J. NEU, NEJM 2011

L’entérocolite ulcéro-nécrosante : point de vue de l’épidémiologiste

J. NEU, NEJM 2011

American Journal of Clinical Nutrition, 2017 * soutien de Nestlé pour le recueil des selles

EPIFLORE* (ANR) CLOSNEC (ANR)

Included preterm infants with gestational age between 24 and 31weeks hospitalized in NICU

n= 3658

Preterm infants alive at day 7 with known information on Necrotizing enterocolitis

N=3151

Without information about Necrotizing enterocolitis

N=59

Infants with Necrotizing enterocolitis

> stade II of Bell

N=106

With spontaneous intestinal perforation

N=54

Deceased before day 7 N=167 dont 3 NEC et 2 perforations

32 without information

Preterm infants alive at day 7

n= 3491

Infants without Necrotizing enterocolitis

> stade II of Bell

N= 3045

Preterm infants alive at day 7 with known information on Necrotizing enterocolitis

N=3378

Hospitalized in small units with less 10 infants enrolled in the EPIPAGE II cohort N=227

Results (1) Description of cases and risk factors

• 106 necrotizing enterocolitis – 62 Bell stage II

– 44 Bell stage III whose 20 treated by surgical intervention.

– Rate of NEC > Bell stage II according to gestational age: • 24-26 weeks : 4.7 % ( 3.3 – 6.7) p<0.001

• 27-29 weeks : 4.0 % ( 3.0 – 5.2)

• 30-31 weeks : 4.0 % ( 1.5 – 3.1)

Nom

brede

CAS

0

5

10

15

20

25

30

W2 W3 W4 W5 W6 W7 W8 W9 W10W11W12>W12

Ageaumomentdel’ECUN(semaines)Post natal age at time of NEC (weeks)

Nombrede

CAS

Agepostmenstruelaumomentdel’ECUN(semaines)

0

2

4

6

8

10

12

14

16

18

27 28 29 30 31 32 33 34 35 36 37 39

Nu

mb

er o

f ca

ses

of

of

NEC

Post menstrual age at time of NEC (weeks)

IMMATURITE

GENETIC

ISCHEMIE / ANOXIE

Risk Factors for NEC, at birth

N ORa 95% Confidence

interval p

GA: 30-31 weeks 1295 1

GA: 27-29 weeks 1224 2.0 1.2 3.2 0.002

GA: 24-26 weeks 632 2.4 1.401 4.181 0.006

Birth weight Z-score 3151 0.8 0.621 0.909 0.003

Female 1499 0.7 0.494 1.091 0.126

APGAR score >6 at 5 min 2446 1

APGAR score not known at 5 min 168 0.6 0.2 1.8 0.430

APGAR score <7 at 5 min 537 1.7 1.1 2.6 0.028

Multiple pregnancy 1017 1.2 0.8 1.8 0.320

Risk Factors for NEC, during the first week n ORa 95% Confidence interval p

RDS 1.7 1.0 2.8 0.042

Hemodynamic treatment during first week • No Hemodynamic Treatment 2698 1

• volume expansion 64 5.7 2.4 13.5 0.001

• Volume expansion and corticosteroids 55 4,312 1.6 12.0 0.005

• Volume expansion and catecholamines 118 11.1 6.1 20.1 0.001

• Catecholamines 129 1.2 0.4 3.4 0.779

• Corticosteroids 13 2.9 0.3 24.7 0.337

• Corticosteroids and catecholamines 53 5.9 2.3 15.0 0.001

Sedation/analgesia during the first week • No sedation/analgesia treatment 2097 1

• Morphin 199 0.4 0.2 0.9 0,024

• Fentanyl 146 1.0 0.4 2.4 0,956

• Surfentanyl 562 0.5 0.3 0.9 0,022

• Ketamine 35 1.5 0.3 6.8 0,640

• Midazolam 89 0.5 0.1 2.2 0,383

• Other treatments* 23 6.4 1.9 21.6 0,003

Instestinal transit during first week

• Good transit 1923 1 • Not known 197 0,22 0,03 1,633 0,139

• Bad transit 1031 2,137 1,382 3,302 0,001

Risk Factors for NEC, during the first week n ORa 95% Confidence interval p

RDS 1.7 1.0 2.8 0.042

Hemodynamic treatment during first week • No Hemodynamic Treatment 2698 1

• volume expansion 64 5.7 2.4 13.5 0.001

• Volume expansion and corticosteroids 55 4,312 1.6 12.0 0.005

• Volume expansion and catecholamines 118 11.1 6.1 20.1 0.001

• Catecholamines 129 1.2 0.4 3.4 0.779

• Corticosteroids 13 2.9 0.3 24.7 0.337

• Corticosteroids and catecholamines 53 5.9 2.3 15.0 0.001

Sedation/analgesia during the first week • No sedation/analgesia treatment 2097 1

