of probioticsupplements in reducing the risk fordeath and for definite NEC in pretermTA
BLE2JadadScoreforAssessmentofTrialQuality
Parameter
Kitajimaetal,33
1997
Danietal,28
2002
Costalosetal,31
2003
BinNunetal,30
2005
Linetal,29
2005
Manzonietal,32
2006
Mohanetal,27
2006
Stratikietal,7
2007
Linetal,8
2008
Samantaetal,9
2009
Rougéetal,10
2009
1.Randomization
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
A.Methodtogeneraterandomization
wasclearandappropriate
Yes
Yes
Yes
NAYes
Yes
Yes
Yes
Yes
Yes
Yes
2.Doubleblind?
NoYes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
A.Wasmethodforblinding
appropriate?
N/A
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
NAYes
3.Descriptionofwithdrawalordropout
Yes
NoYes
NoNo
NoNo
Yes
Yes
NoYes
Totalscore
34
53
44
45
53
5
Yes
�1point;No
�0points;scores:0
�worst,5
�best.NAindicatesnotavailable;N/A,notapplicable.
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VLBW neonates. There is no evidence ofa reduced risk for LOS. The significanteffect size, precision, consistency ofthe results across all trials, extremelylow P values almost ruling out the roleof chance alone, low risk for publica-tion bias, no statistical heterogeneity,critical areas of benefit, and the TSAconclusive of at least 30% reduction inthe incidence of NEC all indicate thatwithholding probiotics from high-riskneonates is now almost unethical.53
Our findings will have a significant im-pact on recruitment in the current/
planned placebo-controlled trials ofprobiotics in preterm neonates, be-cause parents have the right to com-plete and up-to-date information onthis topic in a transparent manner. Onthe basis of our results, we believe thatit will be very difficult to justify theneed for additional placebo-controlledtrials in this population given the sig-nificant reduction in definite NEC andall-cause mortality. Moreover, giventhe sample size and power (Table 3)of the ongoing/planned placebo-controlled trials, it is unlikely that their
individual or cumulative results will af-fect our results significantly. We antic-ipate that the lack of specific data onextremely preterm (�28 weeks’ gesta-tion) neonates and on long-term out-comes and the need to reproducethese results in a setup with possibly ahigher standard of care and lowerbaseline incidence of death and defi-nite NEC could still be pushed forwardas the basis for more placebo-controlled trials; however, the data re-ported by Satoh et al42 indicate thesafety and efficacy of probiotics in ex-
FIGURE 2Effect of probiotics on NEC.
FIGURE 3Effect of probiotics on blood culture–positive sepsis.
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tremely preterm neonates. This obser-vational study from Tokyo comparedthe data before and after introducingprobiotics as a routine therapy in pre-term neonates (epoch I [1994–1998,no probiotic]: ELBW � 101 of 226; ep-och II [1999–2003, supplementationwith Bifidobacteria]: ELBW � 220 of338). Bifidobacteria breve (1 billion)supplementation was mixed in milk/formula and started as early as 7.2hours of age and continued until dis-charge (37 weeks or 2.3 kg). There wassignificant reduction in the incidenceof NEC, sepsis, and sepsis in death in
epoch II versus I (NEC: 6 [2.6%] vs 0[0%]; sepsis: 65 [28.8%] vs 70 [20.7%];sepsis in death: 9 [13.8%] vs 2 [0.6%]).The significant benefit in NEC occurreddespite the low baseline incidence ofthe condition. Manzoni et al54 also re-cently reported the safety of routineuse of probiotics in VLBW neonates(N � 743; mean birth weight: 1056 �88 g; mean gestation: 29.5 � 1.1weeks) during a 6-year period.
