Chinese herbal medicines and nutraceuticals inhibit Pseudomonas aeruginosa biofilm formation.

Pseudomonas aeruginosa is a major biofilm-forming, opportunistic pathogen. Tolerance to antimicrobial agents due to biofilm formation may lead to the emergence of antimicrobial-resistant bacterial strains. Thus, adjunctive agents that can inhibit biofilm formation are necessary to enhance the therapeutic efficacy of antimicrobial agents. In this study, we evaluated the anti-biofilm formation activity of selected Chinese herbal medicines and nutraceuticals, which are commercially available in Japan. Among the eight agents evaluated for their potential to inhibit biofilm formation, Eiekikaryu S, Iribakuga and Hyakujunro significantly reduced P. aeruginosa biofilm formation (P <0.05) without inhibiting bacterial growth. Additionally, the expression of biofilm-associated genes (rhlR, rhlA and lasB) in P. aeruginosa was significantly suppressed by Eiekikaryu S, Iribakuga and Hyakujunro (P <0.001). Our findings indicate that some Chinese herbal medicines and nutraceuticals can be potential adjunctive agents for antimicrobial therapy against P. aeruginosa .

In vitro growth-inhibitory effects of Portulaca oleracea L. formulation on intestinal pathogens.

INTRODUCTION: Empirical evidence suggests that Portulaca oleracea L. treats enteric infections, including dysentery, cholera, and acute infectious gastroenteritis. AIM: The aim of this study is to clarify the growth-inhibitory effects of Portulaca oleracea L. extract against 56 strains of intestinal pathogens. METHODOLOGY: 'Gogyo-so-cha (GSC)' was used as the P. oleracea L. formulation. A growth curve analysis was used to measure the growth-inhibitory effects of GSC, and Shiga toxin induction was measured using the latex agglutination test. RESULTS: GSC demonstrated strong bactericidal effects against Shigella dysenteriae and Vibrio cholerae strains from various isolates. GSC demonstrated weak or no bactericidal effects against intestinal commensal bacteria, including Enterococcus spp. and Escherichia coli . GSC did not induce Shigella toxins. CONCLUSION: GSC significantly inhibited the growth of intestinal pathogens, including S. dysenteriae and V. cholerae , without adversely affecting the intestinal flora, supporting the usage of GSC in traditional Chinese medicine. Taken together, GSC would be of immense value in the developing world, where diarrhoeal infectious diseases continue to pose a major health risk.

Antimicrobial activity and additive effect of the modified Gingyo-san with antimicrobials against Helicobacter pylori.

INTRODUCTION: Helicobacter pylori is an important factor in the development of gastroduodenal ulcers and gastric cancer. Although H. pylori eradication therapy has been employed, the eradication rate has decreased in recent years owing to an increase in clarithromycin-resistant strains. We previously reported the anti-infective effect of herbal medicines against several bacterial species. Here, we evaluated the growth inhibitory activity of herbal medicines alone and in combination with antimicrobials against H. pylori. METHODS AND RESULTS: Nine of 37 herbal medicines inhibited the growth of H. pylori ATCC700392. In particular, modified Gingyo-san showed the strongest growth inhibitory activity with a minimum inhibitory concentration (MIC) of 512 µg/ml for not only ATCC700392 but also clarithromycin-resistant strains having a 23 S rRNA mutation. Results of Time-Kill Kinetics Assay showed that 1 mg/mL modified Gingyo-san treatment for one hour killed 50% of the H. pylori population. Furthermore, modified Gingyo-san showed additive effects with clarithromycin, amoxicillin, and metronidazole against H. pylori ATCC700392 and clarithromycin-resistant strains. CONCLUSIONS: Our findings showed that modified Gingyo-san inhibits the growth of H. pylori and improves antimicrobial susceptibility when used in combination. Therefore, modified Gingyo-san has the potential to enhance the eradication rate of clarithromycin-resistant H. pylori.

Kampo medicines suppress the production of exfoliative toxins causing impetigo in Staphylococcus aureus.

An alternative approach, such as antivirulence therapy that modulates the production of bacterial toxins or virulence factors, is necessary to tackle the emergence of antimicrobial-resistant strains. Here, we investigated the potential antivirulence effects of seven Kampo medicines (Jumihaidokuto, Eppikajutsuto, Jizusoippo, Shomakakkonto, Sammotsuogonto, Hainosankyuto and Inchinkoto) against exfoliative toxin (ET)-positive Staphylococcus aureus, which is the major causative agent of impetigo. A growth inhibition assay showed that all of the selected Kampo medicines inhibited the growth of S. aureus at 1/5 (2.5 mg/mL) or less of the conventionally used concentrations. Among these, Jizusoippo and Inchinkoto (0.25-1 mg/mL) suppressed the production of ET without inhibiting the bacterial growth. Furthermore, Jizusoippo and Inchinkoto significantly suppressed the expression of ET genes in a concentration-dependent manner. Our findings strongly suggest, for the first time, that Kampo medicines, especially Jizusoippo and Inchinkoto, have the potential to serve as antivirulence agents against skin infections caused by S. aureus by suppressing the production of ET.

