A plasmid construct was designed in which the gene of antimicrobial peptide melittin is controlled by the tetracycline-responsive promoter of human cytomegalovirus, aided by a constitutively expressed trans-activator protein gene. Its vaginal administration and induction of melittin gene transcription with doxycycline markedly suppressed subsequent genital tract infection of mice by Mycoplasma hominis and Chlamydia trachomatis. At least half of the melittin-protected animals proved free of either pathogen within 3-4 weeks. Recombinant plasmids expressing genes of antimicrobial peptides hold much promise as agents for prevention and control of urogenital latent infections.
The aims of our research were to investigate the gene expression of the multidrug efflux transporter, CDR1 and the major drug facilitator superfamily transporter, MDR1 gene in azole drug-resistant Candida albicans and Candida glabrata clinical isolates recovered from vaginitis patients; and to identify hotspot mutations that may be present in the C. albicans CaCDR1 gene that could be associated with drug-resistance. The relative expression of the CDR1 and MDR1 transcripts in ketoconazole and clotrimazole-resistant isolates and drug-susceptible ATCC strains were determined by semi-quantitative reverse transcription-polymerase chain reaction. Expression of CaCDR1 transcript was upregulated to varying extents in all three azole-resistant C. albicans isolates studied (1.6-, 3.7- and 3.9-fold) and all three C. glabrata isolates tested (at 1.9-, 2.3- and 2.7-fold). The overexpression level of CaCDR1 in the isolates correlated with the degree of resistance as reflected by the minimum inhibitory concentration (MIC) of the drugs. The messenger RNA for another efflux pump, MDR1, was also overexpressed in one of the azole-resistant C. albicans isolates that overexpressed CDR1. This finding suggests that drug-resistance may involve synergy between energy-dependent drug efflux pumps CDR1p and MDR1p in some but not all isolates. Interestingly, DNA sequence analysis of the promoter region of the CaCDR1 gene revealed several point mutations in the resistant clinical isolates compared to the susceptible isolates at 39, 49 and 151 nucleotides upstream from the ATG start codon. This finding provides new information on point mutations in the promoter region which may be responsible for the overexpression of CDR1 in drug-resistant isolates.