Given that probiotics reduce all-cause mortality significantly, it is im-portant to know whether this benefit
comes at the cost of an increasednumber of survivors with long-termNDI. Chou et al55 recently reportedthe long-term neurodevelopmentaloutcomes of neonates (�32 weeks’gestation) in their RCT of oral probi-otics for NEC. A total of 83.1% of neo-nates (probiotics: 153; placebo: 148)from their trial were assessed byBayley infant developmental assess-ment tool (BSID-II) at 24 months’ cor-rected age; 1 of 153 and 4 of 148 haddied after discharge. There were nosignificant differences in growth(head circumference, length, andweight), cerebral palsy, blindness,deafness, Mental Developmental In-dex (�70), and Psychomotor Devel-opmental Index (�70). Given the im-portance of this issue, it is criticalthat authors of all trials in this areareport long-term neurodevelopmen-tal outcomes of the enrolled neo-nates. Definite NEC and sepsis bothare associated with higher risk forlong-term NDI in preterm VLBW neo-nates.38,56 Given the significant reduc-tion in definite NEC and possiblysevere sepsis8 after probiotic sup-plementation, it is difficult to hypoth-esize that probiotic exposure in earlypostnatal life will be associated withlong-term NDI in preterm neonates. A
FIGURE 4Effect of probiotics on all-cause mortality.
FIGURE 5Funnel plot.
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significantly large sample size (Table4) would be required to documentthe smallest but clinically significantbenefit in centers with higher stan-dards of care and lower baseline in-cidence of definite NEC and death.The potentially preventable numberof deaths and cases of NEC in the pla-cebo arm will be an ethically chal-lenging issue in conducting such tri-als while ignoring the totality ofevidence.
CONCLUSIONS
Considering the robustness of the ev-idence provided and the very signifi-
cant benefits in critical areas thatoutweigh the potential adverse ef-fects, we believe that probioticsshould now be offered as a routinetherapy for preterm neonates andthat additional placebo-controlledtrials are not warranted; however,selection of a safe and suitable prod-uct with documented probiotic prop-erties and close monitoring of thetarget population is a must beforeoffering this therapy as a routine inthis high-risk but most deservingpopulation.57,58 Consistent benefitsdespite significant variations in pro-
biotic strains and protocols indicatethat probiotics “in general” are ben-eficial in this high-risk population inthe context of the broader perspec-tive of meta-analysis.59 It is impor-tant to note that the effect of a pro-biotic bacterium is strain-specificand cannot be extrapolated even toother strains of the same species.60
Other important but as yet unan-swered questions (eg, product/strain(s), dosage, duration, practi-calities of administration) couldeasily be addressed by well-designedand tightly controlled prospective,observational studies or head-on tri-als of various strains/combinations/dosages/protocols etc in collabora-tion with the industry and regulatoryagencies.2,61 Rigorous evaluation ofan available and potentially suitableproduct that has not been tested inthis high-risk population may possi-bly be the only role for additional
FIGURE 6Trial sequential analysis.
TABLE 3 Characteristics of Ongoing Placebo-controlled Trials of Probiotics
Study Primary Outcome Inclusion Criteria EstimatedCompletion
Sample Size Effect Size Power
Braga et al (Brazil)a62
ISRCTN67165178NEC 750–1500 g Completed 630 NA NA
Costeloe et al (UK)63
ISRCTN05511098Sepsis, NEC, or death �31 wk 2013 1300 NA NA
Lozano et al (Colombia)64
NCT00727363Death or neonatal sepsis �2000 g 2011 1110 NA NA
Tobin et al (Australia)65
ACTRN12607000144415LOS �32 wk NA 1100 33% 90%
NA indicates not available.a Completed recruitment results awaited.
TABLE 4 Estimated Sample Sizes for Various Primary Outcomes in ELBW Neonates
placebo-controlled trials in thisarea. Current evidence makes it un-likely that parents would opt for a
50% chance of their infant’s being al-located to a placebo if a suitable pro-biotic product were available.
ACKNOWLEDGMENTWe thank Dr Kristian Thorland for ad-vice on trial sequential analysis.
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