• Morphin 199 0.4 0.2 0.9 0,024

• Fentanyl 146 1.0 0.4 2.4 0,956

• Surfentanyl 562 0.5 0.3 0.9 0,022

• Ketamine 35 1.5 0.3 6.8 0,640

• Midazolam 89 0.5 0.1 2.2 0,383

• Other treatments* 23 6.4 1.9 21.6 0,003

Instestinal transit during first week

• Good transit 1923 1 • Not known 197 0,22 0,03 1,633 0,139

• Bad transit 1031 2,137 1,382 3,302 0,001

CONCEPT du deuxiéme cerveau

MICROBIOTE

IMMATURITE

GENETIC

PRATIQUES DES UNITES

ISCHEMIE / ANOXIE

Risk Factors for NEC, during the first week n ORa 95% Confidence interval p

RDS 1.7 1.0 2.8 0.042

Hemodynamic treatment during first week • No Hemodynamic Treatment 2698 1

• volume expansion 64 5.7 2.4 13.5 0.001

• Volume expansion and corticosteroids 55 4,312 1.6 12.0 0.005

• Volume expansion and catecholamines 118 11.1 6.1 20.1 0.001

• Catecholamines 129 1.2 0.4 3.4 0.779

• Corticosteroids 13 2.9 0.3 24.7 0.337

• Corticosteroids and catecholamines 53 5.9 2.3 15.0 0.001

Sedation/analgesia during the first week • No sedation/analgesia treatment 2097 1

• Morphin 199 0.4 0.2 0.9 0,024

• Fentanyl 146 1.0 0.4 2.4 0,956

• Surfentanyl 562 0.5 0.3 0.9 0,022

• Ketamine 35 1.5 0.3 6.8 0,640

• Midazolam 89 0.5 0.1 2.2 0,383

• Other treatments* 23 6.4 1.9 21.6 0,003

Instestinal transit during first week

• Good transit 1923 1 • Not known 197 0,22 0,03 1,633 0,139

• Bad transit 1031 2,137 1,382 3,302 0,001

*Others analgesia/ treatment

• 7 infants received analgesia by sevofluran (Inhalational anaesthetic ) during the first week (day of birth =2, day 1=2,day2=4). All theses infants were hospitalized in one NICU

• 4/7 developped NEC, 1 during Week2, 1 during Week3, 1 during week4, 1 during Week5.

• Adjusted OR= 24.3 (4.1-145)

*Others analgesia/ treatment

• 65 infants in live at day7 were hospitalized in this unit using sevofluran :

– 24-26weeks : n=2 NEC / 8 infants 1NEC/ 2 sevofluran

– 27-29weeks : n=8 NEC / 34 infants 3NEC/ 5 sevofluran

– 30-31weeks : n=4 NEC /23 infants No sevofluran

10 NEC/48 ( 17.2%) infants without sevofluran versus 4 NEC/7 (57.1% ) infants receving Sevofluran before Day7

P=0.034 Fisher exact test

MICROBIOTE

IMATURITE

GENETICc

ISCHEMIE / ANOXIE

PRATIQUES DES UNITES

n= number of infants, N= number of neonatal intensive care units (NICUs)

24-26 weeks` 27-29 weeks 30-31 weeks

n= number of infants, N= number of neonatal intensive care units (NICUs)

24-26 weeks` 27-29 weeks 30-31 weeks

MICROBIOTE

IMMATURITE

ISCHEMIE / ANOXIE

PRATIQUES DES UNITES

Phyla composition between NEC and healthy preterm infants

0

25

50

75

100

App1 App2 App3 NEC

Bacteroidetes

Actinobacteria

Proteobacteria

Firmicutes

Healthy

NEC NEC

Firmicutes

Proteobacteria

Actinobacteria

Bacteroidetes *

META ANALYSE

Differential bacterial species (delta % are presented (% in NEC - % in healthy controls)

