High-Throughput Sequencing of Campylobacter jejuni Insertion Mutant Libraries Reveals mapA as a Fitness Factor for Chicken Colonization

Abstract

bCampylobacter jejuni is a leading cause of gastrointestinal infections worldwide, due primarily to its ability to asymptomatically colonize the gastrointestinal tracts of agriculturally relevant animals, including chickens. Infection often occurs following consumption of meat that was contaminated by C. jejuni during harvest. Because of this, much interest lies in understanding the mechanisms that allow C. jejuni to colonize the chicken gastrointestinal tract. To address this, we generated a C. jejuni transposon mutant library that is amenable to insertion sequencing and introduced this mutant pool into day-of-hatch chicks. Following deep sequencing of C. jejuni mutants in the cecal outputs, several novel factors required for efficient colonization of the chicken gastrointestinal tract were identified, including the predicted outer membrane protein MapA. A mutant strain lacking mapA was constructed and found to be significantly reduced for chicken colonization in both competitive infections and monoinfections. Further, we found that mapA is required for in vitro competition with wild-type C. jejuni but is dispensable for growth in monoculture.

The PAS Domain-Containing Protein HeuR Regulates Heme Uptake in Campylobacter jejuni

“…Using a Tn-Seq approach, we previously identified several determinants required by C. jejuni for efficient colonization of the chicken gastrointestinal tract (4). One determinant was a component of the Campylobacter heme utilization gene cluster chuB .…”

“…Previously, our group has used both signature-tagged mutagenesis (STM) and insertion sequencing (transposon sequencing [Tn-Seq]) approaches to identify colonization determinants of C. jejuni (4, 5). The earlier STM approach identified genes involved in chemotaxis and flagellar biogenesis, but several other colonization factors were identified using the Tn-Seq approach.…”

“…The earlier STM approach identified genes involved in chemotaxis and flagellar biogenesis, but several other colonization factors were identified using the Tn-Seq approach. One determinant identified to be reduced by 250-fold in cecal outputs by Tn-Seq was a component of the heme utilization gene cluster chuB (Cjj81176_1602), indicating that acquiring iron from heme may be important during colonization of the chicken gastrointestinal tract (4). …”

Narrow-Spectrum Inhibitors of Campylobacter jejuni Flagellar Expression and Growth

“…Infection of humans often occurs following ingestion of undercooked meat or by consuming uncooked food that has been cross-contaminated. As such, much emphasis has been placed on identifying and characterizing factors of C. jejuni with the goal of using those insights to inform the identification and/or development of strategies or compounds that limit C. jejuni loads in the food supply (2,3). One such factor that has repeatedly been shown to be required for full colonization of the chicken gastrointestinal is the flagella of C. jejuni.…”

“…These include signature-tagged mutagenesis and transposon insertion sequencing (Tn-Seq) approaches to identify mutants with decreased cecal abundance; these studies demonstrated that flagella are required for full colonization of the chicken cecum (2,3). Additionally, the regulatory hierarchy of flagellar biosynthesis has been characterized (4).…”