Microbial exposure during early human development primes fetal immune cells
Authors: Nurhidaya Binte Shadan
Gillian Low
Rhea Pai
Charles Antoine Dutertre
Avery Khoo May Lee
Shabnam Khalilnezhad
Norman Pavelka
Regina Men Men Wong
Ravid Straussman
Naomi McGovern
Mahesh Choolani
Florent Ginhoux
Aviva Rotter-Maskowitz
Pei Xiang Hor
Andrea Lee
Thet Tun Aung
Leong Jing Yao
Benoit Malleret
Salvatore Albani
Svetoslav Chakarov
Ghee Chuan Lai
Ankur Sharma
Xiao-Meng Zhang
Phyllis Chen
Yiping Fan
Jerry Kok Yen Chan
Fabien Cottier
Costerwell Khyriem
Adhika Shanti
Edwin Shepherdson
Alrina Shin Min Tan
Gurmit Singh Naranjan Singh
K G Srinivasan
David Vermijlen
Garold Fuks
Archita Mishra
Noam Shental
Journal: Cell (2021)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: Tem; Treg; bacteria; fetal Development; fetal immunity
Affiliations: Singapore Immunology Network (SIgN), A(∗)STAR, 8A Biomedical Grove, Immunos Building, Level 4, Singapore 138648, Singapore., Translational Immunology Institute, Singhealth/Duke-NUS Academic Medical Centre, the Academia, 20 College Road, Discovery Tower Level 8, Singapore 169856, Singapore., Department of Microbiology and Immunology, Immunology Translational Research Programme, Yong Loo Lin School of Medicine, Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore 117597, Singapore., Department of Mathematics and Computer Science, Open University of Israel, Ra'anana 4353701, Israel., Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot 7610001, Israel., Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore 229899, Singapore., Genome Institute of Singapore (GIS), A(∗)STAR, 60 Biopolis Street, Singapore 138672, Singapore., Singapore Immunology Network (SIgN), A(∗)STAR, 8A Biomedical Grove, Immunos Building, Level 4, Singapore 138648, Singapore; Translational Immunology Institute, Singhealth/Duke-NUS Academic Medical Centre, the Academia, 20 College Road, Discovery Tower Level 8, Singapore 169856, Singapore; Program in Emerging Infectious Disease, Duke-NUS Medical School, 8 College Road, Singapore 169857, Singapore., Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore 229899, Singapore; Experimental Fetal Medicine Group, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore., Department of Obstetrics & Gynaecology, Yong Loo Lin School of Medicine, National University of Singapore, NUHS Tower Block, 1E Kent Ridge Road, Singapore 119228, Singpore., Department of Pharmacotherapy and Pharmaceutics, Institute for Medical Immunology, ULB Center for Research in Immunology (U-CRI), Université Libre de Bruxelles (ULB), Brussels 1050, Belgium., Genome Institute of Singapore (GIS), A(∗)STAR, 60 Biopolis Street, Singapore 138672, Singapore; Harry Perkins Institute of Medical Research, QEII Medical Centre and Centre for Medical Research, the University of Western Australia, PO Box 7214, 6 Verdun Street, Nedlands, Perth, WA 6009, Australia; Curtin Medical School, Curtin Health Innovation Research Institute, Curtin University, Perth, WA 6102, Australia., Department of Physics of Complex Systems, Weizmann Institute of Science, Rehovot 7610001, Israel., Singapore Immunology Network (SIgN), A(∗)STAR, 8A Biomedical Grove, Immunos Building, Level 4, Singapore 138648, Singapore; Department of Microbiology and Immunology, Immunology Translational Research Programme, Yong Loo Lin School of Medicine, Immunology Programme, Life Sciences Institute, National University of Singapore, Singapore 117597, Singapore., Department of Pathology and Centre for Trophoblast Research, Tennis Court Road, Cambridge CB2 1QP, UK. Electronic address: ., Translational Immunology Institute, Singhealth/Duke-NUS Academic Medical Centre, the Academia, 20 College Road, Discovery Tower Level 8, Singapore 169856, Singapore. Electronic address: ., Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore 229899, Singapore; Experimental Fetal Medicine Group, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore; OBGYN-Academic Clinical Program, Duke-NUS, Duke-NUS Medical School, 8 College Road, Singapore 169857, Singapore; Cancer and Stem Cell Biology Program, Duke-NUS Graduate Medical School, Singapore 119077, Singapore. Electronic address: ., Singapore Immunology Network (SIgN), A(∗)STAR, 8A Biomedical Grove, Immunos Building, Level 4, Singapore 138648, Singapore; Translational Immunology Institute, Singhealth/Duke-NUS Academic Medical Centre, the Academia, 20 College Road, Discovery Tower Level 8, Singapore 169856, Singapore; Shanghai Institute of Immunology, Shanghai JiaoTong University School of Medicine, 280 South Chongqing Road, Shanghai 200025, China. Electronic address: .

Abstract:  The human fetal immune system begins to develop early during gestation; however, factors responsible for fetal immune-priming remain elusive. We explored potential exposure to microbial agents in utero and their contribution toward activation of memory T cells in fetal tissues. We profiled microbes across fetal organs using 16S rRNA gene sequencing and detected low but consistent microbial signal in fetal gut, skin, placenta, and lungs in the 2nd trimester of gestation. We identified several live bacterial strains including Staphylococcus and Lactobacillus in fetal tissues, which induced in vitro activation of memory T cells in fetal mesenteric lymph node, supporting the role of microbial exposure in fetal immune-priming. Finally, using SEM and RNA-ISH, we visualized discrete localization of bacteria-like structures and eubacterial-RNA within 14th weeks fetal gut lumen. These findings indicate selective presence of live microbes in fetal organs during the 2nd trimester of gestation and have broader implications toward the establishment of immune competency and priming before birth. 

Retracted: No

PubPeer comments:
 Although the authors emphasize their u201cstringentu201d use of negative controls, they lack any control for the most obvious and greatest source of contaminationu2014the maternal vaginal microbiota. The authors note u201ca gradual increase in bacterial diversity in fetal tissue from external or surface-exposed organs in comparison to internal controls,u201d consistent with contamination during sample collection. Given the evident sampling contamination, it is unsurprising that the authors were successful in culturing live bacteria in increasing abundance from internal organs (spleen, thymus) to exposed organs (gut, lung) and external tissue (skin, placenta). The SEM data show  bacteria-like morphological structures that are entirely coccoid. This is inconsistent with their culture and sequencing data, where the majority of bacteria were bacilli, but consistent with contamination by Micrococcus, a genus enriched in PBS (in which fixed tissues were washed and stored). Additionally, the authors finding that u201cmost regions did not show any bacterial localizationu201d by SEM is inconsistent with their RNA-ISH results showing bacterial RNA  tightly packed alongside intestinal epithelium (which is indistinguishable from autofluorescence given the absence of negative controls). , 
 RE the RNA-ish data, as @bioelze pointed out to me on Twitter, the Epcam and 16S signals seem to be inverted. The  16S signal is a very clear Epcam RNA stain (i.e. all intracellular epithelial) whereas the  Epcam signal is spread out punctate, so more likely to be bugs or background as you point out... ,