-6 -4 -2 0 2 4 6

Clostridium butyricum; YE48; AY442812

marine bacterium CS-2; EF040537

Clostridium neonatale; LCDC no.99-A-005; AF275949

bacterium NLAE-zl-C493; JQ608194

Morganella morganii; SSCT63; AB210972

Gram-negative bacterium cL10-2b-4; AY239469

Gram-negative bacterium cTPY-13; AY239461

Staphylococcus haemolyticus JCSC1435; AP006716

Enterococcus sp. AK61; AY098492

Proteus sp. VBC3; JX089650

Escherichia coli; AK108; AY098487

Bifidobacterium breve; JCM 1273; AF491832

Peptoniphilus gorbachii (T); WAL 10418; DQ911241

Streptococcus agalactiae; GY102; HQ658086

Citrobacter sp. DW38; AJ534867

Desulfotomaculum guttoideum (T); DSM 4024; Y11568

Klebsiella sp. YHB; AY517552

Salmonella sp. AHL 6; AY379978

Haemophilus parainfluenzae (T); CCUG 12836; AY362908

Klebsiella sp. D-D; DQ278420

Enterococcus faecalis; Y18293

Staphylococcus sp. CMJ1-7; KC119123

Delta % NEC vs Healthy

Genus

Species

CLOSNEC : analyse des phenotypes des Clostridium Neonatale et Butyricum

Sophia Schönherr-Hellec, Julio Aires dans le laboratoire de MJ Butel

MICROBIOTE

IMMATURITE

GENETIC

PRATIQUES DES UNITES

ISCHEMIE / ANOXIE

?

EPIFLORE

• Analyses microbiotes faites 458 -> 6 clusters?

• Relation

NICU

PRATIQUES • Allaitement • Peau à peau • Ventilation

M I C R O B I O T E

Hôte • AG • Z score Poids • Transit

MICROBIOTE

IMMATURITE

GENETIC

ISCHEMIE / ANOXIE

PRATIQUES DES UNITES

Defects in genes that inhibit intestinal innate immune signaling can contribute to NEC. Adequately powered studies are needed to study the impact of SIGIRR variants and variant–clinical variable interactions in the causation of NEC.

PEDIATRICS 2014

MICROBIOTE

GENETIC

PRATIQUES DES UNITES

BIOMARKER

IMMATURITE

ISCHEMIE / ANOXIE

NEU, NEONATALOGY 2013

PROTEINOME

NEU, NEONATALOGY 2013

MICROBIOTE

GENETIC

PRATIQUES DES UNITES

BIOMARKER

IMMATURITE

ISCHEMIE / ANOXIE

MERCI DE VOTRE ATTENTION

?

?

• les Firmicutes (on y trouve notamment les genres : Ruminococcus, Clostridium, Lactobacillus (dont plusieurs souches utilisées comme probiotiques), et des Eubacterium, Faecalibacterium et Roseburia (productrices de butyrate) ;

• les Bacteroidetes (dans ce groupe, les Bacteroides, Prevotella et Xylanibacter dégradent une grande variété de molécules complexes de glycanes) ;

• les Actinobacteria (ce groupe inclut les genres Collinsella et Bifidobacterium (dont certaines souches de probiotiques connus) ;

• les Proteobacteria (dont communément des Escherichia (de la famille des Enterobacteriaceae et des bactéries du groupe Desulfovibrio (bactéries réductrices de soufre) ;

ISCHEMIE / ANOXIE

Altered microvasculature responses Inflamation

VEGF

Altered metabolome Altered metabolome

NEONATALOGY 2013

PROTEINOME

NEC Cases

Control matched on first propensity score (not including nutritional and antibiotic strategy) Control matched on second propensity score and hospitalizes in the same NICU Control matched on third propensity ( including nutritional and antibiotic strategy)

P<0.001

P<0.001

Neonatal characteristics Cases n=15 Controls=53 p Gestational age (weeks) 27.3 1.8 28.2 1.7 0.08

