Human C. difficile toxin–specific memory B cell repertoires encode poorly neutralizing antibodies
Hemangi B. Shah, … , Jimmy D. Ballard, Mark L. Lang
Hemangi B. Shah, … , Jimmy D. Ballard, Mark L. Lang
Published July 14, 2020
Citation Information: JCI Insight. 2020;5(16):e138137. https://doi.org/10.1172/jci.insight.138137.
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Research Article Immunology Infectious disease

Human C. difficile toxin–specific memory B cell repertoires encode poorly neutralizing antibodies

Abstract

Clostridioides difficile is a leading cause of nosocomial infection responsible for significant morbidity and mortality with limited options for therapy. Secreted C. difficile toxin B (TcdB) is a major contributor to disease pathology, and select TcdB-specific Abs may protect against disease recurrence. However, the high frequency of recurrence suggests that the memory B cell response, essential for new Ab production following C. difficile reexposure, is insufficient. We therefore isolated TcdB-specific memory B cells from individuals with a history of C. difficile infection and performed single-cell deep sequencing of their Ab genes. Herein, we report that TcdB-specific memory B cell–encoded antibodies showed somatic hypermutation but displayed limited isotype class switch. Memory B cell–encoded mAb generated from the gene sequences revealed low to moderate affinity for TcdB and a limited ability to neutralize TcdB. These findings indicate that memory B cells are an important factor in C. difficile disease recurrence.

Authors

Hemangi B. Shah, Kenneth Smith, Edgar J. Scott II, Jason L. Larabee, Judith A. James, Jimmy D. Ballard, Mark L. Lang

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Figure 1

Isolation, sequencing, and repertoire analysis of CTD-specific Bmem cells.

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Isolation, sequencing, and repertoire analysis of CTD-specific Bmem cell...
(A) Blood samples were obtained from subjects with a history of CDI and used as a source of enriched B cells. (B) Enriched B cells were labeled with a cocktail of fluorochrome-conjugated mAbs and Alexa 488–conjugated CTD as described in Methods. Pseudocolor plots 1 through 3 depict the gating strategy, allowing identification and sorting of CD19+CD20+CD27+CD38–CTD+ cells and CD19+CD20+CD27+CD38–CTD– cells. Data shown are from subject 1008. (C) Shows frequency of CTD+ Bmem in 3 healthy controls versus 6 previously infected subjects. The subjects included in the single-cell analysis are denoted by red symbols: subjects 1008 (0.34%), 1009 (0.06%), and 1013 (0.11%). Black dots denote other subjects recruited: subjects 1012 (0%), 1015 (0.11%), and 1018 (0.19%). The line indicates the mean. A 2-tailed unpaired t test with Welch’s correction was applied to determine statistical significance in the differences observed (*P < 0.05). (D) Sorted CTD+ and CTD– Bmem cells were processed as depicted and described in Methods, resulting in the generation of individually barcoded and fully sequenced V(D)J regions for each Bmem cell. FASTA files were generated using the Cell Ranger 3.0.2 pipeline, and the Change-O toolkit was used to analyze the data.