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Maximum depth sequencing reveals an ON/OFF replication slippage switch and apparent in vivo selection for bifdobacterial pilus expression

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posted on 2023-10-10, 09:06 authored by Christophe Penno, Mary O’Connell Motherway, Yuan Fu, Virag SharmaVirag Sharma, Fiona Crispie, Paul D. Cotter, Benoit Houeix, Lokesh Joshi, Francesca Bottacini, Aoife O’Dwyer, Gary LoughranGary Loughran, John F.Atkins, Douwe Van Sinderen

The human gut microbiome, of which the genus Bifidobacterium is a prevalent and abundant member, is thought to sustain and enhance human health. Several surface-exposed structures, including so-called sortase-dependent pili, represent important bifidobacterial gut colonization factors. Here we show that expression of two sortase-dependent pilus clusters of the prototype Bifidobacterium breve UCC2003 depends on replication slippage at an intragenic G-tract, equivalents of which are present in various members of the Bifidobacterium genus. The nature and extent of this slippage is modulated by the host environment. Involvement of such sortase-dependent pilus clusters in microbe-host interactions, including bacterial attachment to the gut epithelial cells, has been shown previously and is corroborated here for one case. Using a Maximum Depth Sequencing strategy aimed at excluding PCR and sequencing errors introduced by DNA polymerase reagents, specific G-tract sequences in B. breve UCC2003 reveal a range of G-tract lengths whose plasticity within the population is functionally utilized. Interestingly, replication slippage is shown to be modulated under in vivo conditions in a murine model. This in vivo modulation causes an enrichment of a G-tract length which appears to allow biosynthesis of these sortase-dependent pili. This work provides the first example of productive replication slippage influenced by in vivo conditions. It highlights the potential for microdiversity generation in “beneficial” gut commensals.


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Scientific Reports, 2022, 12, 9576



Other Funding information

Tis work was supported by Science Foundation Ireland (SFI) through the Irish Government’s National Development Plan [grant numbers SFI/12/RC/2273-P1, SFI/12/RC/2273-P2], and an HRB postdoctoral fellowship [grant no. PDTM/20011/9] awarded to MOCM. AD was supported by an internal APC summer studentship. GL and JFA were supported by Irish Research Council Advanced Laureate award IRCLA/2019/74 to J.F.A.

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