Identification of a <i>Coxiella burnetii</i> outer membrane porin required for intracellular replication

<i>Coxiella burnetii</i> is a gram-negative, obligate intracellular pathogen that causes human Q fever. Within host cells, <i>C. burnetii</i> proliferates in a spacious, acidic, lysosome-derived <i>Coxiella</i>-containing vacuole (CCV) by a process that requires the Dot/Icm type IVB secretion system to deliver effectors that manipulate host cell functions. A previous transposon mutagenesis screen identified the gene <i>cbu0937</i> as being important for intracellular replication of <i>C. burnetii</i>. Here, the function of Cbu0937 was investigated. The <i>cbu0937</i>::Tn mutant had no detectable defect replicating in the axenic acidified citrate cysteine medium 2. Additionally, intracellular replication of the <i>cbu0937</i>::Tn mutant was not restored by co-infection of host cells with an isogenic wild-type strain of <i>C. burnetii</i>. Thus, the <i>cbu0937</i>::Tn mutant has a cell-intrinsic intracellular replication defect. Intracellular replication of the <i>cbu0937</i>::Tn mutant was restored by complementing the gene <i>in trans</i> with a plasmid encoding either untagged or an epitope-tagged version of Cbu0937. Analysis of the predicted structure of the Cbu0937 protein using AlphaFold revealed high similarity between Cbu0937 and several bacterial porins. Fractionation studies and surface labeling of <i>C. burnetii</i> producing a functional epitope-tagged protein indicated the localization of Cbu0937 to the bacterial outer membrane. From these data, we conclude that <i>cbu0937</i> encodes a porin that plays an essential role in supporting <i>C. burnetii</i> intracellular replication, which likely involves the acquisition of an important metabolite in the CCV lumen.