重型β地中海贫血患儿移植前后肠道菌群变化研究

doi:10.12372/jcp.2023.22e1678

摘要/Abstract

摘要：

目的了解重型β地中海贫血（β-TM）患儿造血干细胞移植前后不同阶段肠道菌群改变及其影响因素。方法选择2019年11月至2020年6月在血液肿瘤科行造血干细胞移植的确诊β-TM的患儿作为研究对象。收集移植启动前期（A组）、清洁肠道期（B组）、预处理期（C组）、干细胞输注期（D组）及移植后细胞植入期（E组）的粪便样本。利用16SrDNA基因V4区域的Illumina HiSeq测序系统测序粪便样本。结果纳入37例β-TM患儿，男27例、女10例，中位年龄7.5（3.0~17.3）岁。最终纳入分析共96份样本，A组28份、B组17份、C组18份、D组21份、E组12份。A组和B组肠道菌群构成占比前两位为厚壁菌门、拟杆菌门，C、D、E组菌群占比前两位为变形菌门及拟杆菌门。5组之间厚壁菌门、变形菌门、拟杆菌门、梭菌门、放线菌门、疣状菌门、蓝菌门的丰度差异均有统计学意义（P<0.05）。利用Chao指数比较移植不同时期肠道菌群Alpha丰度。A组Chao指数为225.9（182.0~260.5），B组为234.6（193.0~311.3），C组为127.6（81.1~180.0），D组为96.8（71.1~139.9），E组为122.7（97.5~142.7），5组之间差异有统计学意义（P<0.001）。利用Shannon指数预测移肠道菌群Alpha多样性。A组Shannon指数为2.9（2.4~3.3），B组为2.2（1.6~2.9）、C组为1.4（1.2~2.0）、D组为2.0（1.4~2.3）、E组为0.9（0.6~2.2），5组之间差异有统计学意义（P<0.001）。万古霉素、亚胺培南-西司他丁以及其他类型广谱抗菌药物三组之间肠道菌群丰度与多样性差异无统计学意义（P>0.05）。结论造血干细胞移植术对β-TM患儿肠道菌群的组成有较大影响；清洁肠道方案对肠道菌群的丰度影响不大；肠道菌群丰度及多样性在移植后早期不能恢复重建；在造血干细胞移植过程中暂无法依据肠道菌群来优化选择抗生素种类。

关键词: 肠道菌群, 重型β地中海贫血, 造血干细胞移植, 儿童

Abstract:

Objective To investigate the changes in intestinal flora in children with β-thalassemia major (β-TM) before and after hematopoietic stem cell transplantation and the influencing factors. Methods Children diagnosed with β-TM who underwent HSCT from November 2019 to June 2020 were enrolled as the research subjects. Fecal samples were collected during the pre-transplantation stage (group A), gut decontamination period (group B), conditioning period (group C), hematopoietic stem cell infusion period (group D), and post-transplantation cellular implantation period (group E). The fecal samples were sequenced using Illumina HiSeq sequencing system targeting the V4 region of the 16SrDNA gene. Results The study included 37 children with β-TM, including 27 males and 10 females with a median age of 7.5 (3.0-17.3) years. A total of 96 samples were analyzed, with 28 from group A, 17 from group B, 18 from group C, 21 from group D, and 12 from group E. The two most abundant phyla in groups A and B were Firmicutes and Bacteroidetes. In contrast, the two most abundant phyla in groups C, D, and E were Proteobacteria and Bacteroidetes. The differences in the abundances of Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Verrucomicrobia, and Fusobacteria among the five groups were statistically significant (P<0.05). The Chao was used to compare the alpha diversity of intestinal flora at different transplantation stages. The differences among the five groups were statistically significant (P<0.001). Similarly, the Shannon was used to predict alpha diversity of intestinal flora, and the differences among the five groups were also statistically significant (P<0.001). There were no statistically significant differences in the abundance and diversity of intestinal flora among the three groups treated with vancomycin, imipenem-cilastatin, and other broad-spectrum antibiotics (P>0.05). Conclusions The effect of hematopoietic stem cell transplantation on the composition of intestinal flora in children with β-TM was significant. The gut decontamination had little effect on the abundance of intestinal microbiota. The abundance and diversity of intestinal microbiota could not be restored in the early post-transplantation period. The selection of antibiotic types during HSCT cannot be optimized based on intestinal microbiota data.

Key words: intestinal flora, β-thalassemia major in children, hematopoietic stem cell transplantation, pediatric

闻静, 余阅, 杨春兰, 吕佳忆, 宋欣平, 李越, 张小玲, 王晓东, 刘四喜. 重型β地中海贫血患儿移植前后肠道菌群变化研究[J]. 临床儿科杂志, 2023, 41(11): 833-838.

WEN Jing, YU Yue, YANG Chunlan, LYU Jiayi, SONG Xinping, LI Yue, ZHANG Xiaoling, WANG Xiaodong, LIU Sixi. Changes of intestinal flora in children with β-thalassemia major before and after transplantation[J]. Journal of Clinical Pediatrics, 2023, 41(11): 833-838.