Post natal age (days) 27.9 17.8 24.8 15.0 0.50

Birth weight Z-score 0.16 1.1 -0.44 1.0 0.045

Bad intestinal transit during first 7 days 11 73.3 25 47.2 0.07

APGAR score <7 at 5 minutes 3 20.0 10 18.9 0.83

NICU profiles concerning enteral feeding

slower 2 13.3 8 15.0 0.85

intermediate 11 73.4 35 66.7

faster 2 13.3 10 19.3

NCIU breastfeeding policy

lesss favorable 3 20.0 11 20.8 0.74

intermediate 12 80.0 40 75.5

more favorable 0 0 2 3.8

Family composition between NEC and healthy preterm infants

Healthy

NEC

0

25

50

75

100

App1 App2 App3 NEC

Erysipelotrichaceae

Bacteroidaceae

Corynebacteriaceae

Lactobacillaceae

Peptostreptococcaceae

Bifidobacteriaceae

Clostridiales_Incertae Sedis XI

Porphyromonadaceae

Streptococcaceae

Lachnospiraceae

Pasteurellaceae

Clostridiaceae 1

Enterococcaceae

Staphylococcaceae

Enterobacteriaceae

Healthy Staphylococcus Enterococcus Klebsiella Clostridium sensu stricto Escherichia/ Shigella unclassified_Enterobacteriaceae Citrobacter Haemophilus Enterobacter Clostridium XlVa Streptococcus Peptoniphilus Bifidobacterium Morganella unclassified_Pasteurellaceae Clostridium XI Lactobacillus Barnesiella

Repartition of dominant genera in the overall cohort

IN NEC, increased proportions of Clostridium sensu stricto, Escherichia/Shigella and Morganella (p=0,054) genera but non significant.

Healthy NEC

Prevalence of selected genera

0

25

50

75

100

Stap

hyloco

ccus

Entero

cocc

us

Entero

bacte

riace

ae

Clostr

idium

sens

u str

icto

Bacte

roides

Bifid

obac

teriu

m

Lacto

bacil

lus

Cases Control matched on first propensity score (not including nutritional and antibiotic strategy) Control matched on first propensity score and hospitalizes in the same NICU Control matched on second propensity ( including nutritional and antibiotic strategy)

% P=0.06

Significantly differentially represented bacterial species

0

0,05

0,1

0,15

0,2

0,25

0,3

0,35

0,4

0,45

Esch

erich

ia coli C

FT07

3; A

E014

075

bacte

rium N

LAE-zl-

P257

; JQ60

6938

Stap

hyloco

ccus aureu

s; SU

-BIO

2; JN83

1369

swine

feca

l bac

teriu

m RF3

D-Cel1

; FJ7

5382

5

Bacte

roides sp

. Smar

lab 33

0299

6; A

Y643

081

Esch

erich

ia coli; AF0

7603

7

Stap

hyloco

ccus aureu

s 08B

A0217

6; C

P003

808

Proteu

s mira

bilis; M

1; H

Q25

9934

Esch

erich

ia coli; mothe

r C4;

10C

4; A

M15

7436

Esch

erich

ia co

li; W

D01; E

F620

925

Esch

erich

ia coli; mothe

r C3;

9C3;

AM15

7425

Stap

hyloco

ccus aureu

s; GSA

-44;

JN31

5148

alph

a pr

oteo

bacterium

PI_GH1.

1.B2

; AY1

6203

8

Stap

hyloco

ccus aureu

s; DM

H 7; H

M55

9237

Stap

hyloco

ccus epide

rmidis;

19N

3; H

Q28

4959

Sphin

gobium

yan

oiku

yae; W

1BR1

5; JF4

5993

2

Shigella dysen

teria

e; X

8068

0

Entero

bacter sp

. E4M

-P; G

Q47

8269

Stap

hyloco

ccus sp

. CUL-

LABL

G2X

2; H

Q84

1047

Stap

hyloco

ccus sp

. U13

69-1

0112

2-SW

168;

JQ08

2147

bacterium

OC18

(201

1); H

Q17

8988

App1 App2 App3 NEC

P<0,05 for all App1 +2 +3 versus NEC

App1 App2 App3 NEC

010

20

30

40

50

60

OTU denovo6512

Extrapolation of OTU J Neu (Mai et al, Plos One 2011) representation in our data

Sequencing V1-V3 for data J Neu while V3-V4 for us. 1- Blast OTU_Neu Klebsiella pneumoniae strain B16 (98% coverage 95%) 2- Get full 16S of K.pneumoniae 3- Blast all K. pneumoniae strain B16 full 16S against our OTU representatives 4- e-val 0.0 / 98.8% similarity with denovo6512

The OTU found significantly associated to NEC cases in Mai et al’s paper is not associated to our NEC cases