Tokiinshi, a traditional Japanese medicine (Kampo), suppresses Panton-Valentine leukocidin production in the methicillin-resistant Staphylococcus aureus USA300 clone.

It is necessary to develop agents other than antimicrobials for the treatment of Staphylococcus aureus infections to prevent the emergence of antimicrobial-resistant strains. Particularly, anti-virulence agents against the Panton-Valentine leukocidin (PVL)-positive methicillin-resistant S. aureus (MRSA), USA300 clone, is desired due to its high pathogenicity. Here, we investigated the potential anti-virulence effect of Tokiinshi, which is a traditional Japanese medicine (Kampo) used for skin diseases, against the USA300 clone. A growth inhibition assay showed that a conventional dose (20 mg/ml) of Tokiinshi has bactericidal effects against the clinical USA300 clones. Notably, the growth inhibition effects of Tokiinshi against S. epidermidis strains, which are the major constituents of the skin microbiome, was a bacteriostatic effect. The data suggested that Tokiinshi is unlikely to affect skin flora of S. epidermidis. Furthermore, PVL production and the expression of its gene were significantly suppressed in the USA300 clone by a lower concentration (5 mg/ml) of Tokiinshi. This did not affect the number of viable bacteria. Moreover, Tokiinshi significantly suppressed the expression of the agrA gene, which regulates PVL gene expression. For the first time, our findings strongly suggest that Tokiinshi has the potential to attenuate the virulence of the USA300 clone by suppressing PVL production via agrA gene suppression.

Shiunko and Chuoko, topical Kampo medicines, inhibit the expression of gehA encoding the extracellular lipase in Cutibacterium acnes.

Antimicrobial agents have been used for eradication of Cutibacterium (formerly Propionibacterium) acnes that is an exacerbation factor of the skin disease acne vulgaris. However, the use of antibiotics is associated with an increased risk of promoting the emergence of resistant bacteria and leading to skin dysbiosis. Traditional Japanese Kampo medicines, such as Keigairengyoto, are used to treat acne. However, there is incomplete understanding regarding their functional mechanism in treatment of acne. In this study, we examined the antimicrobial and anti-lipase activity of the Kampo medicines used empirically for acne treatment. Three oral medicines, Keigairengyoto, Seijoboufuto and Jumihaidokuto, were found to inhibit the growth of C. acnes and decrease the lipase activity. Especially, Keigairengyoto caused remarkable decrease of bacterial lipase activity. Furthermore, topical medicines such as Shiunko and Chuoko significantly decreased the lipase activity in a dose-dependent manner, without inhibiting C. acnes growth. The topical medicines were found to inhibit the expression of gehA, which codes for extracellular lipase. Our results indicate that Shiunko and Chuoko have potential as effective acne therapeutic agents, especially because they do not promote the emergence of antimicrobial-resistant bacteria and skin dysbiosis.

Panax Notoginseng Extract Possesses Significant Antibacterial Activity against Pathogenic Streptococci.

BACKGROUND: In traditional Chinese medicine, Panax notoginseng is used to treat inflammation and bleeding but has not been shown to affect bacterial pathogens. OBJECTIVES: Our aim was to assess the antibacterial potential of Panax notoginseng extract (PNE) against bacterial pathogens. METHODS: PNE was dissolved in autoclaved distilled water. Antimicrobial activity was measured by the disc diffusion test and bacterial growth curve assays, in which the concentration of bacterial colony forming units was monitored at several time points in the presence or absence of PNE. RESULTS: Disc diffusion and growth curve assays demonstrated that PNE significantly inhibited the growth of Streptococcus pyogenes, Streptococcus pneumoniae, and Haemophilus influenzae (p < 0.05). In contrast, the growth of the oral commensal bacteria Streptococcus intermedius, Streptococcus salivarius, and Streptococcus anginosus was not inhibited. Therefore, S. pyogenes clinical isolates were analyzed. PNE had antimicrobial effects on all tested isolates in both aerobic and anaerobic conditions. In addition, when S. pyogenes was co-cultured with S. intermedius in the presence of PNE, PNE inhibited the growth of S. pyogenes, but did not inhibit the growth of S. intermedius. CONCLUSION: Our findings suggested that PNE inhibited S. pyogenes without affecting oral commensal bacteria. Therefore, PNE could be used for the treatment of S. pyogenes infections.

