Host Range of Chlamydiaphages φCPAR39 and Chp3

Abstract

The host range of CPAR39 is limited to four Chlamydophila species: C. abortus, C. caviae, C. pecorum, and C. pneumoniae. Chp3 (a newly discovered bacteriophage isolated from C. pecorum) shares three of these hosts (C. abortus, C. caviae, and C. pecorum) but can additionally infect Chlamydophila felis. The ability to support replication was directly correlated with the binding properties of the respective bacteriophages with their host species. Binding studies also show that CPAR39 and Chp3 use different host receptors to infect the same host cells: cell binding is sensitive to proteinase K treatment, confirming that the chlamydiaphage receptors are proteinaceous in nature.Chlamydiae have an obligate intracellular developmental cycle that alternates between the infectious elementary body and the replicative form, the reticulate body (RB). Five bacteriophages have been isolated from the chlamydiae. The first chlamydial bacteriophage to be characterized (13) was found by thin-section transmission electron microscopy (EM) of avian Chlamydophila psittaci RBs (14). Although samples containing Chp1 were later lost, a second bacteriophage (CPG1) (7) was found to infect Chlamydophila caviae (8) and a third bacteriophage, Chp2, was isolated from Chlamydophila abortus (10). The Chlamydophila pneumoniae strain AR39 genomesequencing project revealed the presence of a double-stranded DNA extrachromosomal element (11), which was subsequently shown to be the replicative form of bacteriophage CPAR39 (4). Recently Chp3 was isolated from Chlamydophila pecorum (6) and was found to share 97.1% nucleotide sequence identity with Chp2. Chlamydiaphages CPAR39 and CPG1 also show 97.1% identity, whereas the two groups (Chp2-Chp3 and CPAR39-CPG1) share 93.3% identity. All the chlamydiaphages share similar features; they are small icosahedral T ϭ 1 particles containing circular, single-stranded DNA genomes, and molecular characterization showed that they belong to the virus family Microviridae (9-11, 17).Recently we described the host range of Chp2 and showed that Chp2 was unable to infect C. caviae or C. pneumoniae. It was therefore of interest to investigate the host range of the very closely related Chp3 and the slightly more diverse bacteriophages CPAR39 and CPG1, which were isolated from host chlamydiae resistant to Chp2. In our work, monolayers of BGMK (Buffalo green monkey kidney) or HEp2 cells in 25-cm 2 flasks were infected by centrifugation at 1,000 ϫ g for 1 h in medium containing cycloheximide (1 g/ml) and gentamicin (25 g/ml) with C. pecorum bearing Chp3 or C. pneumoniae (11) bearing CPAR39. At 72 h postinfection the culture medium was replaced with a small volume of phosphate-buffered saline (PBS), and the flasks were frozen at Ϫ70°C. One hundred flasks of phage-infected chlamydiae were prepared, stored frozen, and then processed as a single batch. The flasks were frozen and thawed three times to lyse the chlamydial RBs and release phage. Any remaining monolayer that had not detached after this procedure was scraped off. The s...

Horizontal Gene Transfer and the Evolution of Microvirid Coliphage Genomes

“…They have been well studied in molecular biology (20) and in experimental evolution (5,6,9,24,53). The 12 previously sequenced microvirid phages were isolated on Escherichia coli, Salmonella, Bdellovibrio (2), Chlamydia, (15,16,28,45), and Spiroplasma (41). The initial studies of microvirid genomes seemed to confirm differences in genome evolution from what has been observed in the dsDNA phages.…”

Lateral Gene Transfer In Vitro in the Intracellular PathogenChlamydia trachomatis

“…Phage-mediated transduction in C. trachomatis appears to be unlikely because a C. trachomatis phage has not been reported, although phages are present in other species of Chlamydia (12). Conjugation-mediated DNA transfer occurs in two main ways: (i) an extrachromosomal plasmid is transferred from a donor to a recipient bacterium, and (ii) a chromosome-mobilizing plasmid interacts with the DNA donor chromosome in a way that initiates transfer of chromosomal DNA into a recipient bacterium that lacks the plasmid.…”

The plasmids of Chlamydia trachomatis and Chlamydophila pneumoniae (N16): accurate determination of copy number and the paradoxical effect of plasmid-curing agents

“…Extrachromosomal elements have been detected in several chlamydial species: both a cryptic plasmid (Lovett et al, 1980;Thomas et al, 1997) and a chlamydiaphage of the microvirus family (Everson et al, 2003;Richmond et al, 1982) have been described. Almost all strains of C. trachomatis harbour the plasmid, but some plasmid-free isolates have been described Farencena et al, 1997;Matsumoto et al, 1998;Peterson et al, 1990;Stothard et al, 1998Stothard et al, , 1999.…”