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参考文献 24

[1]	文飞球, 刘四喜. 启动重型β地中海贫血防控的攻坚战[J]. 中华儿科杂志, 2018, 56(10): 721-723.
[2]	广东省地中海贫血防治协会, 中国实用儿科杂志编辑委员会. 造血干细胞移植治疗重型β地中海贫血儿科专家共识[J]. 中国实用儿科杂志, 2018, 33(12): 935-939.
[3]	Qin J, Li R, Raes J, et al. A human gut microbial gene catalogue established by metagenomic sequencing[J]. Nature, 2010, 464(7285): 59-65. doi: 10.1038/nature08821
[4]	Peled JU, Gomes ALC, Devlin SM, et al. Microbiota as predictor of mortality in allogeneic hematopoietic-cell transplantation[J]. N Engl J Med, 2020, 382(9): 822-834. doi: 10.1056/NEJMoa1900623
[5]	许吕宏, 方建培. 《重型β地中海贫血的诊断和治疗指南(2017年版)》解读[J]. 中国实用儿科杂志, 2018, 33(12): 940-943.
[6]	D'Souza A, Fretham C, Lee SJ, et al. Current use of and trends in hematopoietic cell transplantation in the United States[J]. Biol Blood Marrow Transplant, 2020, 26(8): e177-e182. doi: 10.1016/j.bbmt.2020.04.013
[7]	Penack O, Peczynski C, Mohty M, et al. How much has allogeneic stem cell transplant-related mortality improved since the 1980s? A retrospective analysis from the EBMT[J]. Blood Adv, 2020, 4(24): 6283-6290. doi: 10.1182/bloodadvances.2020003418 pmid: 33351121
[8]	Chang YJ, Pei XY, Huang XJ. Haematopoietic stem-cell transplantation in China in the era of targeted therapies: current advances, challenges, and future directions[J]. Lancet Haematol, 2022, 9(12): e919-e929. doi: 10.1016/S2352-3026(22)00293-9
[9]	Ruff WE, Greiling TM, Kriegel MA. Host-microbiota interactions in immune-mediated diseases[J]. Nat Rev Microbiol, 2020, 18(9): 521-538. doi: 10.1038/s41579-020-0367-2 pmid: 32457482
[10]	Lozupone CA, Stombaugh JI, Gordon JI, et al. Diversity, stability and resilience of the human gut microbiota[J]. Nature, 2012, 489(7415): 220-230. doi: 10.1038/nature11550
[11]	Gałązka P, Styczyński J, Czyżewski K, et al. Impact of decontamination therapy on gastrointestinal acute graft-versus-host disease after allogeneic hematopoietic cell transplantation in children: decontamination therapy in allo-HCT[J]. Curr Res Transl Med, 2021, 69(3): 103298.
[12]	Severyn CJ, Siranosian BA, Kong ST, et al. Microbiota dynamics in a randomized trial of gut decontamination during allogeneic hematopoietic cell transplantation[J]. JCI Insight, 2022, 7(7): e154344. doi: 10.1172/jci.insight.154344
[13]	Bekker V, Zwittink RD, Knetsch CW, et al. Dynamics of the gut microbiota in children receiving selective or total gut decontamination treatment during hematopoietic stem cell transplantation[J]. Biol Blood Marrow Transplant, 2019, 25(6): 1164-1171. doi: 10.1016/j.bbmt.2019.01.037
[14]	Taur Y, Xavier JB, Lipuma L, et al. Intestinal domination and the risk of bacteremia in patients undergoing allogeneic hematopoietic stem cell transplantation[J]. Clin Infect Dis, 2012, 55(7): 905-914. doi: 10.1093/cid/cis580 pmid: 22718773
[15]	Holler E, Butzhammer P, Schmid K, et al. Metagenomic analysis of the stool microbiome in patients receiving allogeneic stem cell transplantation: loss of diversity is associated with use of systemic antibiotics and more pronounced in gastrointestinal graft-versus-host disease[J]. Biol Blood Marrow Transplant, 2014, 20(5): 640-645. doi: 10.1016/j.bbmt.2014.01.030
[16]	Wei L, Wen XS, Xian CJ. Chemotherapy-induced intestinal microbiota dysbiosis impairs mucosal homeostasis by modulating toll-like receptor signaling pathways[J]. Int J Mol Sci, 2021, 22(17): 9474. doi: 10.3390/ijms22179474
[17]	Rajagopala SV, Singh H, Yu Y, et al. Persistent gut microbial dysbiosis in children with acute lymphoblastic leukemia (all) during chemotherapy[J]. Microb Ecol, 2020, 79(4): 1034-1043. doi: 10.1007/s00248-019-01448-x pmid: 31754744
[18]	Yixia Y, Sripetchwandee J, Chattipakorn N, et al. The alterations of microbiota and pathological conditions in the gut of patients with colorectal cancer undergoing chemotherapy[J]. Anaerobe, 2021, 68: 102361. doi: 10.1016/j.anaerobe.2021.102361
[19]	陈森敏, 刘四喜, 陈芬, 等. 儿童急性淋巴细胞白血病化疗前后肠道菌群变化特点[J]. 中国当代儿科杂志, 2022, 24(5): 550-560.
[20]	Harris B, Morjaria SM, Littmann ER, et al. Gut microbiota predict pulmonary infiltrates after allogeneic hematopoietic cell transplantation[J]. Am J Respir Crit Care Med, 2016, 194(4): 450-463. doi: 10.1164/rccm.201507-1491OC
[21]	Le Bastard Q, Chevallier P, Montassier E. Gut microbiome in allogeneic hematopoietic stem cell transplantation and specific changes associated with acute graft vs host disease[J]. World J Gastroenterol, 2021, 27(45): 7792-7800. doi: 10.3748/wjg.v27.i45.7792
[22]	Ingham AC, Kielsen K, Mordhorst H, et al. Microbiota long-term dynamics and prediction of acute graft-versus-host disease in pediatric allogeneic stem cell transplantation[J]. Microbiome, 2021, 9(1): 148. doi: 10.1186/s40168-021-01100-2 pmid: 34183060
[23]	Hayase E, Hayase T, Jamal MA, et al. Mucus-degrading bacteroides link carbapenems to aggravated graft-versus-host disease[J]. Cell, 2022, 185(20): 3705-3719. doi: 10.1016/j.cell.2022.09.007 pmid: 36179667
[24]	Elgarten CW, Li Y, Getz KD, et al. Broad-spectrum antibiotics and risk of graft-versus-host disease in pediatric patients undergoing transplantation for acute leukemia: association of carbapenem use with the risk of acute graft-versus-host disease[J]. Transplant Cell Ther, 2021, 27(2): 177.

