Herpes simplex virus co-infection-induced Chlamydia trachomatis persistence is not mediated by any known persistence inducer or anti-chlamydial pathway, by J. Vanover, J. Sun, S. Deka, J. Kintner, M. M. Duffourc and R. V. Schoborg
Microbiology vol. 154, part 3, pp. 971 - 978
Table S1. Semi-quantitative RT-PCR primer and synthetic DNA target sequences. [PDF] (57 kb)
Fig. S1. C. trachomatis/HSV co-infected cells do not produce nitric oxide synthase. Total cellular RNA from mock-, C. trachomatis and HSV-2 singly- and co-infected HeLa cells was used for semiquantitative RT-PCR with primers specific to human inos (a) and 18S rRNA (b). A dilution series of synthetic DNA targets or host genomic DNA served as an amplification control for each gene. n=8. [PDF] (405 kb)
Fig. S2. HSV-2 co-infection does not stimulate ido or trpA expression. (a) C. trachomatis-infected HeLa cultures were either exposed to diluent (-IFN-γ) or 50 U ml-1 of IFN-γ (+IFN-γ) beginning immediately after infection. Total RNA was isolated at 48 h post-infection as described and subjected to RT-PCR using primers specific for human ido. (b, c)Total cellular RNA was isolated from mock-, C. trachomatis and HSV-2 singly- and co-infected HeLa cultures for semiquantitative RT-PCR using primers specific to human ido (b) and chlamydial trpA (c). A dilution series of synthetic DNA targets (ido) or chlamydial genomic DNA (trpA) served as an amplification control for each gene. Chlamydial trpA and human ido amplimers were quantified as described and normalized to chlamydial (trpA) or host (ido) genome copy number (data not shown). n=8. [PDF] (581 kb)
Fig. S3. HSV-2 induction of chlamydial persistence is not mediated by nutrient deprivation. (a) Cultures of HeLa cells were mock-, C. trachomatis singly-infected +/- Desferal or co-infected. Protein concentrations were standardized using the MicroBCA kit (Pierce) prior to analysis by ELISA. Ferritin concentration is expressed as ng per ml sampleħSEM; n=3. Asterisks (*) indicate ferritin concentrations that are significantly different (by t-test) compared to mock-infected cells (P<0.05). (b, c) Cultures of mock-, C. trachomatis and HSV-2 singly- and co-infected HeLa cells were refed with either MEM (controls) or MEM+6 mg hTF ml-1, 5X (1.4 mM) essential and non-essential amino acids or 450 mg ml-1 glucose following HSV-2 adsorption and harvested for EB titration at 20 h post HSV-2 infection. EB titres are expressed as IFU per ml sampleħSEM; n=3. Asterisks (*) indicate titres that are significantly different (by t-test) compared to those from C. trachomatis singly-infected cells (P<0.05). The data shown are representative of three independent experiments. [PDF] (727 kb)
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