Oldenlandia diffusa Extract Inhibits Biofilm Formation by Haemophilus influenzae Clinical Isolates.

Oldenlandia diffusa has been empirically used as a therapeutic adjunct for the treatment of respiratory infections. To establish the basic evidence of its clinical usefulness, antimicrobial and biofilm inhibitory activities of an O. diffusa extract were examined against clinical isolates of Haemophilus influenzae, a major causative pathogen of respiratory and sensory organ infections. No significant growth inhibitory activity was observed during incubation for more than 6 h after the extract addition into a culture of H. influenzae. On the other hand, biofilm formation by H. influenzae, evaluated by a crystal violet method, was significantly and dose-dependently inhibited by the O. diffusa extract. Furthermore, the mRNA level of the biofilm-associated gene luxS of H. influenzae significantly decreased soon after the extract addition, and the suppressive effect continued for at least 2 h. At 2 h after the addition of the O. diffusa extract, the autoinducer in the culture supernatant was also significantly reduced by the O. diffusa extract in a dose-dependent manner. These results revealed that O. diffusa extract shows inhibitory activity against luxS-dependent biofilm formation but has no antimicrobial activity against planktonic cells of H. influenzae. Thus, O. diffusa extract might be useful as an adjunctive therapy for the treatment of respiratory infections caused by H. influenzae.

The modified Gingyo-san, a Chinese herbal medicine, has direct antibacterial effects against respiratory pathogens.

BACKGROUND: Modified Gingyo-san (MGS) is empirically used to treat various respiratory infections. MGS has been reported to have antiinflammatory and antiviral activities; however, it is not known if it has an antibacterial activity. Therefore, in this study, we aimed to investigate the antimicrobial activity of MGS against respiratory pathogens. METHODS: MGS, which is sold as an over-the-counter drug in Japan, was used for the study. Antimicrobial activity was evaluated using the disk diffusion method. Growth inhibitory activity was evaluated by measuring colony-forming units of the pathogens in the presence of MGS. RESULTS: MGS inhibited the growth of Bacillus subtilis, Streptococcus pneumoniae, and Streptococcus pyogenes, which are gram-positive bacteria. Although the growth of most gram-negative bacteria was not inhibited by MGS, interestingly, the growth of Haemophilus influenzae was inhibited. MGS did not show any activity against Candida albicans or bacteriophage φX174. CONCLUSIONS: In addition to the antiinflammatory and antiviral activities of MGS, which have already been reported, the data obtained from this study indicates that MGS has an antibacterial activity.

Effects of a 5-HT7 receptor antagonist DR4004 on the exploratory behavior in a novel environment and on brain monoamine dynamics in mice.

The present study examined whether serotonin (5-hydroxytryptamine; 5-HT)7 receptors play a role in the modulation of emotionality in mice using the selective 5-HT7 receptor antagonist 2a-[4-(4-phenyl-1,2,3,6-tetrahydropyridyl)butyl]-2a,3,4,5-tetrahydrobenzo (c,d)indol-2-(1H)-one (DR4004). The emotionality of mice was evaluated in terms of exploratory activity in the hole-board test. The mice treated with DR4004 (2.5-10 mg/kg, i.p.) displayed a dose-dependent decrease in locomotor activity by moving less distance in the hole-board, and statistically significant decreases were observed at 5 and 10 mg/kg. On the other hand, DR4004 (10 mg/kg, i.p.) did not affect spontaneous motor activity. In a neurochemical study, decreases in amygdaloid dopamine and 5-HT turnover were observed in mice in which locomotor activity in the hole-board test was attenuated following the administration of DR4004 (10 mg/kg, i.p.). Also, a simple linear regression analysis revealed that locomotor activity on the hole-board was significantly correlated with dopamine and 5-HT turnover in amygdala. Furthermore, co-injection of the selective dopamine reuptake inhibitor 1-(2-[bis(4-fluorophenyl)methoxy]ethyl)-4-(3-phenylpropyl)piperazine (GBR12909; 1.25-5 mg/kg, i.p.) or the selective 5-HT reuptake inhibitor fluvoxamine (20 mg/kg, i.p.) significantly reversed the DR4004 (10 mg/kg, i.p.)-induced decrease in locomotor activity in the hole-board test. These findings constitute the behavioral evidence that 5-HT7 receptors may play a role in the modulation of emotionality. Furthermore, it is also suggested that amygdaloid dopamine and 5-HT neuronal systems may be involved in this modulation.