组别	A组(n=28)	B组(n=17)	C组(n=18)	D组(n=21)	E组(n=12)	统计量	P
年龄[M(P₂₅~P₇₅)]/岁	7.3(5.0~9.9)	6.6(4.6~9.3)	7.6(6.3~10.0)	7.5(6.3~9.3)	8.2(5.9~16.6)	H=1.98	0.740
女性[n(%)]	8(28.6)	5(29.4)	5(27.8)	4(19.0)	4(33.3)	－	0.908¹⁾
供者类型[n(%)]						－	0.831¹⁾
同胞全相和	3(10.7)	4(23.5)	2(11.1)	3(14.3)	2(16.7)
无关全相和	0(0)	0(0)	1(5.6)	1(4.8)	0(0)
单倍体	25(89.0)	13(76.5)	15(83.3)	17(80.9)	10(83.3)
预处理方案[n(%)]						－	0.808¹⁾
Cy+Bu+Flu+TT+ATG	25(89.3)	13(76.5)	16(88.9)	18(85.7)	10(83.3)
Cy+CSA+Bu+Flu+TT+ATG	3(10.7)	4(23.5)	2(11.1)	3(14.3)	2(16.7)
预防GVHD[n(%)]						－	0.808¹⁾
CSA+MMF	3(10.7)	4(23.5)	2(11.1)	3(14.3)	2(16.7)
Pt-Cy+MMF+FK506	25(89.3)	13(76.5)	16(88.9)	18(85.7)	10(83.3)

组别	A组(n=28)	B组(n=17)	C组(n=18)	D组(n=21)	E组(n=12)	H值	P
厚壁菌门	40.7(24.1~50.9)	21.0(6.6~38.8)	9.9(1.1~28.2)	18.3(13.8~25.6)	48.6(20.4~92.0)	20.57	<0.001
变形菌门	73.6(2.7~15.8)	28.3(19.0~61.8)	43.6(14.3~80.8)	14.3(6.2~46.1)	3.8(0.4~54.0)	21.01	<0.001
拟杆菌门	44.5(27.8~52.8)	25.4(10.3~82.7)	0.6(0.0~39.4)	50.1(12.9~60.7)	0.4(0.1~39.0)	18.46	0.001
梭菌门	0.1(0.0~1.5)	0.1(0.0~18.7)	8.0(0.1~15.3)	0.2(0.0~20.1)	0.0(0.0~0.0)	11.22	0.024
放线菌门	0.4(0.1~13.7)	1.0(0.3~4.1)	0.0(0.0~0.1)	0.1(0.0~0.3)	0.0(0.0~0.7)	68.27	<0.001
疣状菌门	0.0(0.0~0.1)	0.0(0.0~0.1)	0.0(0.0~2.0)	0.0(0.0~0.0)	0.0(0.0~0.0)	15.41	0.040
蓝菌门	0.0(0.0~0.0)	0.0(0.0~2.7)	0.0(0.0~0.0)	0.0(0.0~0.0)	0.0(0.0~0.2)	42.65	<0.001