BackgroundPhellodendron amurense, exhibits antifungal activity mainly by bioactive components including berberine hydrochloride and palmatine hydrochloride. This study was conducted to evaluate the antifungal effects of berberine hydrochloride, palmatine hydrochloride, and a mixture of both substances against Microsporum canis in vivo and in vitro.MethodsThe minimal inhibitory concentrations (MICs) of monomers and clotrimazole were determined using 1.5 % tryptic soy agar. The effects of these drugs on Microsporum canis growth was detected by determining dry weight. Transmission electron microscopy (TEM) was performed to observe the effect of chemicals on cell ultrastructure. Differential mRNA expressions of eight genes of M. canis treated with berberine or palmatine or their combination at different time points were determined by real-time PCR. NADH enzyme concentration was also detected. Clinical evaluation via in-vivo antifungal assay was also performed. Skin histology PAS staining was also carried out.ResultsResults showed that MICs of berberine, palmatine and clotrimazole were 1, 1, and 0.015 mg/mL, respectively. No significant difference was observed among the growth curves of the three groups before 18 h was reached. TEM showed that these drugs could destroy the cell membrane and organelles of M. canis at different time points. After 30 h of incubation, relative mRNA expressions of the genes in the combined group were significantly higher than those in the other groups including the clotrimazole group (P < 0.05); Palmatine initially induced the mRNA up-regulation of PGAL4, FSH1, PQ-LRP, NADH1 and NDR in M. canis; by contrast, berberine maintained a high expression level of these genes to shorten fungal life cycle and eradicate M. canis. Clinical results showed that combined treatment was more effective than single administration of each monomer or clotrimazole. Hence, berberine mixed with palmatine significantly elicited antifungal activities and could be used to treat M. canis in rabbits.ConclusionThese results provide a comprehensive view of the mechanism of berberine and palmatine in anti-M. canis activity.
BackgroundThe Gram-negative pathogen Bordetella bronchiseptica causes acute and chronic respiratory infection in a variety of animals. Currently, there is no vaccine to prevent these infections. To identify useful candidate antigens for such a vaccine, five B. bronchiseptica genes including amino acid ATP-binding cassette transporter substrate-binding protein (ABC), lipoprotein (PL), outer membrane porin protein (PPP), leu/ile/val-binding protein (BPP), and conserved hypothetical protein (CHP) were cloned and the recombinant proteins were expressed. The immune responses of mice to vaccination with individual recombinant proteins were measured.ResultsEach of the tested recombinant proteins induced a high antibody titer. PPP and PL showed protective indices against challenges with B. bronchiseptica. The protection ratios were 62.5 and 50 %, respectively, compared with 12.5 % for control vaccinations. The protection ratios of ABC, BPP, and CHP were not significantly different from the controls. IgG-subtype and cytokine analysis demonstrated that PPP and PL can induce two immune responses: a humoral immune response and a cell-mediated immune response. The humoral immunity-mediated, Th2-type response dominated.ConclusionThe identification of PPP and PL, which offer immune-protective potential, identifies them as candidates for the development of a diagnostic test or a vaccine for B. bronchiseptica.
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