Intestinal microbiome changes in an infant with right atrial isomerism and recurrent necrotizing enterocolitis: A case report and review of literature

Table 1 Baseline characteristics of the patient

Characteristics	Description
CHD (according to echocardiography and CT results)	Right-sided heart. Double-inlet single ventricle. Common atrium. Malposition of the great arteries. Complete, unbalanced common atrioventricular valve. Right pulmonary artery stenosis. Double superior vena cava. Total, anomalous pulmonary venous return to the left superior vena cava (supracardiac type)
Noncardiac congenital anomalies (according to CT and abdominal ultrasound imaging)	Tracheal and esophageal displacement to the vertebral column. Tracheal narrowing to 4 mm × 2 mm at the level of the brachiocephalic trunk. Symmetrical liver. Asplenia. Common origin of the celiac trunk and superior mesenteric artery from the abdominal aorta. Right-sided aortic arch, right descending aorta

CHD: Congenital heart disease; CT: Computed tomography.

Table 2 Characteristics of cardiac surgery and the postoperative period

Characteristics	Values
Cardiopulmonary bypass duration (min)	105
Aortic cross-clamp during operation	Not performed
Mechanical ventilation duration after cardiac surgery (h)	95
Inotrope therapy duration (number of postoperative days)	4
Length of stay in the cardiac intensive care unit (d)	6
Length of stay after cardiac surgery (number of postoperative days)	55

Table 3 Resistive index of the visceral arteries according to Doppler ultrasonography

Parameters	Postoperative day (day of life)
	- (1)	1 (15)	5 (19)
		Course of the first episode of NEC stage IIA
RI AbdAo	> 1.0	> 1.0	1.17
RI celiac trunk	0.9	1.05	Not evaluated because patient exhibited anxiety
RI SMA	0.86	1.05
Conclusion	Holodiastolic flow reversal in AbdAo	Holodiastolic flow reversal in AbdAo, celiac trunk, SMA	Holodiastolic flow reversal in AbdAo

DOL: Day of life; POD: Postoperative day; RI: Resistive index; AbdAo: Abdominal aorta; SMA: Superior mesenteric artery; NEC: Necrotizing enterocolitis.

Table 4 Complete blood count and C-reactive protein values before cardiac surgery and during the first episode of necrotizing enterocolitis

Parameters	- (9)	Postoperative day (day of life)
		1 (15)	2 (16)	3 (17)		4 (18)	5 (19)	6 (20)	7 (21)
		Course of the first episode of NEC IIA
Hemoglobin, g/L	147	171	148		135	134	148	157	154
Red blood cells, 1012/L	3.95	5.54	4.85		4.39	4.38	4.83	5.19	5.11
Platelets, 109/L	559	154	144		152	158	208	263	289
White blood cells, 109/L	13.4	19.5	24.7		18.7	11.1	12.7	17.6	18.4
Normoblasts per 100 white blood cells					2	6		2
Metamyelocytes, %						1
Myelocytes, %								1
Stab neutrophils, %		11	9			1		2	2
Segmented neutrophils, %	25	66	80		79	50	40	35	41
Eosinophils, %	3					1	9	2	2
Basophils, %							1
Monocytes, %	16	13	3		7	10	13	11	26
Lymphocytes, %	53	10	8		14	37	37	49	30
С-reactive protein, mg/L	4	78	181		61	38	21	16	16

The reference range of C-reactive protein is 0-10 mg/L. NEC: Necrotizing enterocolitis.

Table 5 Time points of the microbiological examinations and fecal sampling for 16S rRNA gene-based microbiota profiling

Examinations	Postoperative day (day of life)
	- (2)	- (7)	- (10)	- (14)	5 (19)	8 (22)	18 (32)	21 (35)	29 (43)	35 (49)	40 (54)
16S rRNA gene-based microbiota profiling	-	+	-	+	-	+	-	+	+	-	-
Fecal culture	-	-	+	-	+	+	+	-	-	+	+
Gastric content culture	-	-	-	-	+	-	+	-	-	-	-
Pharynx smear culture	-	-	+	+	+	-	+	-	-	+	+
Blood culture	+	-	-	+	+	-	-	-	+	-	-

Table 6 Results of 16S rRNA gene-based fecal microbiota profiling in serial fecal samples of the neonate

POD (DOL)	Class level	Family level	Genus level
Before surgical treatment of CHD (DOL 7) – feeding with fortified BM with preterm formula added (because of low BM supply)
- (7)	Negativicutes 44.9%	Veillonellaceae 44.9%	Veillonella 44.9%
	Gammaproteobacteria 41.1%	Enterobacteriaceae 38.8% Pasteurellaceae 2.3%	Escherichia/Shigella 38.8%
	Bacilli 12.3%	Streptococcaceae 8.0% Enterococcaceae 4.3%	Streptococcus 8.0% Enterococcus 4.3%
Before surgical treatment of CHD (DOL 14) – before the first episode of NEC stage IIA on POD 1 (DOL 15) Feeding with fortified BM with preterm formula added (because of low BM supply)
- (14)	Clostridia 57.8%	Clostridiaceae 57.8%	Clostridium sensu stricto I 57.8% 1Clostridium paraputrificum 56.1%
	Gammaproteobacteria 29.6%	Enterobacteriaceae 24.8% Pasteurellaceae 4.8%	Escherichia/Shigella 24.8% Actinobacillus 4.8%
	Bacilli 5.7%	Streptococcaceae 5.7%	Streptococcus 5.7%
	Actinobacteria 3.8%	Bifidobacteriaceae 3.8%	Bifidobacterium 3.8%
	Negativicutes 3.1%	Veillonellaceae 3.1%	Veillonella 3.1%
POD 8 (DOL 22) – after ampicillin/sulbactam therapy (POD 1-7) and metronidazole therapy (POD 2-6) had been completed and piperacillin/tazobactam therapy was started (POD 7-14) (nil per os)
8 (22)	Gammaproteobacteria 99.8%	Enterobacteriaceae 99.8%	Escherichia/Shigella 99.8%
	Clostridia 0.2%	Ruminococcaceae 0.2%	1Ruminococcus bucirculans 0.2%
POD 21 (DOL 35) – 7 d after the antibacterial therapy was completed on POD 14 Feeding with hydrolyzed formula
21 (35)	Clostridia 58.0%	Clostridiaceae 56.9% Lachnospiraceae 1.1%	Clostridium sensu stricto I 56.9% Anaerostripes 1.1%
	Gammaproteobacteria 38.7%	Enterobacteriaceae 38.7%	Escherichia/Shigella 38.7%
	Bacilli 3.2%	Enterococcaceae 3.2%	Enterococcus 3.2%
POD 29 (DOL 43) – on the day after onset of the second episode of NEC IIA Feeding before NEC IIA onset: Expressed BM (from DOL 37) with hydrolyzed formula dotation
29 (43)	Clostridia 72.0%	Clostridiaceae 72.0%	Clostridium sensu stricto I 72.0%
	Gammaproteobacteria 22.7%	Enterobacteriaceae 19.8% Pasteurellaceae 2.9%	Escherichia/Shigella 18.1%
	Negativicutes 5.3%	Veillonellaceae 5.3%	Veillonella 5.3%

¹Species level.

POD: Postoperative day; DOL: Day of life; CHD: Congenital heart disease; BM: Breast milk; NEC: Necrotizing enterocolitis.

Table 7 Results of bacterial cultures and 16S rRNA gene-based fecal microbiota profiling

POD (DOL)	Fecal microbiome at the genus level, 16S rRNA	Fecal culture	Gastric culture	Pharynx smear culture	Blood culture	Antibacterial therapy
- (2)	-	-	-	-	No growth	No
- (7)	Escherichia/Shigella, Veillonella, Streptococcus, Enterococcus	-	-	-	-	No
- (10)	-	No growth	-	Staphylococcus hominis	-	No
- (14)	Clostridium sensu stricto I, Escherichia/Shigella, Streptococcus, Actinobacillus, Bifidobacterium, Veillonella	-	-	No growth	No growth	CUR: POD 0-1 (perioperative prophylaxis)
5 (19)	-	No growth	Enterococcus faecalis R: TET	Escherichia coli S: CUR, PIT, GEN R: AMS	No growth	NEC IIA AMS: POD 1-7; MET: POD 2-6 GEN+PIT: POD 7-14
8 (22)	Escherichia/Shigella, Ruminococcus	No growth	-	-	-
18 (32)	-	No growth	No growth	Streptococcus oralis	-	No
21 (35)	Clostridium sensu stricto I, Escherichia/Shigella, Enterococcus	-	-	-	-	No
29 (43)	Clostridium sensu stricto I, Escherichia/Shigella, Veillonella	-	-	-	No growth	NEC IIA No
35 (49)	-	No growth	-	Streptococcus mitis, Rothia mucilaginosa	-	No
40 (54)	-	No growth	-	No growth	-	No

“-“: Examination was not carried out; POD: Postoperative day; DOL: Day of life; S: Sensitivity; R: Resistance; NEC: Necrotizing enterocolitis; antibiotics (according to EUCAST, ver. 10.0, 2020): CUR: Cefuroxime; PIT: Piperacillin/tazobactam; GEN: Gentamicin; AMS: Ampicillin/sulbactam; TET: Tetracycline; MET: Metronidazole.

Table 8 Complete blood count and C-reactive protein values before and during the second episode of necrotizing enterocolitis

Parameters	Postoperative day (day of life)/day after the pericardial drainage procedure
	26 (40)	29 (43)	32 (46)	36 (50)	40 (54)/1	41 (55)/2	43 (57)/4
		Course of the second episode of NEC IIA
Hemoglobin, g/L	136	132	139	155	124	115	122
Red blood cells, 1012/L	4.6	4.4	4.6	5.2	4.1	3.8	4.1
Platelets, 109/L	461	554	139	331	216	141	236
White blood cells, 109/L	7.6	9.7	13.6	8.8	17.5	21.9	14.3
Stab neutrophils, %				1	1		1
Segmented neutrophils, %	27	60	20	19	55	50	28
Eosinophils, %	3	4	3	1		2	1
Basophils, %				1
Monocytes, %	13	8	10	11		9	7
Lymphocytes, %	57	28	67	67	39	39	63
С-reactive protein, mg/L	4	1	NA	1	53	203	54

The reference range of C-reactive protein is 0-10 mg/L. NEC: Necrotizing enterocolitis